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Sample records for adjacent cell types

  1. Sound insulation property of membrane-type acoustic metamaterials carrying different masses at adjacent cells

    NASA Astrophysics Data System (ADS)

    Zhang, Yuguang; Wen, Jihong; Zhao, Honggang; Yu, Dianlong; Cai, Li; Wen, Xisen

    2013-08-01

    We present the experimental realization and theoretical understanding of membrane-type acoustic metamaterials embedded with different masses at adjacent cells, capable of increasing the transmission loss at low frequency. Owing to the reverse vibration of adjacent cells, Transmission loss (TL) peaks appear, and the magnitudes of the TL peaks exceed the predicted results of the composite wall. Compared with commonly used configuration, i.e., all cells carrying with identical mass, the nonuniformity of attaching masses causes another much low TL peak. Finite element analysis was employed to validate and provide insights into the TL behavior of the structure.

  2. Type-1 Collagen differentially alters β-catenin accumulation in primary Dupuytren's Disease cord and adjacent palmar fascia cells

    PubMed Central

    Vi, Linda; Njarlangattil, Anna; Wu, Yan; Gan, Bing Siang; O'Gorman, David B

    2009-01-01

    Background Dupuytren's Disease (DD) is a debilitating contractile fibrosis of the palmar fascia characterised by excess collagen deposition, contractile myofibroblast development, increased Transforming Growth Factor-β levels and β-catenin accumulation. The aim of this study was to determine if a collagen-enriched environment, similar to in vivo conditions, altered β-catenin accumulation by primary DD cells in the presence or absence of Transforming Growth Factor-β. Methods Primary DD and patient matched, phenotypically normal palmar fascia (PF) cells were cultured in the presence or absence of type-1 collagen and Transforming Growth Factor-β1. β-catenin and α-smooth muscle actin levels were assessed by western immunoblotting and immunofluorescence microscopy. Results DD cells display a rapid depletion of cellular β-catenin not evident in patient-matched PF cells. This effect was not evident in either cell type when cultured in the absence of type-1 collagen. Exogenous addition of Transforming Growth Factor-β1 to DD cells in collagen culture negates the loss of β-catenin accumulation. Transforming Growth Factor-β1-induced α-smooth muscle actin, a marker of myofibroblast differentiation, is attenuated by the inclusion of type-1 collagen in cultures of DD and PF cells. Conclusion Our findings implicate type-1 collagen as a previously unrecognized regulator of β-catenin accumulation and a modifier of TGF-β1 signaling specifically in primary DD cells. These data have implications for current treatment modalities as well as the design of in vitro models for research into the molecular mechanisms of DD. PMID:19545383

  3. Interior building details of Building A, dungeon cell adjacent to ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Interior building details of Building A, dungeon cell adjacent to northwest cell: granite and brick threshold, poured concrete floors, plastered finished walls, vaulted veiling; northwesterly view - San Quentin State Prison, Building 22, Point San Quentin, San Quentin, Marin County, CA

  4. Human breast cancer biopsies induce eosinophil recruitment and enhance adjacent cancer cell proliferation.

    PubMed

    Szalayova, Gabriela; Ogrodnik, Aleksandra; Spencer, Brianna; Wade, Jacqueline; Bunn, Janice; Ambaye, Abiy; James, Ted; Rincon, Mercedes

    2016-06-01

    Chronic inflammation is known to facilitate cancer progression and metastasis. Less is known about the effect of acute inflammation within the tumor microenvironment, resulting from standard invasive procedures. Recent studies in mouse models have shown that the acute inflammatory response triggered by a biopsy in mammary cancer increases the frequency of distal metastases. Although tumor biopsies are part of the standard clinical practice in breast cancer diagnosis, no studies have reported their effect on inflammatory response. The objective of this study is to (1) determine whether core needle biopsies in breast cancer patients trigger an inflammatory response, (2) characterize the type of inflammatory response present, and (3) evaluate the potential effect of any acute inflammatory response on residual tumor cells. The biopsy wound site was identified in the primary tumor resection tissue samples from breast cancer patients. The inflammatory response in areas adjacent (i.e., immediately around previous biopsy site) and distant to the wound biopsy was investigated by histology and immunohistochemistry analysis. Proliferation of tumor cells was also assayed. We demonstrate that diagnostic core needle biopsies trigger a selective recruitment of inflammatory cells at the site of the biopsy, and they persist for extended periods of time. While macrophages were part of the inflammatory response, an unexpected accumulation of eosinophils at the edge of the biopsy wound was also identified. Importantly, we show that biopsy causes an increase in the proliferation rate of tumor cells located in the area adjacent to the biopsy wound. Diagnostic core needle biopsies in breast cancer patients do induce a unique acute inflammatory response within the tumor microenvironment and have an effect on the surrounding tumor cells. Therefore, biopsy-induced inflammation could have an impact on residual tumor cell progression and/or metastasis in human breast cancer. These findings

  5. Proteome analysis of proliferative response of bystander cells adjacent to cells exposed to ionizing radiation

    PubMed Central

    Gerashchenko, Bogdan I.; Yamagata, Akira; Oofusa, Ken; Yoshizato, Katsutoshi; de Toledo, Sonia M.; Howell, Roger W.

    2010-01-01

    Recently (Cytometry 2003, 56A, 71–80), we reported that direct cell-to-cell contact is required for stimulating proliferation of bystander rat liver cells (WB-F344) cocultured with irradiated cells, and neither functional gap junction intercellular communication nor long-range extracellular factors appear to be involved in this proliferative bystander response (PBR). The molecular basis for this response is unknown. Confluent monolayers of WB-F344 cells were exposed to 5-Gray (Gy) of γ-rays. Irradiated cells were mixed with unirradiated cells and co-cultured for 24 h. Cells were harvested and protein expression was examined using 2-DE. Protein expression was also determined in cultures of unirradiated and 5-Gy irradiated cells. Proteins were identified by MS. Nucleophosmin (NPM)-1, a multifunctional nucleolar protein, was more highly expressed in bystander cells than in either unirradiated or 5-Gy irradiated cells. Enolase-α, a glycolytic enzyme, was present in acidic and basic variants in unirradiated cells. In bystander and 5-Gy irradiated cells, the basic variant was weakly expressed, whereas the acidic variant was overwhelmingly present. These data indicate that the presence of irradiated cells can affect NPM-1 and enolase-α in adjacent bystander cells. These proteins appear to participate in molecular events related to the PBR and suggest that this response may involve cellular defense, proliferation, and metabolism. PMID:17514680

  6. Proteome analysis of proliferative response of bystander cells adjacent to cells exposed to ionizing radiation.

    PubMed

    Gerashchenko, Bogdan I; Yamagata, Akira; Oofusa, Ken; Yoshizato, Katsutoshi; de Toledo, Sonia M; Howell, Roger W

    2007-06-01

    Recently (Cytometry 2003, 56A, 71-80), we reported that direct cell-to-cell contact is required for stimulating proliferation of bystander rat liver cells (WB-F344) cocultured with irradiated cells, and neither functional gap junction intercellular communication nor long-range extracellular factors appear to be involved in this proliferative bystander response (PBR). The molecular basis for this response is unknown. Confluent monolayers of WB-F344 cells were exposed to 5-Gray (Gy) of gamma-rays. Irradiated cells were mixed with unirradiated cells and co-cultured for 24 h. Cells were harvested and protein expression was examined using 2-DE. Protein expression was also determined in cultures of unirradiated and 5-Gy irradiated cells. Proteins were identified by MS. Nucleophosmin (NPM)-1, a multifunctional nucleolar protein, was more highly expressed in bystander cells than in either unirradiated or 5-Gy irradiated cells. Enolase-alpha, a glycolytic enzyme, was present in acidic and basic variants in unirradiated cells. In bystander and 5-Gy irradiated cells, the basic variant was weakly expressed, whereas the acidic variant was overwhelmingly present. These data indicate that the presence of irradiated cells can affect NPM-1 and enolase-alpha in adjacent bystander cells. These proteins appear to participate in molecular events related to the PBR and suggest that this response may involve cellular defense, proliferation, and metabolism.

  7. Type- and Subcomplex-Specific Neutralizing Antibodies against Domain III of Dengue Virus Type 2 Envelope Protein Recognize Adjacent Epitopes▿

    PubMed Central

    Sukupolvi-Petty, Soila; Austin, S. Kyle; Purtha, Whitney E.; Oliphant, Theodore; Nybakken, Grant E.; Schlesinger, Jacob J.; Roehrig, John T.; Gromowski, Gregory D.; Barrett, Alan D.; Fremont, Daved H.; Diamond, Michael S.

    2007-01-01

    Neutralization of flaviviruses in vivo correlates with the development of an antibody response against the viral envelope (E) protein. Previous studies demonstrated that monoclonal antibodies (MAbs) against an epitope on the lateral ridge of domain III (DIII) of the West Nile virus (WNV) E protein strongly protect against infection in animals. Based on X-ray crystallography and sequence analysis, an analogous type-specific neutralizing epitope for individual serotypes of the related flavivirus dengue virus (DENV) was hypothesized. Using yeast surface display of DIII variants, we defined contact residues of a panel of type-specific, subcomplex-specific, and cross-reactive MAbs that recognize DIII of DENV type 2 (DENV-2) and have different neutralizing potentials. Type-specific MAbs with neutralizing activity against DENV-2 localized to a sequence-unique epitope on the lateral ridge of DIII, centered at the FG loop near residues E383 and P384, analogous in position to that observed with WNV-specific strongly neutralizing MAbs. Subcomplex-specific MAbs that bound some but not all DENV serotypes and neutralized DENV-2 infection recognized an adjacent epitope centered on the connecting A strand of DIII at residues K305, K307, and K310. In contrast, several MAbs that had poor neutralizing activity against DENV-2 and cross-reacted with all DENV serotypes and other flaviviruses recognized an epitope with residues in the AB loop of DIII, a conserved region that is predicted to have limited accessibility on the mature virion. Overall, our experiments define adjacent and structurally distinct epitopes on DIII of DENV-2 which elicit type-specific, subcomplex-specific, and cross-reactive antibodies with different neutralizing potentials. PMID:17881453

  8. Type- and subcomplex-specific neutralizing antibodies against domain III of dengue virus type 2 envelope protein recognize adjacent epitopes.

    PubMed

    Sukupolvi-Petty, Soila; Austin, S Kyle; Purtha, Whitney E; Oliphant, Theodore; Nybakken, Grant E; Schlesinger, Jacob J; Roehrig, John T; Gromowski, Gregory D; Barrett, Alan D; Fremont, Daved H; Diamond, Michael S

    2007-12-01

    Neutralization of flaviviruses in vivo correlates with the development of an antibody response against the viral envelope (E) protein. Previous studies demonstrated that monoclonal antibodies (MAbs) against an epitope on the lateral ridge of domain III (DIII) of the West Nile virus (WNV) E protein strongly protect against infection in animals. Based on X-ray crystallography and sequence analysis, an analogous type-specific neutralizing epitope for individual serotypes of the related flavivirus dengue virus (DENV) was hypothesized. Using yeast surface display of DIII variants, we defined contact residues of a panel of type-specific, subcomplex-specific, and cross-reactive MAbs that recognize DIII of DENV type 2 (DENV-2) and have different neutralizing potentials. Type-specific MAbs with neutralizing activity against DENV-2 localized to a sequence-unique epitope on the lateral ridge of DIII, centered at the FG loop near residues E383 and P384, analogous in position to that observed with WNV-specific strongly neutralizing MAbs. Subcomplex-specific MAbs that bound some but not all DENV serotypes and neutralized DENV-2 infection recognized an adjacent epitope centered on the connecting A strand of DIII at residues K305, K307, and K310. In contrast, several MAbs that had poor neutralizing activity against DENV-2 and cross-reacted with all DENV serotypes and other flaviviruses recognized an epitope with residues in the AB loop of DIII, a conserved region that is predicted to have limited accessibility on the mature virion. Overall, our experiments define adjacent and structurally distinct epitopes on DIII of DENV-2 which elicit type-specific, subcomplex-specific, and cross-reactive antibodies with different neutralizing potentials.

  9. Correlative light and electron microscopy of the frog adrenal gland cells using adjacent epon-embedded sections.

    PubMed

    Nakai, Y; Iwashita, T

    1976-07-01

    Correlative light and electron microscopy on the same cells of the adrenal gland of the frog, Rana nigromaculata, fixed in glutaraldehyde followed by osmium tetroxide, was done using the adjacent Epon embedded sections. Electron microscope observation revealed three different types of granule-filled secretory cells; the noradrenaline-storing cells (NA cells) filled with intensely dense and varying shaped granules, the adrenaline-strong cells (A cells) filled with relatively less dense granules and the summer cells (STILLING, 1898) containing very large, round or polygonal granules (0.2-1.3 mu in diameter). Light microscopically, an essential difference could be observed in the affinity to ammoniacal silver solution between NA and A cells. It was clarified that the granules of NA cells stained in black and were clearly distinguishable from the yellow- or brown-stained granules in both A cells and summer cells. This silver method can be applied for the light microscopic identification of the NA cells in the Epon-embedded sections. Furthermore, after immersing the thick sections in toluidine blue or methylene blue, the granules of NA cells showed much stronger affinity to both dyes than those of A cells and became dark blue and occasionally stained greenish blue in methylene blue, while the summer cells became blue and the granules of the A cells stained light blue.

  10. Characterization of humus microbial communities in adjacent forest types that differ in nitrogen availability.

    PubMed

    Leckie, S E; Prescott, C E; Grayston, S J; Neufeld, J D; Mohn, W W

    2004-07-01

    To address the link between soil microbial community composition and soil processes, we investigated the microbial communities in forest floors of two forest types that differ substantially in nitrogen availability. Cedar-hemlock (CH) and hemlock-amabilis fir (HA) forests are both common on northern Vancouver Island, B.C., occurring adjacently across the landscape. CH forest floors have low nitrogen availability and HA high nitrogen availability. Total microbial biomass was assessed using chloroform fumigation-extraction and community composition was assessed using several cultivation-independent approaches: denaturing gradient gel electrophoresis (DGGE) of the bacterial communities, ribosomal intergenic spacer analysis (RISA) of the bacterial and fungal communities, and phospholipid fatty acid (PLFA) profiles of the whole microbial community. We did not detect differences in the bacterial communities of each forest type using DGGE and RISA, but differences in the fungal communities were detected using RISA. PLFA analysis detected subtle differences in overall composition of the microbial community between the forest types, as well as in particular groups of organisms. Fungal PLFAs were more abundant in the nitrogen-poor CH forests. Bacteria were proportionally more abundant in HA forests than CH in the lower humus layer, and Gram-positive bacteria were proportionally more abundant in HA forests irrespective of layer. Bacterial and fungal communities were distinct in the F, upper humus, and lower humus layers of the forest floor and total biomass decreased in deeper layers. These results indicate that there are distinct patterns in forest floor microbial community composition at the landscape scale, which may be important for understanding nutrient availability to forest vegetation.

  11. Molecular heterogeneity in adjacent cells in triple-negative breast cancer

    PubMed Central

    Huebschman, Michael L; Lane, Nancy L; Liu, Huaying; Sarode, Venetia R; Devlin, Judith L; Frenkel, Eugene P

    2015-01-01

    Purpose This study interrogates the molecular status of individual cells in patients with triple-negative breast cancers and explores the molecular identification and characterization of these tumors to consider the exploitation of a potential-targeted therapeutic approach. Patients and methods Hyperspectral immunologic cell by cell analysis was applied to touch imprint smears obtained from fresh tumors of breast cancer patients. Results Cell by cell analysis confirms significant intratumoral molecular heterogeneity in cancer markers with differences from polymerase chain reaction marker reporting. The individual cell heterogeneity was recognized in adjacent cells examined with panels of ten molecular markers in each single cell and included some markers that are considered to express “stem-cell” character. In addition, heterogeneity did not relate either to the size or stage of the primary tumor or to the site from within the cancer. Conclusion There is a very significant molecular heterogeneity when “adjacent cells” are examined in triple-negative breast cancer, thereby making a successful targeted approach unlikely. In addition, it is not reasonable to consider that these changes will provide an answer to tumor dormancy. PMID:26316815

  12. Preferential localization of IgG memory B cells adjacent to contracted germinal centers

    PubMed Central

    Aiba, Yuichi; Kometani, Kohei; Hamadate, Megumi; Moriyama, Saya; Sakaue-Sawano, Asako; Tomura, Michio; Luche, Hervé; Fehling, Hans Jörg; Casellas, Rafael; Kanagawa, Osami; Miyawaki, Atsushi; Kurosaki, Tomohiro

    2010-01-01

    It has long been presumed that after leaving the germinal centers (GCs), memory B cells colonize the marginal zone or join the recirculating pool. Here we demonstrate the preferential localization of nitrophenol-chicken γ-globulin-induced CD38+IgG1+ memory B cells adjacent to contracted GCs in the spleen. The memory B cells in this region proliferated after secondary immunization, a response that was abolished by depletion of CD4+ T cells. We also found that these IgG1+ memory B cells could present antigen on their surface, and that this activity was required for their activation. These results implicate this peri-GC region as an important site for survival and reactivation of memory B cells. PMID:20547847

  13. Star cell type core configuration for structural sandwich materials

    DOEpatents

    Christensen, Richard M.

    1995-01-01

    A new pattern for cellular core material used in sandwich type structural materials. The new pattern involves star shaped cells intermixed with hexagonal shaped cells. The new patterned cellular core material includes star shaped cells interconnected at points thereof and having hexagonal shape cells positioned adjacent the star points. The new pattern allows more flexibility and can conform more easily to curved shapes.

  14. Iterative Convergence Acceleration of Neutral Particle Transport Methods via Adjacent-Cell Preconditioners

    NASA Astrophysics Data System (ADS)

    Azmy, Y. Y.

    1999-06-01

    We propose preconditioning as a viable acceleration scheme for the inner iterations of transport calculations in slab geometry. In particular we develop Adjacent-Cell Preconditioners (AP) that have the same coupling stencil as cell-centered diffusion schemes. For lowest order methods, e.g., Diamond Difference, Step, and 0-order Nodal Integral Method (0NIM), cast in a Weighted Diamond Difference (WDD) form, we derive AP for thick (KAP) and thin (NAP) cells that for model problems are unconditionally stable and efficient. For the First-Order Nodal Integral Method (1NIM) we derive a NAP that possesses similarly excellent spectral properties for model problems. [Note that the order of NIM refers to the truncated order of the local expansion of the cell and edge fluxes in Legendre series.] The two most attractive features of our new technique are: (1) its cell-centered coupling stencil, which makes it more adequate for extension to multidimensional, higher order situations than the standard edge-centered or point-centered Diffusion Synthetic Acceleration (DSA) methods; and (2) its decreasing spectral radius with increasing cell thickness to the extent that immediate pointwise convergence, i.e., in one iteration, can be achieved for problems with sufficiently thick cells. We implemented these methods, augmented with appropriate boundary conditions and mixing formulas for material heterogeneities, in the test code AP1D that we use to successfully verify the analytical spectral properties for homogeneous problems. Furthermore, we conduct numerical tests to demonstrate the robustness of the KAP and NAP in the presence of sharp mesh or material discontinuities. We show that the AP for WDD is highly resilient to such discontinuities, but for 1NIM a few cases occur in which the scheme does not converge; however, when it converges, AP greatly reduces the number of iterations required to achieve convergence.

  15. Iterative convergence acceleration of neutral particle transport methods via adjacent-cell preconditioners

    SciTech Connect

    Azmy, Y.Y.

    1999-06-10

    The author proposes preconditioning as a viable acceleration scheme for the inner iterations of transport calculations in slab geometry. In particular he develops Adjacent-Cell Preconditioners (AP) that have the same coupling stencil as cell-centered diffusion schemes. For lowest order methods, e.g., Diamond Difference, Step, and 0-order Nodal Integral Method (ONIM), cast in a Weighted Diamond Difference (WDD) form, he derives AP for thick (KAP) and thin (NAP) cells that for model problems are unconditionally stable and efficient. For the First-Order Nodal Integral Method (INIM) he derives a NAP that possesses similarly excellent spectral properties for model problems. The two most attractive features of the new technique are:(1) its cell-centered coupling stencil, which makes it more adequate for extension to multidimensional, higher order situations than the standard edge-centered or point-centered Diffusion Synthetic Acceleration (DSA) methods; and (2) its decreasing spectral radius with increasing cell thickness to the extent that immediate pointwise convergence, i.e., in one iteration, can be achieved for problems with sufficiently thick cells. He implemented these methods, augmented with appropriate boundary conditions and mixing formulas for material heterogeneities, in the test code APID that he uses to successfully verify the analytical spectral properties for homogeneous problems. Furthermore, he conducts numerical tests to demonstrate the robustness of the KAP and NAP in the presence of sharp mesh or material discontinuities. He shows that the AP for WDD is highly resilient to such discontinuities, but for INIM a few cases occur in which the scheme does not converge; however, when it converges, AP greatly reduces the number of iterations required to achieve convergence.

  16. Types of Stem Cells

    MedlinePlus

    ... PDF) Download an introduction to stem cells and stem cell research. Stem Cell Glossary Stem cell terms to know. ... stem cells blog from the International Society for Stem Cell Research. Learn About Stem Cells From Lab to You ...

  17. Spontaneous Activity of Cochlear Hair Cells Triggered by Fluid Secretion Mechanism in Adjacent Support Cells.

    PubMed

    Wang, Han Chin; Lin, Chun-Chieh; Cheung, Rocky; Zhang-Hooks, YingXin; Agarwal, Amit; Ellis-Davies, Graham; Rock, Jason; Bergles, Dwight E

    2015-12-01

    Spontaneous electrical activity of neurons in developing sensory systems promotes their maturation and proper connectivity. In the auditory system, spontaneous activity of cochlear inner hair cells (IHCs) is initiated by the release of ATP from glia-like inner supporting cells (ISCs), facilitating maturation of central pathways before hearing onset. Here, we find that ATP stimulates purinergic autoreceptors in ISCs, triggering Cl(-) efflux and osmotic cell shrinkage by opening TMEM16A Ca(2+)-activated Cl(-) channels. Release of Cl(-) from ISCs also forces K(+) efflux, causing transient depolarization of IHCs near ATP release sites. Genetic deletion of TMEM16A markedly reduces the spontaneous activity of IHCs and spiral ganglion neurons in the developing cochlea and prevents ATP-dependent shrinkage of supporting cells. These results indicate that supporting cells in the developing cochlea have adapted a pathway used for fluid secretion in other organs to induce periodic excitation of hair cells. PMID:26627734

  18. Schottky barrier amorphous silicon solar cell with thin doped region adjacent metal Schottky barrier

    DOEpatents

    Carlson, David E.; Wronski, Christopher R.

    1979-01-01

    A Schottky barrier amorphous silicon solar cell incorporating a thin highly doped p-type region of hydrogenated amorphous silicon disposed between a Schottky barrier high work function metal and the intrinsic region of hydrogenated amorphous silicon wherein said high work function metal and said thin highly doped p-type region forms a surface barrier junction with the intrinsic amorphous silicon layer. The thickness and concentration of p-type dopants in said p-type region are selected so that said p-type region is fully ionized by the Schottky barrier high work function metal. The thin highly doped p-type region has been found to increase the open circuit voltage and current of the photovoltaic device.

  19. Star cell type core configuration for structural sandwich materials

    DOEpatents

    Christensen, R.M.

    1995-08-01

    A new pattern for cellular core material used in sandwich type structural materials is disclosed. The new pattern involves star shaped cells intermixed with hexagonal shaped cells. The new patterned cellular core material includes star shaped cells interconnected at points thereof and having hexagonal shape cells positioned adjacent the star points. The new pattern allows more flexibility and can conform more easily to curved shapes. 3 figs.

  20. Periostin differentially induces proliferation, contraction and apoptosis of primary Dupuytren's disease and adjacent palmar fascia cells

    SciTech Connect

    Vi, Linda; Feng, Lucy; Zhu, Rebecca D.; Wu, Yan; Satish, Latha; Gan, Bing Siang; O'Gorman, David B.

    2009-12-10

    Dupuytren's disease, (DD), is a fibroproliferative condition of the palmar fascia in the hand, typically resulting in permanent contracture of one or more fingers. This fibromatosis is similar to scarring and other fibroses in displaying excess collagen secretion and contractile myofibroblast differentiation. In this report we expand on previous data demonstrating that POSTN mRNA, which encodes the extra-cellular matrix protein periostin, is up-regulated in Dupuytren's disease cord tissue relative to phenotypically normal palmar fascia. We demonstrate that the protein product of POSTN, periostin, is abundant in Dupuytren's disease cord tissue while little or no periostin immunoreactivity is evident in patient-matched control tissues. The relevance of periostin up-regulation in DD was assessed in primary cultures of cells derived from diseased and phenotypically unaffected palmar fascia from the same patients. These cells were grown in type-1 collagen-enriched culture conditions with or without periostin addition to more closely replicate the in vivo environment. Periostin was found to differentially regulate the apoptosis, proliferation, {alpha} smooth muscle actin expression and stressed Fibroblast Populated Collagen Lattice contraction of these cell types. We hypothesize that periostin, secreted by disease cord myofibroblasts into the extra-cellular matrix, promotes the transition of resident fibroblasts in the palmar fascia toward a myofibroblast phenotype, thereby promoting disease progression.

  1. Dynamics of re-constitution of the human nuclear proteome after cell division is regulated by NLS-adjacent phosphorylation

    PubMed Central

    Róna, Gergely; Borsos, Máté; Ellis, Jonathan J; Mehdi, Ahmed M; Christie, Mary; Környei, Zsuzsanna; Neubrandt, Máté; Tóth, Judit; Bozóky, Zoltán; Buday, László; Madarász, Emília; Bodén, Mikael; Kobe, Bostjan; Vértessy, Beáta G

    2014-01-01

    Phosphorylation by the cyclin-dependent kinase 1 (Cdk1) adjacent to nuclear localization signals (NLSs) is an important mechanism of regulation of nucleocytoplasmic transport. However, no systematic survey has yet been performed in human cells to analyze this regulatory process, and the corresponding cell-cycle dynamics have not yet been investigated. Here, we focused on the human proteome and found that numerous proteins, previously not identified in this context, are associated with Cdk1-dependent phosphorylation sites adjacent to their NLSs. Interestingly, these proteins are involved in key regulatory events of DNA repair, epigenetics, or RNA editing and splicing. This finding indicates that cell-cycle dependent events of genome editing and gene expression profiling may be controlled by nucleocytoplasmic trafficking. For in-depth investigations, we selected a number of these proteins and analyzed how point mutations, expected to modify the phosphorylation ability of the NLS segments, perturb nucleocytoplasmic localization. In each case, we found that mutations mimicking hyper-phosphorylation abolish nuclear import processes. To understand the mechanism underlying these phenomena, we performed a video microscopy-based kinetic analysis to obtain information on cell-cycle dynamics on a model protein, dUTPase. We show that the NLS-adjacent phosphorylation by Cdk1 of human dUTPase, an enzyme essential for genomic integrity, results in dynamic cell cycle-dependent distribution of the protein. Non-phosphorylatable mutants have drastically altered protein re-import characteristics into the nucleus during the G1 phase. Our results suggest a dynamic Cdk1-driven mechanism of regulation of the nuclear proteome composition during the cell cycle. PMID:25483092

  2. Self-Association Is Required for Occupation of Adjacent Binding Sites in Pseudomonas aeruginosa Type III Secretion System Promoters

    PubMed Central

    Marsden, Anne E.; Schubot, Florian D.

    2014-01-01

    ExsA is a member of the AraC/XylS family of transcriptional regulators and is required for expression of the Pseudomonas aeruginosa type III secretion system (T3SS). All P. aeruginosa T3SS promoters contain two adjacent binding sites for monomeric ExsA. The amino-terminal domain of ExsA (NTD) is thought to mediate interactions between the ExsA monomers bound to each site. Threading the NTD onto the AraC backbone revealed an α-helix that likely serves as the primary determinant for dimerization. In this study, we performed alanine scanning mutagenesis of the ExsA α-helix (residues 136 to 152) to identify determinants required for self-association. Residues L137, C139, L140, K141, and L148 exhibited self-association defects and were required for maximal activation by ExsA. Disruption of self-association resulted in decreased binding to T3SS promoters, particularly loss of binding by the second ExsA monomer. Removing the NTD or increasing the space between the ExsA-binding sites restored the ability of the second ExsA monomer to bind the PexsC promoter. This finding indicated that, in the absence of self-association, the NTD prevents binding by a second monomer. Similar findings were seen with the PexoT promoter; however, binding of the second ExsA monomer in the absence of self-association also required the presence of a high-affinity site 2. Based on these data, ExsA self-association is necessary to overcome inhibition by the NTD and to compensate for low-affinity binding sites, thereby allowing for full occupation and activation of ExsA-dependent promoters. Therefore, ExsA self-association is indispensable and provides an attractive target for antivirulence therapies. PMID:25070741

  3. [Species specificity of morphogenetic factors of Acetabularia, localized in the cytoplasmic zone adjacent to the cell membrane].

    PubMed

    Naumova, G A; Naumova, L P; Puchkova, L I; Savchenko, S M; Sandakhchiev, L S

    1976-01-01

    The species specificity of the factors controlling the cap development was established in the experiments with the transplantation of both the intact and centrifuged in the basal direction apical regions of Acetabularia meditteranea on nuclear basal regions of A. crenulata. These factors are found at the stage of 72 hrs of regeneration primarily in the cytoplasmic zone adjacent to the cell membrane which is not displaced during centrifugation. Using direct measurements and radiochemical method, we have shown that the accumulation of proteins proceeded in this zone due, mainly, to their transition from the cytoplasmic zone displaced during centrifugation.

  4. Distinguishing cell type using epigenotype

    NASA Astrophysics Data System (ADS)

    Wytock, Thomas; Motter, Adilson E.

    Recently, researchers have proposed that unique cell types are attractors of their epigenetic dynamics including gene expression and chromatin conformation patterns. Traditionally, cell types have been classified by their function, morphology, cytochemistry, and other macroscopically observable properties. Because these properties are the result of many proteins working together, it should be possible to predict cell types from gene expression or chromatin conformation profiles. In this talk, I present a maximum entropy approach to identify and distinguish cell type attractors on the basis of correlations within these profiles. I will demonstrate the flexibility of this method through its separate application to gene expression and chromatin conformation datasets. I show that our method out-performs other machine-learning techniques and uncorrelated benchmarks. We adapt our method to predict growth rate from gene expression in E. coli and S. cerevisiae and compare our predictions with those from metabolic models. In addition, our method identifies a nearly convex region of state-space associated with each cell type attractor basin. Estimates of the growth rate and attractor basin make it possible to rationally control gene regulatory networks independent of a model. This research was supported by NSF-GRFP, NSF-GK12, GAANN, and Northwestern's NIH-NIGMS Molecular Biophysics Training Grant.

  5. Identification of a New Stromal Cell Type Involved in the Regulation of Inflamed B Cell Follicles

    PubMed Central

    Mionnet, Cyrille; Mondor, Isabelle; Jorquera, Audrey; Loosveld, Marie; Maurizio, Julien; Arcangeli, Marie-Laure; Ruddle, Nancy H.; Nowak, Jonathan; Aurrand-Lions, Michel; Luche, Hervé; Bajénoff, Marc

    2013-01-01

    Lymph node (LN) stromal cells provide survival signals and adhesive substrata to lymphocytes. During an immune response, B cell follicles enlarge, questioning how LN stromal cells manage these cellular demands. Herein, we used a murine fate mapping system to describe a new stromal cell type that resides in the T cell zone of resting LNs. We demonstrated that upon inflammation, B cell follicles progressively trespassed into the adjacent T cell zone and surrounded and converted these stromal cells into CXCL13 secreting cells that in return delineated the new boundaries of the growing follicle. Acute B cell ablation in inflamed LNs abolished CXCL13 secretion in these cells, while LT-β deficiency in B cells drastically affected this conversion. Altogether, we reveal the existence of a dormant stromal cell subset that can be functionally awakened by B cells to delineate the transient boundaries of their expanding territories upon inflammation. PMID:24130458

  6. Galvanic cell type oxygen sensor

    SciTech Connect

    Fujita, Y.; Kudo, H.; Tanigawa, I.

    1985-01-22

    A galvanic cell type oxygen sensor comprising a galvanic cell comprised of a cathode made up of metal effective for the electrolytic reduction of oxygen, an anode made up of lead material and an electrolyte made up of an aqueous mixed solution of organic acid and organic acid salt, which has a long life and a high output voltage, is not at all affected by carbon dioxide and which can prevent the generation of hydrogen from the cathode, is disclosed.

  7. The vascular plant-pathogenic bacterium Ralstonia solanacearum produces biofilms required for its virulence on the surfaces of tomato cells adjacent to intercellular spaces.

    PubMed

    Mori, Yuka; Inoue, Kanako; Ikeda, Kenichi; Nakayashiki, Hitoshi; Higashimoto, Chikaki; Ohnishi, Kouhei; Kiba, Akinori; Hikichi, Yasufumi

    2016-08-01

    The mechanism of colonization of intercellular spaces by the soil-borne and vascular plant-pathogenic bacterium Ralstonia solanacearum strain OE1-1 after invasion into host plants remains unclear. To analyse the behaviour of OE1-1 cells in intercellular spaces, tomato leaves with the lower epidermis layers excised after infiltration with OE1-1 were observed under a scanning electron microscope. OE1-1 cells formed microcolonies on the surfaces of tomato cells adjacent to intercellular spaces, and then aggregated surrounded by an extracellular matrix, forming mature biofilm structures. Furthermore, OE1-1 cells produced mushroom-type biofilms when incubated in fluids of apoplasts including intercellular spaces, but not xylem fluids from tomato plants. This is the first report of biofilm formation by R. solanacearum on host plant cells after invasion into intercellular spaces and mushroom-type biofilms produced by R. solanacearum in vitro. Sugar application led to enhanced biofilm formation by OE1-1. Mutation of lecM encoding a lectin, RS-IIL, which reportedly exhibits affinity for these sugars, led to a significant decrease in biofilm formation. Colonization in intercellular spaces was significantly decreased in the lecM mutant, leading to a loss of virulence on tomato plants. Complementation of the lecM mutant with native lecM resulted in the recovery of mushroom-type biofilms and virulence on tomato plants. Together, our findings indicate that OE1-1 produces mature biofilms on the surfaces of tomato cells after invasion into intercellular spaces. RS-IIL may contribute to biofilm formation by OE1-1, which is required for OE1-1 virulence. PMID:26609568

  8. The vascular plant-pathogenic bacterium Ralstonia solanacearum produces biofilms required for its virulence on the surfaces of tomato cells adjacent to intercellular spaces.

    PubMed

    Mori, Yuka; Inoue, Kanako; Ikeda, Kenichi; Nakayashiki, Hitoshi; Higashimoto, Chikaki; Ohnishi, Kouhei; Kiba, Akinori; Hikichi, Yasufumi

    2016-08-01

    The mechanism of colonization of intercellular spaces by the soil-borne and vascular plant-pathogenic bacterium Ralstonia solanacearum strain OE1-1 after invasion into host plants remains unclear. To analyse the behaviour of OE1-1 cells in intercellular spaces, tomato leaves with the lower epidermis layers excised after infiltration with OE1-1 were observed under a scanning electron microscope. OE1-1 cells formed microcolonies on the surfaces of tomato cells adjacent to intercellular spaces, and then aggregated surrounded by an extracellular matrix, forming mature biofilm structures. Furthermore, OE1-1 cells produced mushroom-type biofilms when incubated in fluids of apoplasts including intercellular spaces, but not xylem fluids from tomato plants. This is the first report of biofilm formation by R. solanacearum on host plant cells after invasion into intercellular spaces and mushroom-type biofilms produced by R. solanacearum in vitro. Sugar application led to enhanced biofilm formation by OE1-1. Mutation of lecM encoding a lectin, RS-IIL, which reportedly exhibits affinity for these sugars, led to a significant decrease in biofilm formation. Colonization in intercellular spaces was significantly decreased in the lecM mutant, leading to a loss of virulence on tomato plants. Complementation of the lecM mutant with native lecM resulted in the recovery of mushroom-type biofilms and virulence on tomato plants. Together, our findings indicate that OE1-1 produces mature biofilms on the surfaces of tomato cells after invasion into intercellular spaces. RS-IIL may contribute to biofilm formation by OE1-1, which is required for OE1-1 virulence.

  9. Syncytial-Type Cell Plates

    PubMed Central

    Otegui, Marisa; Staehelin, L. Andrew

    2000-01-01

    Cell wall formation in the syncytial endosperm of Arabidopsis was studied by using high-pressure-frozen/freeze-substituted developing seeds and immunocytochemical techniques. The endosperm cellularization process begins at the late globular embryo stage with the synchronous organization of small clusters of oppositely oriented microtubules (∼10 microtubules in each set) into phragmoplast-like structures termed mini-phragmoplasts between both sister and nonsister nuclei. These mini-phragmoplasts produce a novel kind of cell plate, the syncytial-type cell plate, from Golgi-derived vesicles ∼63 nm in diameter, which fuse by way of hourglass-shaped intermediates into wide (∼45 nm in diameter) tubules. These wide tubules quickly become coated and surrounded by a ribosome-excluding matrix; as they grow, they branch and fuse with each other to form wide tubular networks. The mini-phragmoplasts formed between a given pair of nuclei produce aligned tubular networks that grow centrifugally until they merge into a coherent wide tubular network with the mini-phragmoplasts positioned along the network margins. The individual wide tubular networks expand laterally until they meet and eventually fuse with each other at the sites of the future cell corners. Transformation of the wide tubular networks into noncoated, thin (∼27 nm in diameter) tubular networks begins at multiple sites and coincides with the appearance of clathrin-coated budding structures. After fusion with the syncytial cell wall, the thin tubular networks are converted into fenestrated sheets and cell walls. Immunolabeling experiments show that the cell plates and cell walls of the endosperm differ from those of the embryo and maternal tissue in two features: their xyloglucans lack terminal fucose residues on the side chain, and callose persists in the cell walls after the cell plates fuse with the parental plasma membrane. The lack of terminal fucose residues on xyloglucans suggests that these cell wall

  10. Replication Fork Velocities at Adjacent Replication Origins Are Coordinately Modified during DNA Replication in Human Cells

    PubMed Central

    Conti, Chiara; Saccà, Barbara; Herrick, John; Lalou, Claude; Pommier, Yves

    2007-01-01

    The spatial organization of replicons into clusters is believed to be of critical importance for genome duplication in higher eukaryotes, but its functional organization still remains to be fully clarified. The coordinated activation of origins is insufficient on its own to account for a timely completion of genome duplication when interorigin distances vary significantly and fork velocities are constant. Mechanisms coordinating origin distribution with fork progression are still poorly elucidated, because of technical difficulties of visualizing the process. Taking advantage of a single molecule approach, we delineated and compared the DNA replication kinetics at the genome level in human normal primary and malignant cells. Our results show that replication forks moving from one origin, as well as from neighboring origins, tend to exhibit the same velocity, although the plasticity of the replication program allows for their adaptation to variable interorigin distances. We also found that forks that emanated from closely spaced origins tended to move slower than those associated with long replicons. Taken together, our results indicate a functional role for origin clustering in the dynamic regulation of genome duplication. PMID:17522385

  11. Clara cell adenomas of the mouse lung. Interaction with alveolar type 2 cells.

    PubMed

    Palmer, K C

    1985-09-01

    Multiple pulmonary adenomas were induced in the offspring of pregnant Swiss-Webster mice by transplacental exposure to ethylnitrosourea (ENU) on the 15th day of gestation. Development and growth of tumors were followed for up to a year after birth. Morphologic assessment indicated that the majority of adenomas were of Clara-cell origin and were relatively normal on the basis of structural features. Histochemical studies, utilizing nitroblue tetrazolium reductase activity as a marker for normal Clara cells demonstrated that the Clara-cell-derived tumors possessed nearly normal enzyme activity. Microscopic studies of the tumors and adjacent parenchyma revealed a unique Type 2 cell response to the presence of Clara-cell adenomas occurring in the alveoli beyond the margins of the tumor. Otherwise normal-appearing Type 2 cells, in a narrow zone around the Clara-cell tumors, accumulated large amounts of surfactantlike osmiophilic lamellar material within cytoplasmic vacuoles as early as 30 days after birth. These changes were clearly a Clara-cell-tumor-related response, and not seen in association with other non-Clara-cell adenomas of the same lung. Furthermore, the alterations occurred exclusively in Type 2 cells. The extent of Type 2 cell change was correlated with tumor size and age. Autoradiographic studies with tritiated choline showed marked incorporation of the labeled precursor by the altered Type 2 cells. By electron microscopy, these inclusions were membrane-limited and contained osmiophilic lamellar structures similar to lamellar bodies in normal Type 2 cells. Because these Clara cell adenomas may act as a concentrated focus of normal Clara cells, the alterations seen in Type 2 cells may reflect an amplification of a normal interaction between bronchiolar Clara cells and alveolar Type 2 cells in the centriacinar and juxtabronchiolar alveoli.

  12. Mitochondria “fuel” breast cancer metabolism: Fifteen markers of mitochondrial biogenesis label epithelial cancer cells, but are excluded from adjacent stromal cells

    PubMed Central

    Sotgia, Federica; Whitaker-Menezes, Diana; Martinez-Outschoorn, Ubaldo E.; Salem, Ahmed F.; Tsirigos, Aristotelis; Lamb, Rebecca; Sneddon, Sharon; Hulit, James; Howell, Anthony; Lisanti, Michael P.

    2012-01-01

    Here, we present new genetic and morphological evidence that human tumors consist of two distinct metabolic compartments. First, re-analysis of genome-wide transcriptional profiling data revealed that > 95 gene transcripts associated with mitochondrial biogenesis and/or mitochondrial translation were significantly elevated in human breast cancer cells, as compared with adjacent stromal tissue. Remarkably, nearly 40 of these upregulated gene transcripts were mitochondrial ribosomal proteins (MRPs), functionally associated with mitochondrial translation of protein components of the OXPHOS complex. Second, during validation by immunohistochemistry, we observed that antibodies directed against 15 markers of mitochondrial biogenesis and/or mitochondrial translation (AKAP1, GOLPH3, GOLPH3L, MCT1, MRPL40, MRPS7, MRPS15, MRPS22, NRF1, NRF2, PGC1-α, POLRMT, TFAM, TIMM9 and TOMM70A) selectively labeled epithelial breast cancer cells. These same mitochondrial markers were largely absent or excluded from adjacent tumor stromal cells. Finally, markers of mitochondrial lipid synthesis (GOLPH3) and mitochondrial translation (POLRMT) were associated with poor clinical outcome in human breast cancer patients. Thus, we conclude that human breast cancers contain two distinct metabolic compartments—a glycolytic tumor stroma, which surrounds oxidative epithelial cancer cells—that are mitochondria-rich. The co-existence of these two compartments is indicative of metabolic symbiosis between epithelial cancer cells and their surrounding stroma. As such, epithelial breast cancer cells should be viewed as predatory metabolic “parasites,” which undergo anabolic reprogramming to amplify their mitochondrial “power.” This notion is consistent with the observation that the anti-malarial agent chloroquine may be an effective anticancer agent. New anticancer therapies should be developed to target mitochondrial biogenesis and/or mitochondrial translation in human cancer cells. PMID

  13. The growth and structure of double-diffusive cells adjacent to a cooled sidewall in a salt-stratified environment

    NASA Astrophysics Data System (ADS)

    Malki-Epshtein, Liora; Phillips, Owen M.; Huppert, Herbert E.

    2004-11-01

    Observations and measurements are reported on the patterns and rates of growth in time of the double-diffusive cells that form adjacent to a cooled sidewall in a saltstratified environment. Fluid near the wall is cooled and sinks a distance h where its density, increased by cooling, matches that of the salt-stratified ambient. The fluid separates from the wall, moving outwards as a cool, fresher layer beneath a warmer, more saline region. This leads to growing double-diffusive cells that advance outward at a rate, found by dimensional reasoning, to initially be proportional to N_{0}h, where N_{0} is the initial buoyancy frequency in the ambient and h is the intrusion's vertical thickness. Near the wall at the top of each cell, the sinking colder fluid is continually replaced by selective withdrawal from the ambient ‘far field’. The fluid being withdrawn from the ambient is always the least dense in the cell, and as the experiment proceeds, the straining of the fluid in the ambient region reduces the stratification. The vertical density gradient inside the cell relaxes by continuous hydrostatic adjustment (CHA) to match the ambient and the speed of advance reduces. Measurements of the rate of advance of the cell nose were made in tanks of different lengths L with a range of initial salinity gradients and temperature differences. A simple two-dimensional model is developed to describe the rate of extension of the cells and the internal density gradient as functions of time in which the tank length appears as an important variable. This effect does not seem to have been recognized previously. The rates of evolution in each run involve the time scale tau {=} L /( {C_H hN_0 }), where C_H {≈} 10({) - 2} is a heat transfer coefficient. The mean length of the cells skew2bar {l}(t)and the internal buoyancy frequency as functions of time are given by [ skew2bar {l}(t) / L = t/tau - ( t/2tau)^2,quad N = N_0 (1 - t / 2tau ). ] Inversion of the first of these expressions

  14. Periostin differentially induces proliferation, contraction and apoptosis of primary Dupuytren’s disease and adjacent palmar fascia cells

    PubMed Central

    Vi, Linda; Feng, Lucy; Zhu, Rebecca D.; Wu, Yan; Satish, Latha; Gan, Bing Siang; O’Gorman, David B.

    2016-01-01

    Dupuytren’s disease, (DD), is a fibroproliferative condition of the palmar fascia in the hand, typically resulting in permanent contracture of one or more fingers. This fibromatosis is similar to scarring and other fibroses in displaying excess collagen secretion and contractile myofibroblast differentiation. In this report we expand on previous data demonstrating that POSTN mRNA, which encodes the extra-cellular matrix protein periostin, is up-regulated in Dupuytren’s disease cord tissue relative to phenotypically normal palmar fascia. We demonstrate that the protein product of POSTN, periostin, is abundant in Dupuytren’s disease cord tissue while little or no periostin immunoreactivity is evident in patient-matched control tissues. The relevance of periostin up-regulation in DD was assessed in primary cultures of cells derived from diseased and phenotypically unaffected palmar fascia from the same patients. These cells were grown in type-1 collagen-enriched culture conditions with or without periostin addition to more closely replicate the in vivo environment. Periostinwas found to differentially regulate the apoptosis, proliferation, α smooth muscle actin expression and stressed Fibroblast Populated Collagen Lattice contraction of these cell types. We hypothesize that periostin, secreted by disease cord myofibroblasts into the extra-cellular matrix, promotes the transition of resident fibroblasts in the palmar fascia toward a myofibroblast phenotype, thereby promoting disease progression. PMID:19619531

  15. Border of Notch activity establishes a boundary between the two dorsal appendage tube cell types.

    PubMed

    Ward, Ellen J; Zhou, Xiaofeng; Riddiford, Lynn M; Berg, Celeste A; Ruohola-Baker, Hannele

    2006-09-15

    Boundaries establish and maintain separate populations of cells critical for organ formation. We show that Notch signaling establishes the boundary between two types of post-mitotic epithelial cells, the Rhomboid- and the Broad-positive cells. These cells will undergo morphogenetic movements to generate the two sides of a simple organ, the dorsal appendage tube of the Drosophila egg chamber. The boundary forms due to a difference in Notch levels in adjacent cells. The Notch expression pattern mimics the boundary; Notch levels are high in Rhomboid cells and low in Broad cells. Notch(-) mutant clones generate an ectopic boundary: ectopic Rhomboid cells arise in Notch(+) cells adjacent to the Notch(-) mutant cells but not further away from the clonal border. Pangolin, a component of the Wingless pathway, is required for Broad expression and for rhomboid repression. We further show that Broad represses rhomboid cell autonomously. Our data provide a foundation for understanding how a single row of Rhomboid cells arises adjacent to the Broad cells in the dorsal appendage primordia. Generating a boundary by the Notch pathway might constitute an evolutionarily conserved first step during organ formation in many tissues. PMID:16828735

  16. Cell-Type-Specific Optogenetics in Monkeys.

    PubMed

    Namboodiri, Vijay Mohan K; Stuber, Garret D

    2016-09-01

    The recent advent of technologies enabling cell-type-specific recording and manipulation of neuronal activity spurred tremendous progress in neuroscience. However, they have been largely limited to mice, which lack the richness in behavior of primates. Stauffer et al. now present a generalizable method for achieving cell-type specificity in monkeys. PMID:27610562

  17. A Cell-type-resolved Liver Proteome*

    PubMed Central

    Ding, Chen; Li, Yanyan; Guo, Feifei; Jiang, Ying; Ying, Wantao; Li, Dong; Yang, Dong; Xia, Xia; Liu, Wanlin; Zhao, Yan; He, Yangzhige; Li, Xianyu; Sun, Wei; Liu, Qiongming; Song, Lei; Zhen, Bei; Zhang, Pumin; Qian, Xiaohong; Qin, Jun; He, Fuchu

    2016-01-01

    Parenchymatous organs consist of multiple cell types, primarily defined as parenchymal cells (PCs) and nonparenchymal cells (NPCs). The cellular characteristics of these organs are not well understood. Proteomic studies facilitate the resolution of the molecular details of different cell types in organs. These studies have significantly extended our knowledge about organogenesis and organ cellular composition. Here, we present an atlas of the cell-type-resolved liver proteome. In-depth proteomics identified 6000 to 8000 gene products (GPs) for each cell type and a total of 10,075 GPs for four cell types. This data set revealed features of the cellular composition of the liver: (1) hepatocytes (PCs) express the least GPs, have a unique but highly homogenous proteome pattern, and execute fundamental liver functions; (2) the division of labor among PCs and NPCs follows a model in which PCs make the main components of pathways, but NPCs trigger the pathways; and (3) crosstalk among NPCs and PCs maintains the PC phenotype. This study presents the liver proteome at cell resolution, serving as a research model for dissecting the cell type constitution and organ features at the molecular level. PMID:27562671

  18. NK Cells and Type 1 Diabetes

    PubMed Central

    Rodacki, Melanie; Milech, Adolpho; de Oliveira, José Egídio Paulo

    2006-01-01

    Type 1 diabetes (T1D) is characterized by an immuno-mediated progressive destruction of the pancreatic β cells. Due to the ability of NK cells to kill target cells as well as to interact with antigen-presenting and T cells, it has been suggested that they could be involved in one or multiple steps of the immune-mediated attack that leads to T1D. Abnormalities in the frequency and activity of NK cells have been described both in animal models and patients with T1D. Some of these alterations are linked to its onset while others seem to be a consequence of the disease. Here, we discuss the main characteristics of NK cells and review the studies that investigated the role of NK cells in T1D, both in mouse models and humans. PMID:17162353

  19. Adjacent segment disease.

    PubMed

    Virk, Sohrab S; Niedermeier, Steven; Yu, Elizabeth; Khan, Safdar N

    2014-08-01

    EDUCATIONAL OBJECTIVES As a result of reading this article, physicians should be able to: 1. Understand the forces that predispose adjacent cervical segments to degeneration. 2. Understand the challenges of radiographic evaluation in the diagnosis of cervical and lumbar adjacent segment disease. 3. Describe the changes in biomechanical forces applied to adjacent segments of lumbar vertebrae with fusion. 4. Know the risk factors for adjacent segment disease in spinal fusion. Adjacent segment disease (ASD) is a broad term encompassing many complications of spinal fusion, including listhesis, instability, herniated nucleus pulposus, stenosis, hypertrophic facet arthritis, scoliosis, and vertebral compression fracture. The area of the cervical spine where most fusions occur (C3-C7) is adjacent to a highly mobile upper cervical region, and this contributes to the biomechanical stress put on the adjacent cervical segments postfusion. Studies have shown that after fusion surgery, there is increased load on adjacent segments. Definitive treatment of ASD is a topic of continuing research, but in general, treatment choices are dictated by patient age and degree of debilitation. Investigators have also studied the risk factors associated with spinal fusion that may predispose certain patients to ASD postfusion, and these data are invaluable for properly counseling patients considering spinal fusion surgery. Biomechanical studies have confirmed the added stress on adjacent segments in the cervical and lumbar spine. The diagnosis of cervical ASD is complicated given the imprecise correlation of radiographic and clinical findings. Although radiological and clinical diagnoses do not always correlate, radiographs and clinical examination dictate how a patient with prolonged pain is treated. Options for both cervical and lumbar spine ASD include fusion and/or decompression. Current studies are encouraging regarding the adoption of arthroplasty in spinal surgery, but more long

  20. Cell culture models using rat primary alveolar type I cells

    PubMed Central

    Downs, Charles A.; Montgomery, David W.; Merkle, Carrie J.

    2011-01-01

    There is a lack of cell culture models using primary alveolar type I (AT I) cells. The purpose of this study was to develop cell culture models using rat AT I cells and microvascular endothelial cells from the lung (MVECL). Two types of model systems were developed: single and co-culture systems; additionally a 3-dimensional model system was developed. Pure AT I cell (96.3 ±2.7%) and MVECL (97.9 ±1.1 %) preparations were used. AT I cell morphology, mitochondrial number and distribution, actin filament arrangement and number of apoptotic cells at confluence, and telomere attrition were characterized. AT I cells maintained their morphometric characteristics through at least population doubling (PD) 35, while demonstrating telomere attrition through at least PD 100. Furthermore, AT I cells maintained the expression of their specific markers, T1α and AQ-5, through PD 42. For the co-cultures, AT I cells were grown on the top and MVECL were grown on the bottom of fibronectin coated 24 well Transwell Fluroblok™ filter inserts. Neither cell type transmigrated the 1 micron pores. Additionally AT I cells were grown in a thick layer of Matrigel® to create a 3-dimensional model in which primary AT I cells form ring-like structures that resemble an alveolus. The development of these model systems offers the opportunities to investigate AT I cell cells and their interactions with MVECL in response to pharmacological interventions and in the processes of disease, repair and regeneration. PMID:21624488

  1. Cell elasticity with altered cytoskeletal architectures across multiple cell types.

    PubMed

    Grady, Martha E; Composto, Russell J; Eckmann, David M

    2016-08-01

    The cytoskeleton is primarily responsible for providing structural support, localization and transport of organelles, and intracellular trafficking. The structural support is supplied by actin filaments, microtubules, and intermediate filaments, which contribute to overall cell elasticity to varying degrees. We evaluate cell elasticity in five different cell types with drug-induced cytoskeletal derangements to probe how actin filaments and microtubules contribute to cell elasticity and whether it is conserved across cell type. Specifically, we measure elastic stiffness in primary chondrocytes, fibroblasts, endothelial cells (HUVEC), hepatocellular carcinoma cells (HUH-7), and fibrosarcoma cells (HT 1080) subjected to two cytoskeletal destabilizers: cytochalasin D and nocodazole, which disrupt actin and microtubule polymerization, respectively. Elastic stiffness is measured by atomic force microscopy (AFM) and the disruption of the cytoskeleton is confirmed using fluorescence microscopy. The two cancer cell lines showed significantly reduced elastic moduli values (~0.5kPa) when compared to the three healthy cell lines (~2kPa). Non-cancer cells whose actin filaments were disrupted using cytochalasin D showed a decrease of 60-80% in moduli values compared to untreated cells of the same origin, whereas the nocodazole-treated cells showed no change in elasticity. Overall, we demonstrate actin filaments contribute more to elastic stiffness than microtubules but this result is cell type dependent. Cancer cells behaved differently, exhibiting increased stiffness as well as stiffness variability when subjected to nocodazole. We show that disruption of microtubule dynamics affects cancer cell elasticity, suggesting therapeutic drugs targeting microtubules be monitored for significant elastic changes. PMID:26874250

  2. Interneuron cell types are fit to function.

    PubMed

    Kepecs, Adam; Fishell, Gordon

    2014-01-16

    Understanding brain circuits begins with an appreciation of their component parts - the cells. Although GABAergic interneurons are a minority population within the brain, they are crucial for the control of inhibition. Determining the diversity of these interneurons has been a central goal of neurobiologists, but this amazing cell type has so far defied a generalized classification system. Interneuron complexity within the telencephalon could be simplified by viewing them as elaborations of a much more finite group of developmentally specified cardinal classes that become further specialized as they mature. Our perspective emphasizes that the ultimate goal is to dispense with classification criteria and directly define interneuron types by function. PMID:24429630

  3. Identification and characterization of a T-cell-specific enhancer adjacent to the murine CD4 gene.

    PubMed Central

    Sawada, S; Littman, D R

    1991-01-01

    Expression of the CD4 and CD8 glycoproteins is a tightly regulated process tied to the maturation of functionally distinct classes of thymocytes. Therefore, understanding of the mechanism of expression of the genes encoding CD4 and CD8 is likely to yield important insight into regulation of the differentiated functions of T cells. Here, we report the identification of a T-cell-specific enhancer in a DNase I-hypersensitive region about 13 kb 5' of the transcription initiation site of the murine CD4 gene. Within the minimal enhancer element, at least three nuclear protein binding sites were identified by DNase I footprint analysis. One site contains the consensus motif for TCF-1 alpha/LEF-1, a recently identified HMG box transcription factor primarily expressed in pre-B and T cells. By Southwestern (DNA-protein) blotting and binding competition analyses, the protein binding to this site was found to be indistinguishable from TCF-1 alpha/LEF-1. Mutagenesis of this site resulted in loss of factor binding but had a relatively minor effect on enhancer activity. In contrast, mutations in another site, containing two consensus binding motifs for basic helix-loop-helix proteins, abolished factor binding and dramatically reduced enhancer activity. None of the protein binding sites had activity on its own, suggesting that the CD4 enhancer requires the interaction of multiple regulatory sites. Images PMID:1922061

  4. Versatile UHV compatible Knudsen type effusion cell

    SciTech Connect

    Shukla, A.K.; Banik, S.; Dhaka, R.S.; Biswas, C.; Barman, S.R.; Haak, H.

    2004-11-01

    A versatile Knudsen type effusion cell has been fabricated for growing nanostructures and epitaxial layers of metals and semiconductors. The cell provides excellent vacuum compatibility (10{sup -10} mbar range during operation), efficient water cooling, uniform heating, and moderate input power consumption (100 W at 1000 deg. C). The thermal properties of the cell have been determined. The performance of the cell has been assessed by x-ray photoemission spectroscopy (XPS) for Mn adlayer growth on Al(111). We find that this Knudsen cell has a stable deposition rate of 0.17 monolayer per minute at 550 deg. C. From the XPS spectra, we show that the Mn adlayers are completely clean, i.e., devoid of any surface contamination.

  5. Redox modulation of adjacent thiols in VLA-4 by AS101 converts myeloid leukemia cells from a drug-resistant to drug-sensitive state.

    PubMed

    Layani-Bazar, Adi; Skornick, Itai; Berrebi, Alain; Pauker, Maor H; Noy, Elad; Silberman, Alon; Albeck, Michael; Longo, Dan L; Kalechman, Yona; Sredni, Benjamin

    2014-06-01

    Interaction between the integrin VLA-4 on acute myelogenous leukemia (AML) cells with stromal fibronectin is a decisive factor in chemotherapeutic resistance. In this study, we provide a rationale for a drug repositioning strategy to blunt integrin activation in AML cells and restore their sensitivity to chemotherapy. Specifically, we demonstrate that the nontoxic tellurium compound AS101, currently being evaluated in clinical trials, can abrogate the acquired resistance of AML. Mechanistic investigations revealed that AS101 caused redox inactivation of adjacent thiols in the exofacial domain of VLA-4 after its ligation to stromal fibronectin. This effect triggered cytoskeletal conformational changes that decreased PI3K/Akt/Bcl2 signaling, an obligatory step in chemosensitization by AS101. In a mouse xenograft of AML derived from patient leukemic cells with high VLA-4 expression and activity, we demonstrated that AS101 abrogated drug resistance and prolonged survival in mice receiving chemotherapy. Decreased integrin activity was confirmed on AML cells in vivo. The chemosensitizing activity of AS101 persisted in hosts with defective adaptive and innate immunity, consistent with evidence that integrin deactivation was not mediated by heightening immune attack. Our findings provide a mechanistic rationale to reposition the experimental clinical agent, AS101, to degrade VLA-4-mediated chemoresistance and improve clinical responses in patients with AML.

  6. Overcoming the Refractory Expression of Secreted Recombinant Proteins in Mammalian Cells through Modification of the Signal Peptide and Adjacent Amino Acids

    PubMed Central

    Güler-Gane, Gülin; Kidd, Sara; Sridharan, Sudharsan; Vaughan, Tristan J.; Wilkinson, Trevor C. I.

    2016-01-01

    The expression and subsequent purification of mammalian recombinant proteins is of critical importance to many areas of biological science. To maintain the appropriate tertiary structure and post-translational modifications of such proteins, transient mammalian expression systems are often adopted. The successful utilisation of these systems is, however, not always forthcoming and some recombinant proteins prove refractory to expression in mammalian hosts. In this study we focussed on the role of different N-terminal signal peptides and residues immediately downstream, in influencing the level of secreted recombinant protein obtained from suspension HEK293 cells. Using secreted alkaline phosphatase (SEAP) as a model protein, we identified that the +1/+2 downstream residues flanking a heterologous signal peptide significantly affect secreted levels. By incorporating these findings we conducted a comparison of different signal peptide sequences and identified the most productive as secrecon, a computationally-designed sequence. Importantly, in the context of the secrecon signal peptide and SEAP, we also demonstrated a clear preference for specific amino acid residues at the +1 position (e.g. alanine), and a detrimental effect of others (cysteine, proline, tyrosine and glutamine). When proteins that naturally contain these “undesirable” residues at the +1 position were expressed with their native signal peptide, the heterologous secrecon signal peptide, or secrecon with an additional alanine at the +1 or +1 and +2 position, the level of expression differed significantly and in an unpredictable manner. For each protein, however, at least one of the panel of signal peptide/adjacent amino acid combinations enabled successful recombinant expression. In this study, we highlight the important interplay between a signal peptide and its adjacent amino acids in enabling protein expression, and we describe a strategy that could enable recombinant proteins that have so far

  7. Tumor-induced solid stress activates β-catenin signaling to drive malignant behavior in normal, tumor-adjacent cells

    PubMed Central

    Ou, Guanqing; Weaver, Valerie Marie

    2016-01-01

    Recent work by Fernández-Sánchez and coworkers examining the impact of applied pressure on the malignant phenotype of murine colon tissue in vivo revealed that mechanical perturbations can drive malignant behavior in genetically normal cells. Their findings build upon an existing understanding of how the mechanical cues experienced by cells within a tissue become progressively modified as the tissue transforms. Using magnetically stimulated ultra-magnetic liposomes to mimic tumor growth -induced solid stress, Fernández-Sánchez and coworkers were able to stimulate β-catenin to promote the cancerous behavior of both a normal and genetically modified colon epithelium. In this perspective, we discuss their findings in the context of what is currently known regarding the role of the mechanical landscape in cancer progression and β-catenin as a mechanotransducer. We review data that suggest that mechanically regulated activation of β-catenin fosters development of a malignant phenotype in tissue and predict that mechanical cues may contribute to tumor heterogeneity. PMID:26439949

  8. Lipid Profiles of Canine Invasive Transitional Cell Carcinoma of the Urinary Bladder and Adjacent Normal Tissue by Desorption Electrospray Ionization Imaging Mass Spectrometry

    PubMed Central

    Dill, Allison L.; Ifa, Demian R.; Manicke, Nicholas E.; Costa, Anthony B.; Ramos-Vara, José A.; Knapp, Deborah W.; Cooks, R. Graham

    2009-01-01

    Desorption electrospray ionization (DESI) mass spectrometry (MS) was used in an imaging mode to interrogate the lipid profiles of thin tissue sections of canine spontaneous invasive transitional cell carcinoma (TCC) of the urinary bladder (a model of human invasive bladder cancer) as well as adjacent normal tissue from four different dogs. The glycerophospholipids and sphingolipids that appear as intense signals in both the negative ion and positive ion modes were identified by tandem mass spectrometry (MS/MS) product ion scans using collision-induced dissociation. Differences in the relative distributions of the lipid species were present between the tumor and adjacent normal tissue in both the negative and positive ion modes. DESI-MS images showing the spatial distributions of particular glycerophospholipids, sphinoglipids and free fatty acids in both the negative and positive ion modes were compared to serial tissue sections that were stained with hematoxylin and eosin (H&E). Increased absolute and relative intensities for at least five different glycerophospholipids and three free fatty acids in the negative ion mode and at least four different lipid species in the positive ion mode were seen in the tumor region of the samples in all four dogs. In addition, one sphingolipid species exhibited increased signal intensity in the positive ion mode in normal tissue relative to the diseased tissue. Principal component analysis (PCA) was also used to generate unsupervised statistical images from the negative ion mode data and these images are in excellent agreement with the DESI images obtained from the selected ions and also the H&E stained tissue PMID:19810710

  9. Cytotoxic Effects and Osteogenic Activity of Calcium Sulfate with and without Recombinant Human Bone Morphogenetic Protein 2 and Nano-Hydroxyapatite Adjacent to MG-63 Cell Line

    PubMed Central

    Ghorbanzadeh, Abdollah; Aminsobhani, Mohsen; Khoshzaban, Ahad; Abbaszadeh, Armin; Ghorbanzadeh, Atiyeh; Shamshiri, Ahmad Reza

    2015-01-01

    Objectives: The aim of this study was to assess the cytotoxic effects and osteogenic activity of recombinant human bone morphogenetic protein (rhBMP2) and nano-hydroxyapatite (n-HA) adjacent to MG-63 cell line. Materials and Methods: To assess cytotoxicity, the 4,5-dimethyl thiazolyl-2,5-diphenyl tetrazolium bromide (MTT) assay was used. Alkaline phosphatase (ALP) activity and osteogenic activity were evaluated using Alizarin red and the von Kossa staining and analyzed by one-way ANOVA followed by Tukey’s post hoc test. Results: The n-HA/calcium sulfate (CS) mixture significantly promoted cell growth in comparison to pure CS. Moreover, addition of rhBMP2 to CS (P=0.02) and also mixing CS with n-HA led to further increase in extracellular calcium production and ALP activity (P=0.03). Conclusion: This in vitro study indicates that a scaffold material in combination with an osteoinductive material is effective for bone matrix formation. PMID:26877731

  10. Functionally deficient mesenchymal stem cells reside in the bone marrow niche with M2-macrophages and amyloid-β protein adjacent to loose total joint implants.

    PubMed

    Margulies, Bryan S; DeBoyace, Sean D; Parsons, Adrienne M; Policastro, Connor G; Ee, Jessica S S; Damron, Timothy S

    2015-05-01

    We sought to demonstrate whether there is a difference in the local mesenchymal stem cells (MSC) niche obtained from patients undergoing their first total joint replacement surgery versus those patients undergoing a revision surgery for an failing total joint implant. Bone marrow aspirates collected from patients undergoing revision total joint arthroplasty were observed to be less clonal and the expression of PDGFRα, CD51, ALCAM, endoglin, CXCL12, nestin, and nucleostemin were decreased. Revision MSC were also less able to commit to an osteoblast-lineage or an adipocyte-lineage. Further, in revision MSC, OPG, and IL6 expression were increased. Monocytes, derived from revision whole marrow aspirates, were less capable of differentiating into osteoclasts, the cells implicated in the pathologic degradation of bone. Osteoclasts were also not observed in tissue samples collected adjacent to the implants of revision patients; however, the alternatatively activated M2-macrophage phenotype was observed in parallel with pathologic accumulations of amyloid-β, τ-protien and 3-nitrotyrosine. Despite the limited numbers of patients examined, our data suggest that nucleostemin may be a useful functional marker for MSC while the observation of M2-macrophage infiltration around the implant lays the foundation for future investigation into a novel mechanism that we propose is associated with loose total joint implants.

  11. Cell type-specific bipolar cell input to ganglion cells in the mouse retina.

    PubMed

    Neumann, S; Hüser, L; Ondreka, K; Auler, N; Haverkamp, S

    2016-03-01

    Many distinct ganglion cell types, which are the output elements of the retina, were found to encode for specific features of a visual scene such as contrast, color information or movement. The detailed composition of retinal circuits leading to this tuning of retinal ganglion cells, however, is apart from some prominent examples, largely unknown. Here we aimed to investigate if ganglion cell types in the mouse retina receive selective input from specific bipolar cell types or if they sample their synaptic input non-selectively from all bipolar cell types stratifying within their dendritic tree. To address this question we took an anatomical approach and immunolabeled retinae of two transgenic mouse lines (GFP-O and JAM-B) with markers for ribbon synapses and type 2 bipolar cells. We morphologically identified all green fluorescent protein (GFP)-expressing ganglion cell types, which co-stratified with type 2 bipolar cells and assessed the total number of bipolar input synapses and the proportion of synapses deriving from type 2 bipolar cells. Only JAM-B ganglion cells received synaptic input preferentially from bipolar cell types other than type 2 bipolar cells whereas the other analyzed ganglion cell types sampled their bipolar input most likely from all bipolar cell terminals within their dendritic arbor.

  12. The 1993 cell typings of the International Cell Exchange.

    PubMed

    Lau, M; Terasaki, P I; Park, M S

    1993-01-01

    1. This is a summary of the typings for 40 cells for Class I antigens and 20 cultured cell lines for Class II antigens through the International Cell Exchange. Serological typings were compared with DNA typing reports for Class II specificities. Presently, 283 laboratories participate in the monthly Class I exchange. Class II results were received from 124 serology labs and 81 DNA labs on a monthly basis. 2. In 1993, 12 A-locus antigens were typed and 8 specificities reached levels of 95% or greater average detection. Thirteen of the 33 B-locus antigens showed 95% or better mean agreement levels. There was an improvement in detection of B76 and B7801. 3. Discrepancy rates of 7 A-locus and 9 B-locus antigens typed 3 or more times were compared with the overall rates for each respective locus. The discrepancy rate of false negatives, ie, how often the antigen was missed for the recognized B-locus specificities, continued to be greater than those for the A-locus antigens. The discrepancy rates, especially the percent false-positive, decreased for A33 during the recent 6-year period. 4. We showed the number of labs with their total of false-negatives and false-positives. Twelve labs attained a final total of no misses for all antigens. In 1993, 11 labs achieved impressive perfect records (zero false negative and false positive) for all analyzed antigens. 5. Retyping results of 2 donors showed improved antigen detection, particularly of A2403, B70, and B76. 6. Eleven cells typed in previous cell exchanges as having new or rare variants were sequenced recently. The B*5102 and B*5901 cells were retyped as reference cells. A new A-locus variant detected in previous exchanges was recently confirmed by sequence work as A*8001. New variants of B5 and B22 were discussed. 7. In addition to the mean detection rates, the low and high levels were determined for 15 broad (11 DR & 4 DQ) specificities by serology and compared with those attained for the respective generic (low

  13. Plant single-cell and single-cell-type metabolomics.

    PubMed

    Misra, Biswapriya B; Assmann, Sarah M; Chen, Sixue

    2014-10-01

    In conjunction with genomics, transcriptomics, and proteomics, plant metabolomics is providing large data sets that are paving the way towards a comprehensive and holistic understanding of plant growth, development, defense, and productivity. However, dilution effects from organ- and tissue-based sampling of metabolomes have limited our understanding of the intricate regulation of metabolic pathways and networks at the cellular level. Recent advances in metabolomics methodologies, along with the post-genomic expansion of bioinformatics knowledge and functional genomics tools, have allowed the gathering of enriched information on individual cells and single cell types. Here we review progress, current status, opportunities, and challenges presented by single cell-based metabolomics research in plants.

  14. Defining cell types and states with single-cell genomics

    PubMed Central

    Trapnell, Cole

    2015-01-01

    A revolution in cellular measurement technology is under way: For the first time, we have the ability to monitor global gene regulation in thousands of individual cells in a single experiment. Such experiments will allow us to discover new cell types and states and trace their developmental origins. They overcome fundamental limitations inherent in measurements of bulk cell population that have frustrated efforts to resolve cellular states. Single-cell genomics and proteomics enable not only precise characterization of cell state, but also provide a stunningly high-resolution view of transitions between states. These measurements may finally make explicit the metaphor that C.H. Waddington posed nearly 60 years ago to explain cellular plasticity: Cells are residents of a vast “landscape” of possible states, over which they travel during development and in disease. Single-cell technology helps not only locate cells on this landscape, but illuminates the molecular mechanisms that shape the landscape itself. However, single-cell genomics is a field in its infancy, with many experimental and computational advances needed to fully realize its full potential. PMID:26430159

  15. Type specimens of Heteroptera (Insecta: Hemiptera) collected from North Korea and adjacent regions deposited at Insect Collections of Chungnam National University (CNU) in Daejeon, Republic of Korea.

    PubMed

    Jung, Sunghoon; Kim, Junggon; Oh, Sumin; Heiss, Ernst

    2015-07-06

    A list of type specimens of Heteroptera (Insecta: Hemiptera) collected from North Korea (mostly by the late Dr. Michail Josifov, Sofia, Bulgaria) acquired earlier by E. Heiss, now donated to and deposited in the insect collections of Chungnam National University (CNU), Deajeon, Korea, is presented. A total of 31 holotypes and 694 paratypes of 41 species and 1 subspecies in 6 families and 9 subfamilies are presented: Miridae (Deraeocorinae, Mirinae, Orthotylinae, Phylinae), Tingidae (Tinginae), Piesmatidae (Piesmatinae), Berytidae (Metacanthinae), Cymidae (Cyminae), Pentatomidae (Asopinae).

  16. Screen printed SIS-type solar cells

    NASA Astrophysics Data System (ADS)

    Avaritsiotis, J. N.; Caroubalos, C.; Campbell, D. S.

    Experimentation was performed on the formation of an SIS junction on n-type monocrystalling Si wafers using thick film techniques and an indium-tin oxide thick film paste. A SiO layer 20 A thick grew on the optically smooth side of monocrystalline wafers during the firing of a screen printed Au back contact. An ITO film 2000 A thick was grown on the oxidized surface and a silver front thick-film grid was fired on the other side. Various firing temperatures were examined to test the effects on the open circuit voltage and the current density. The best samples displayed an open circuit voltage of 293 mV for an illumination of 55-90 mW/sq cm, after which the short circuit current fell off. The best cell efficiencies were less than one, although the performance degraded less than 10 pct in 6 mos.

  17. Activation of Type II Cells into Regenerative Stem Cell Antigen-1+ Cells during Alveolar Repair

    PubMed Central

    Kumar, Varsha Suresh; Zhang, Wei; Rehman, Jalees; Malik, Asrar B.

    2015-01-01

    The alveolar epithelium is composed of two cell types: type I cells comprise 95% of the gas exchange surface area, whereas type II cells secrete surfactant, while retaining the ability to convert into type I cells to induce alveolar repair. Using lineage-tracing analyses in the mouse model of Pseudomonas aeruginosa–induced lung injury, we identified a population of stem cell antigen (Sca)-1–expressing type II cells with progenitor cell properties that mediate alveolar repair. These cells were shown to be distinct from previously reported Sca-1–expressing bronchioalveolar stem cells. Microarray and Wnt reporter studies showed that surfactant protein (Sp)-C+Sca-1+ cells expressed Wnt signaling pathway genes, and inhibiting Wnt/β-catenin signaling prevented the regenerative function of Sp-C+Sca-1+ cells in vitro. Thus, P. aeruginosa–mediated lung injury induces the generation of a Sca-1+ subset of type II cells. The progenitor phenotype of the Sp-C+Sca-1+ cells that mediates alveolar epithelial repair might involve Wnt signaling. PMID:25474582

  18. Patterns of Distribution of Macro-fauna in Different Types of Estuarine, Soft Sediment Habitats Adjacent to Urban and Non-urban Areas

    NASA Astrophysics Data System (ADS)

    Lindegarth, M.; Hoskin, M.

    2001-02-01

    Urban development typically creates a large number of potentially interacting disturbances that may cause impacts on assemblages of animals and plans in estuarine habitats. We tested predictions from the general model that intertidal areas exposed to different types of disturbances have different types of assemblages of benthic macrofauna. Different parts of the Port Hacking Estuary (New South Wales, Australia) are exposed to varying degrees of disturbance by human activities. We predicted that the average structure of assemblages of intertidal animals, and patterns of variability would differ between urban and non-urban areas of Port Hacking. Consistent with previous observations from the literature, there were differences in average structure between urban and non-urban sandy areas. Qualitative differences between abundances of individual taxa in urban and non-urban areas were generally not consistent with previous observations. Differences between assemblages in urban and non-urban areas were not observed in muddy sediments, nor in sediments among mangroves and seagrass. No significant differences in variability was observed between urban and non-urban areas. Two general models may be proposed to explain the observed differences in response to urbanization in different habitats: (1) animals are exposed to different levels or combinations of disturbances in different habitats; or (2) assemblages of animals differ in sensitivity to disturbances among habitats.

  19. The tadpole of Amazophrynella manaos Rojas, Carvalho, Gordo, Ávila, Farias and Hrbek, 2014 (Anura, Bufonidae) from the type locality and adjacent regions at Central Amazonia, Brazil.

    PubMed

    Menin, Marcelo; Pegorini, Reysi Jhayne; De Carvalho, Vinicius Tadeu; Rojas, Rommel Roberto; Gordo, Marcelo

    2014-06-30

    The genus Amazophrynella, as currently recognized (Fouquet et al. 2012a, b), is represented by four nominal species (Frost 2014; Rojas et al. 2014) but the tadpoles of only one species, Amazophrynella minuta (Melin) from Ecuador, have been described (Duellman & Lynch 1969; Duellman 1978). Amazophrynella manaos Rojas, Carvalho, Gordo, Ávila, Farias and Hrbek, 2014 occurs in the leaf litter of terra firme forest in the southwestern part of the Brazilian Guiana region (Rojas et al. 2014). The tadpole of this species was briefly described in diagrammatic drawings by Hero (1990) as Dendrophryniscus minutus. Herein, we provide a detailed description of this tadpole based on individuals at 12 stages of development collected in five different sites, including the type locality, at Central Amazonia, Brazil.

  20. Identification of positive and negative regulatory elements governing cell-type-specific expression of the neural cell adhesion molecule gene.

    PubMed Central

    Hirsch, M R; Gaugler, L; Deagostini-Bazin, H; Bally-Cuif, L; Goridis, C

    1990-01-01

    The neural cell adhesion molecule (NCAM) is one of the most prevalent cell adhesion molecules in vertebrates. Its expression is subject to complex cell-type- and developmental-stage-dependent regulation. To study this regulation at the level of transcription, we analyzed the promoter region of the mouse NCAM gene. The NCAM promoter did not contain a typical TATA box. Transcription started at several sites that were used indiscriminately by different cell types, implying that the different NCAM isoforms are expressed from a single promoter. Sequences responsible for both promotion and inhibition of transcription resided within 840 base pairs upstream of the main transcriptional start site. The sequence from positions -645 to -37 relative to the translation initiation site directed high levels of expression in NCAM-expressing N2A cells. The same fragment was six times less active but still significantly active in L cells, but this activity was repressed by inclusion of an additional upstream segment. We mapped eight domains of interactions with nuclear proteins within the 840-base-pair region. The segment with maximum promoter activity contained two adjacent footprints, the occupation of which appeared to be mutually exclusive. One of them corresponded to an Sp1-factor-binding consensus site, the other one bound a factor with nuclear factor I activity. The single protected domain in the fragment harboring a repressor activity consisted of a GGA repeat resembling negative regulatory elements in other promoters. Three adjacent binding sites occupied an A + T-rich segment and contained ATTA motifs also found in the recognition elements of homeodomain proteins. These results show that negative and positive elements interact to regulate the tissue-specific patterns of expression of the NCAM gene and indicate that a factor related to nuclear factor I is involved in its transcriptional control. Images PMID:2325642

  1. Cell type-specific transcriptome profiling in mammalian brains

    PubMed Central

    LoVerso, Peter R.; Cui, Feng

    2016-01-01

    A mammalian brain contains numerous types of cells. Advances in neuroscience in the past decade allow us to identify and isolate neural cells of interest from mammalian brains. Recent developments in high-throughput technologies, such as microarrays and next-generation sequencing (NGS), provide detailed information on gene expression in pooled cells on a genomic scale. As a result, many novel genes have been found critical in cell type-specific transcriptional regulation. These differentially expressed genes can be used as molecular signatures, unique to a particular class of neural cells. Use of this gene expression-based approach can further differentiate neural cell types into subtypes, potentially linking some of them with neurological diseases. In this article, experimental techniques used to purify neural cells are described, followed by a review on recent microarray- or NGS-based transcriptomic studies of common neural cell types. The future prospects of cell type-specific research are also discussed. PMID:27100485

  2. A web-server of cell type discrimination system.

    PubMed

    Wang, Anyou; Zhong, Yan; Wang, Yanhua; He, Qianchuan

    2014-01-01

    Discriminating cell types is a daily request for stem cell biologists. However, there is not a user-friendly system available to date for public users to discriminate the common cell types, embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and somatic cells (SCs). Here, we develop WCTDS, a web-server of cell type discrimination system, to discriminate the three cell types and their subtypes like fetal versus adult SCs. WCTDS is developed as a top layer application of our recent publication regarding cell type discriminations, which employs DNA-methylation as biomarkers and machine learning models to discriminate cell types. Implemented by Django, Python, R, and Linux shell programming, run under Linux-Apache web server, and communicated through MySQL, WCTDS provides a friendly framework to efficiently receive the user input and to run mathematical models for analyzing data and then to present results to users. This framework is flexible and easy to be expended for other applications. Therefore, WCTDS works as a user-friendly framework to discriminate cell types and subtypes and it can also be expended to detect other cell types like cancer cells. PMID:24578634

  3. A web-server of cell type discrimination system.

    PubMed

    Wang, Anyou; Zhong, Yan; Wang, Yanhua; He, Qianchuan

    2014-01-01

    Discriminating cell types is a daily request for stem cell biologists. However, there is not a user-friendly system available to date for public users to discriminate the common cell types, embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and somatic cells (SCs). Here, we develop WCTDS, a web-server of cell type discrimination system, to discriminate the three cell types and their subtypes like fetal versus adult SCs. WCTDS is developed as a top layer application of our recent publication regarding cell type discriminations, which employs DNA-methylation as biomarkers and machine learning models to discriminate cell types. Implemented by Django, Python, R, and Linux shell programming, run under Linux-Apache web server, and communicated through MySQL, WCTDS provides a friendly framework to efficiently receive the user input and to run mathematical models for analyzing data and then to present results to users. This framework is flexible and easy to be expended for other applications. Therefore, WCTDS works as a user-friendly framework to discriminate cell types and subtypes and it can also be expended to detect other cell types like cancer cells.

  4. Should tumor with direct adjacent lobe invasion (Tdali) be assigned to T2 or T3 in non-small cell lung cancer: a meta-analysis

    PubMed Central

    Xiao, Zhilan; Cao, Christphor; Mei, Jiandong; Liao, Hu; Yan, Tristan

    2016-01-01

    Background The staging of tumor with direct adjacent lobe invasion (Tdali) or interlobar invasion pleural 3 (ILI PL3) in TNM system of non-small cell lung cancer (NSCLC) is still in controversy. We conducted a meta-analysis to compare the prognosis of Tdali with T2 or T3 disease. Methods PubMed and Embase were searched for relevant studies. Ln hazard ratio (HR) and its standard error (SE) of each study were estimated in the comparison of overall survival (OS) between Tdali and T2 or T3 respectively. Forest plots were used to show the combined HRs. Results The meta-analysis for comparison of OS of Tdali and T2 or T3 disease both showed a significant HR [Tdali versus T2, 1.39 (1.21, 1.61), P<0.000, Tdali versus T3, 0.73 (0.57, 0.93), P=0.01]. Comparisons of OS of Tdali specified to T2 (Tdali-T2) and that of all patients of T2 or T3 disease also both showed significant HRs [Tdali-T2 versus T2, 1.44 (1.23, 1.69), P<0.000, Tdali-T2 versus T3, 0.77 (0.64, 0.94), P=0.008]. When only analyzing the patients with N0 status, those with Tdali-T2N0 compared to the T2N0 group had a HR of 1.79 (1.37, 2.34) (P<0.000). For those with Tdali-T2N0 compared to the T3N0 group, the HR was 0.98 (0.71, 1.35) (P=0.91). Conclusions Our meta-analysis showed that the prognosis of Tdali is poorer than T2 disease but similar to T3 disease after controlled for T and N status. We suggest that Tdali should be considered to be upgraded to T3. Our work challenges the current staging system regarding staging of Tdali, which might be important evidence of future revision of Tdali staging. As the malignancy of Tdali has been underrated till now, more attention needs to be drawn to proper treatment of Tdali patients. PMID:27621847

  5. Regulation of antiviral T cell responses by type I interferons.

    PubMed

    Crouse, Josh; Kalinke, Ulrich; Oxenius, Annette

    2015-04-01

    Type I interferons (IFNs) are pro-inflammatory cytokines that are rapidly induced in different cell types during viral infections. The consequences of type I IFN signalling include direct antiviral activity, innate immune cell activation and regulation of adaptive immune responses. In this Review, we discuss recent conceptual advances in our understanding of indirect and direct regulation of T cell immunity by type I IFNs, which can either promote or inhibit T cell activation, proliferation, differentiation and survival. This regulation depends, to a large extent, on the timing of type I IFN exposure relative to T cell receptor signalling. Type I IFNs also provide activated T cells with resistance to natural killer cell-mediated elimination. PMID:25790790

  6. Comparison of Types of Cell Death: Apoptosis and Necrosis.

    ERIC Educational Resources Information Center

    Manning, Francis; Zuzel, Katherine

    2003-01-01

    Cell death is an essential factor in many biological processes including development. Discusses two types of cell death: (1) necrosis (induced by sodium azide); and (2) apoptosis (induced by sodium chromate). Illustrates key features that differ between these two types of cells death including loss of membrane integrity and internucleosomal DNA…

  7. Cell-type specific and combinatorial usage of diverse transcription factors revealed by genome-wide binding studies in multiple human cells

    PubMed Central

    Lee, Bum-Kyu; Bhinge, Akshay A.; Battenhouse, Anna; McDaniell, Ryan M.; Liu, Zheng; Song, Lingyun; Ni, Yunyun; Birney, Ewan; Lieb, Jason D.; Furey, Terrence S.; Crawford, Gregory E.; Iyer, Vishwanath R.

    2012-01-01

    Cell-type diversity is governed in part by differential gene expression programs mediated by transcription factor (TF) binding. However, there are few systematic studies of the genomic binding of different types of TFs across a wide range of human cell types, especially in relation to gene expression. In the ENCODE Project, we have identified the genomic binding locations across 11 different human cell types of CTCF, RNA Pol II (RNAPII), and MYC, three TFs with diverse roles. Our data and analysis revealed how these factors bind in relation to genomic features and shape gene expression and cell-type specificity. CTCF bound predominantly in intergenic regions while RNAPII and MYC preferentially bound to core promoter regions. CTCF sites were relatively invariant across diverse cell types, while MYC showed the greatest cell-type specificity. MYC and RNAPII co-localized at many of their binding sites and putative target genes. Cell-type specific binding sites, in particular for MYC and RNAPII, were associated with cell-type specific functions. Patterns of binding in relation to gene features were generally conserved across different cell types. RNAPII occupancy was higher over exons than adjacent introns, likely reflecting a link between transcriptional elongation and splicing. TF binding was positively correlated with the expression levels of their putative target genes, but combinatorial binding, in particular of MYC and RNAPII, was even more strongly associated with higher gene expression. These data illuminate how combinatorial binding of transcription factors in diverse cell types is associated with gene expression and cell-type specific biology. PMID:22090374

  8. Potassium currents in rat type II alveolar epithelial cells.

    PubMed Central

    DeCoursey, T E; Jacobs, E R; Silver, M R

    1988-01-01

    1. Type II alveolar epithelial cells isolated from adult rats and grown in primary culture were studied using the whole-cell configuration of the gigohm-seal voltage clamp technique. 2. The average specific capacitance of type II cells was 2.5 microF/cm2, suggesting that type II cell membranes in vitro are irregular, with an actual area more than twice the apparent area. 3. Most type II cells have time- and voltage-dependent outward currents carried by potassium ions. Potassium currents activate with a sigmoid time course upon membrane depolarization, and inactivate during maintained depolarization. The average maximum whole-cell K+ conductance was 1.6 nS. 4. Two distinct types of K+-selective channels underlie outward currents in type II cells. Most cells have currents resembling delayed rectifier K+ currents in skeletal muscle, nerve and immune cells. A few cells had a different type of K+ conductance which is more sensitive to block by tetraethylammonium ions, has faster 'tail currents', and activates at more positive potentials. 5. In some experiments, individual type II cells were identified by staining with phosphine, a fluorescent dye which is concentrated in lamellar bodies. Both types of K+ channels were seen in type II cells identified with this dye. 6. Phosphine added to the bathing solution reversibly reduced K+ currents and shifted K+ channel activation to more positive potentials. Excitation of phosphine to fluoresce reduced irreversibly K+ currents in type II cells. The usefulness of phosphine as a means of identifying cells for study is discussed. PMID:2457683

  9. Neuronal cell types and connectivity: lessons from the retina

    PubMed Central

    Seung, H. Sebastian; Sümbül, Uygar

    2014-01-01

    We describe recent progress towards defining neuronal cell types in the mouse retina, and attempt to extract lessons that may be generally useful in the mammalian brain. Achieving a comprehensive catalog of retinal cell types now appears within reach, because researchers have achieved consensus concerning two fundamental challenges. The first is accuracy—defining pure cell types rather than settling for neuronal classes that are mixtures of types. The second is completeness—developing methods guaranteed to eventually identify all cell types, as well as criteria for determining when all types have been found. Case studies illustrate how these two challenges are handled by combining state-of-the-art molecular, anatomical and physiological techniques. Progress is also being made in observing and modeling connectivity between cell types. Scaling up to larger brain regions, such as the cortex, will require not only technical advances but careful consideration of the challenges of accuracy and completeness. PMID:25233310

  10. Transmission and Scanning Electron Microscopy of the Accessory Cells and Chorion During Development of Ciona intestinalis Type B Embryos and the Impact of Their Removal on Cell Morphology.

    PubMed

    Thompson, Helen; Shimeld, Sebastian M

    2015-06-01

    Spawned ascidian oocytes are surrounded by a membrane called the chorion (or vitelline coat) and associated with two populations of maternally-supplied cells. Outside the chorion are follicle cells, which may affect the buoyancy of eggs. Inside the chorion are test cells, which during oogenesis provision the egg and which after fertilisation contribute to the larval tunic. The structure of maternal cells may vary between species. The model ascidian Ciona intestinalis has been recently split into two species, currently named type A and type B. The ultrastructure of extraembryonic cells and structures from type A embryos has been reported. Here we describe the ultrastructure of follicle and test cells from C. intestinalis type B embryos. Test cells are about 5 µm in diameter and line the inside of the chorion of developing embryos in a dense sheet. Follicle cells are large (> 100 µm long) and spike-shaped, with many large vesicles. Terminal electron dense granules are found towards the tips of spikes, adjacent to cytoplasm containing numerous small electron dense bodies connected by filaments. These are probably vesicles containing material for the terminal granules. Removal of maternal structures and cells just after fertilisation, as commonly used in many experiments manipulating C. intestinalis development, has been reported to affect embryonic patterning. We examined the impact of this on embryonic ectoderm cells by scanning electron microscopy. Cells of embryos that developed without maternal structures still developed cilia, but had indistinct cell boundaries and a more flattened appearance than those that developed within the chorion.

  11. Distinguishing epigenetic features of preneoplastic testis tissues adjacent to seminomas and nonseminomas

    PubMed Central

    Skvortsova, Yulia V.; Zinovyeva, Marina V.; Stukacheva, Elena A.; Klimov, Alexey; Tryakin, Alexey A.; Azhikina, Tatyana L.

    2016-01-01

    PIWI pathway proteins are expressed during spermatogenesis where they play a key role in germ cell development. Epigenetic loss of PIWI proteins expression was previously demonstrated in testicular germ cell tumors (TGCTs), implying their involvement in TGCT development. In this work, apart from studying only normal testis and TGCT samples, we also analyzed an intermediate stage, i.e. preneoplastic testis tissues adjacent to TGCTs. Importantly, in this study, we minimized the contribution of patient-to-patient heterogeneity by using matched preneoplastic/TGCT samples. Surprisingly, expression of germ cell marker DDX4 suggests that spermatogenesis is retained in premalignant testis tissues adjacent to nonseminoma, but not those adjacent to seminoma. Moreover, this pattern is followed by expression of PIWI pathway genes, which impacts one of their functions: DNA methylation level over LINE-1 promoters is higher in preneoplastic testis tissues adjacent to nonseminomas than those adjacent to seminomas. This finding might imply distinct routes for development of the two types of TGCTs and could be used as a novel diagnostic marker, possibly, noninvasively. Finally, we studied the role of CpG island methylation in expression of PIWI genes in patient samples and using in vitro experiments in cell line models: a more complex interrelation between DNA methylation and expression of the corresponding genes was revealed. PMID:26843623

  12. Freedom of expression: cell-type-specific gene profiling.

    PubMed

    Otsuki, Leo; Cheetham, Seth W; Brand, Andrea H

    2014-01-01

    Cell fate and behavior are results of differential gene regulation, making techniques to profile gene expression in specific cell types highly desirable. Many methods now enable investigation at the DNA, RNA and protein level. This review introduces the most recent and popular techniques, and discusses key issues influencing the choice between these such as ease, cost and applicability of information gained. Interdisciplinary collaborations will no doubt contribute further advances, including not just in single cell type but single-cell expression profiling.

  13. [Dendritic cells and interaction with other cell types. Immune tolerance].

    PubMed

    Guerder, S

    2001-07-01

    T cell tolerance to self antigen is mainly established in the thymus were self-reactive T cells are deleted. Interdigitating dendritic cells and medulary epithelial cells are directly involved in the deletion process. Some self-reactive T cells escape, however this thymic censorship and enter the peripheral pool of naive T cells. Multiple mechanisms are also at play in the periphery to control this potentially armfull T cells, this include deletion and immune deviation.

  14. Improved fuel-cell-type hydrogen sensor

    NASA Technical Reports Server (NTRS)

    Rudek, F. P.; Rutkowski, M. D.

    1968-01-01

    Modified hydrogen sensor replaces oxygen cathode with a cathode consisting of a sealed paste of gold hydroxide and a pure gold current collector. The net reaction which occurs during cell operation is the reduction of the gold hydroxide to gold and water, with a half-cell potential of 1.4 volts.

  15. Pathogenic memory type Th2 cells in allergic inflammation.

    PubMed

    Endo, Yusuke; Hirahara, Kiyoshi; Yagi, Ryoji; Tumes, Damon J; Nakayama, Toshinori

    2014-02-01

    Immunological memory is a hallmark of adaptive immunity. Memory CD4 T helper (Th) cells are central to acquired immunity, and vaccines for infectious diseases are developed based on this concept. However, memory Th cells also play a critical role in the pathogenesis of various chronic inflammatory diseases, including asthma. We refer to these populations as 'pathogenic memory Th cells.' Here, we review recent developments highlighting the functions and characteristics of several pathogenic memory type Th2 cell subsets in allergic inflammation. Also discussed are the similarities and differences between pathogenic memory Th2 cells and recently identified type 2 innate lymphoid cells (ILC2), focusing on cytokine production and phenotypic profiles.

  16. The immunoregulatory role of type I and type II NKT cells in cancer and other diseases

    PubMed Central

    Terabe, Masaki; Berzofsky, Jay A.

    2014-01-01

    NKT cells are CD1d-restricted T cells that recognize lipid antigens. They also have been shown to play critical roles in the regulation of immune responses. In the immune responses against tumors, two subsets of NKT cells, type I and type II, play opposing roles and cross-regulate each other. As members of both the innate and adaptive immune systems, which form a network of multiple components, they also interact with other immune components. Here we discuss the function of NKT cells in tumor immunity and their interaction with other regulatory cells, especially CD4+CD25+Foxp3+ regulatory T cells. PMID:24384834

  17. Pancreas β cell regeneration and type 1 diabetes (Review)

    PubMed Central

    WU, JINXIAO; YANG, XIYAN; CHEN, BIN; XU, XIUPING

    2015-01-01

    Diabetes mellitus, which may cause hyperglycemia and a number of complications, mostly results from a deficiency of β cell mass (type 1 diabetes) or a limitation of β cell function (type 2 diabetes). Currently, enhancing β cell regeneration and increasing cell proliferation have not only been described in experimental diabetes models, but have also been proven to improve outcomes for patients with diabetes. Therefore, understanding the mechanisms controlling the development and regeneration of β cells in the human pancreas may be helpful for the treatment of β cell-deficient disease. In this review, we first introduce the various cell types in the adult pancreas and thereby clarify their functions and origins. Then, the known mechanisms of β cell development and expansion in the normal human pancreas are described. The potential mechanisms of β cell regeneration, including β cell self-replication, neogenesis from non-β cell precursors and transdifferentiation from α cells, are discussed in the next part. Finally, the ability of the pancreas to regenerate mature β cells is explored in pathological conditions, including type 1 diabetes, chronic pancreatitis and persistent hyperinsulinemic hypoglycemia of infancy. PMID:25667609

  18. High Cell Surface Death Receptor Expression Determines Type I Versus Type II Signaling*

    PubMed Central

    Meng, Xue Wei; Peterson, Kevin L.; Dai, Haiming; Schneider, Paula; Lee, Sun-Hee; Zhang, Jin-San; Koenig, Alexander; Bronk, Steve; Billadeau, Daniel D.; Gores, Gregory J.; Kaufmann, Scott H.

    2011-01-01

    Previous studies have suggested that there are two signaling pathways leading from ligation of the Fas receptor to induction of apoptosis. Type I signaling involves Fas ligand-induced recruitment of large amounts of FADD (FAS-associated death domain protein) and procaspase 8, leading to direct activation of caspase 3, whereas type II signaling involves Bid-mediated mitochondrial perturbation to amplify a more modest death receptor-initiated signal. The biochemical basis for this dichotomy has previously been unclear. Here we show that type I cells have a longer half-life for Fas message and express higher amounts of cell surface Fas, explaining the increased recruitment of FADD and subsequent signaling. Moreover, we demonstrate that cells with type II Fas signaling (Jurkat or HCT-15) can signal through a type I pathway upon forced receptor overexpression and that shRNA-mediated Fas down-regulation converts cells with type I signaling (A498) to type II signaling. Importantly, the same cells can exhibit type I signaling for Fas and type II signaling for TRAIL (TNF-α-related apoptosis-inducing ligand), indicating that the choice of signaling pathway is related to the specific receptor, not some other cellular feature. Additional experiments revealed that up-regulation of cell surface death receptor 5 levels by treatment with 7-ethyl-10-hydroxy-camptothecin converted TRAIL signaling in HCT116 cells from type II to type I. Collectively, these results suggest that the type I/type II dichotomy reflects differences in cell surface death receptor expression. PMID:21865165

  19. Characterization of cloned cells from an immortalized fetal pulmonary type II cell line

    SciTech Connect

    Henderson, R.F.; Waide, J.J.; Lechner, J.F.

    1995-12-01

    A cultured cell line that maintained expression of pulmonary type II cell markers of differentiation would be advantageous to generate a large number of homogenous cells in which to study the biochemical functions of type II cells. Type II epithelial cells are the source of pulmonary surfactant and a cell of origin for pulmonary adenomas. Last year our laboratory reported the induction of expression of two phenotypic markers of pulmonary type II cells (alkaline phosphatase activity and surfactant lipid synthesis) in cultured fetal rat lung epithelial (FRLE) cells, a spontaneously immortalized cell line of fetal rat lung type II cell origin. Subsequently, the induction of the ability to synthesize surfactant lipid became difficult to repeat. We hypothesized that the cell line was heterogenuous and some cells were more like type II cells than others. The purpose of this study was to test this hypothesis and to obtain a cultured cell line with type II cell phenotypic markers by cloning several FRLE cells and characterizing them for phenotypic markers of type II cells (alkaline phosphatase activity and presence of surfactant lipids). Thirty cloned cell lines were analyzed for induced alkaline phosphatase activity (on x-axis) and for percent of phospholipids that were disaturated (i.e., surfactant).

  20. Identification and quantitation of morphological cell types in electrophoretically separated human embryonic kidney cell cultures

    NASA Technical Reports Server (NTRS)

    Williams, K. B.; Kunze, M. E.; Todd, P. W.

    1985-01-01

    Four major cell types were identified by phase microscopy in early passage human embryonic kidney cell cultures. They are small and large epithelioid, domed, and fenestrated cells. Fibroblasts are also present in some explants. The percent of each cell type changes with passage number as any given culture grows. As a general rule, the fraction of small epithelioid cells increases, while the fraction of fenestrated cells, always small, decreases further. When fibroblasts are present, they always increase in percentage of the total cell population. Electrophoretic separation of early passage cells showed that the domed cells have the highest electrophoretic mobility, fibroblasts have an intermediate high mobility, small epithelioid cells have a low mobility, broadly distributed, and fenestrated cells have the lowest mobility. All cell types were broadly distributed among electrophoretic subfractions, which were never pure but only enriched with respect to a given cell type.

  1. Asymmetrical cell division in Blepharisma japonicum: difference between daughter cells in mating-type expression.

    PubMed

    Miyake, A; Harumoto, T

    1990-09-01

    In cell division of high-frequency-selfers in the ciliate Blepharisma japonicum, daughter cells are different in mating-type expression. The anterior daughter cell is mating type I. The posterior daughter cell is mating type II at first and then changes to mating type I after about 24 h. The anteroposterior polarity of predivision cells appears to correlate with the asymmetrical cell division. This work introduces a unicellular organism about the size of microscopic metazoa as a model system for the study of asymmetrical cell division, which is particularly important in developmental processes.

  2. Neuronal Neuregulin 1 type III directs Schwann cell migration

    PubMed Central

    Perlin, Julie R.; Lush, Mark E.; Stephens, W. Zac; Piotrowski, Tatjana; Talbot, William S.

    2011-01-01

    During peripheral nerve development, each segment of a myelinated axon is matched with a single Schwann cell. Tight regulation of Schwann cell movement, proliferation and differentiation is essential to ensure that these glial cells properly associate with axons. ErbB receptors are required for Schwann cell migration, but the operative ligand and its mechanism of action have remained unknown. We demonstrate that zebrafish Neuregulin 1 (Nrg1) type III, which signals through ErbB receptors, controls Schwann cell migration in addition to its previously known roles in proliferation and myelination. Chimera analyses indicate that ErbB receptors are required in all migrating Schwann cells, and that Nrg1 type III is required in neurons for migration. Surprisingly, expression of the ligand in a few axons is sufficient to induce migration along a chimeric nerve constituted largely of nrg1 type III mutant axons. These studies also reveal a mechanism that allows Schwann cells to fasciculate axons regardless of nrg1 type III expression. Time-lapse imaging of transgenic embryos demonstrated that misexpression of human NRG1 type III results in ectopic Schwann cell migration, allowing them to aberrantly enter the central nervous system. These results demonstrate that Nrg1 type III is an essential signal that controls Schwann cell migration to ensure that these glia are present in the correct numbers and positions in developing nerves. PMID:21965611

  3. Heparin- and sulfatide-binding peptides from the type I repeats of human thrombospondin promote melanoma cell adhesion.

    PubMed Central

    Guo, N H; Krutzsch, H C; Nègre, E; Vogel, T; Blake, D A; Roberts, D D

    1992-01-01

    Peptides from the three type I repeats of human endothelial cell thrombospondin, containing the consensus sequence-Trp-Ser-Xaa-Trp-, bind to sulfated glycoconjugates including heparin and sulfatide. The peptides are potent inhibitors for the binding of thrombospondin, laminin, or apolipoprotein E to these ligands. The thrombospondin peptides that inhibit heparin binding, but not adjacent peptides from the thrombospondin sequence containing the previously identified adhesive motif Val-Thr-Cys-Gly, promote melanoma cell adhesion when immobilized on plastic. Melanoma cell adhesion to the immobilized peptides is inhibited by soluble recombinant heparin-binding fragment of thrombospondin. The peptides also inhibit heparin-dependent binding of thrombospondin or laminin to human melanoma cells. The active peptides lack any previously identified heparin-binding consensus sequences and most do not contain any basic amino acids. Studies with homologous peptides showed that the tryptophan residues are required for binding. Adjacent basic residues in the second type I repeat enhance binding to heparin but not to sulfatide. Thus the type I peptides of thrombospondin define a distinct class of heparin-binding peptides. Images PMID:1557410

  4. Novel Cell Types, Neurosecretory Cells and Body Plan of the Early-Diverging Metazoan, Trichoplax adhaerens

    PubMed Central

    Smith, Carolyn L.; Varoqueaux, Frédérique; Kittelmann, Maike; Azzam, Rita N.; Cooper, Benjamin; Winters, Christine A.; Eitel, Michael; Fasshauer, Dirk; Reese, Thomas S.

    2014-01-01

    Summary Background Trichoplax adhaerens is the best-known member of the Phylum Placozoa, one of the earliest-diverging metazoan phyla. It is a small disk-shaped animal that glides on surfaces in warm oceans to feed on algae. Prior anatomical studies of Trichoplax revealed that it has a simple three-layered organization with four somatic cell types. Results We reinvestigate the cellular organization of Trichoplax using advanced freezing and microscopy techniques to identify localize and count cells. Six somatic cell types are deployed in stereotyped positions. A thick ventral plate, comprising the majority of the cells, includes ciliated epithelial cells, newly identified lipophil cells packed with large lipid granules, and gland cells. Lipophils project deep into the interior where they alternate with regularly spaced fiber cells whose branches contact all other cell types, including cells of the dorsal and ventral epithelium. Crystal cells, each containing a birefringent crystal, are arrayed around the rim. Gland cells express several proteins typical of neurosecretory cells, and a subset of them, around the rim, also expresses an FMRFamide-like neuropeptide. Conclusions Structural analysis of Trichoplax with significantly improved techniques provides an advance in understanding its cell types and their distributions. We find two previously undetected cell types, lipohil and crystal cells, and an organized body plan in which different cell types are arranged in distinct patterns. The composition of gland cells suggests that they are neurosecretory cells and could control locomotor and feeding behavior. PMID:24954051

  5. Barrier Epithelial Cells and the Control of Type 2 Immunity.

    PubMed

    Hammad, Hamida; Lambrecht, Bart N

    2015-07-21

    Type-2-cell-mediated immunity, rich in eosinophils, basophils, mast cells, CD4(+) T helper 2 (Th2) cells, and type 2 innate lymphoid cells (ILC2s), protects the host from helminth infection but also drives chronic allergic diseases like asthma and atopic dermatitis. Barrier epithelial cells (ECs) represent the very first line of defense and express pattern recognition receptors to recognize type-2-cell-mediated immune insults like proteolytic allergens or helminths. These ECs mount a prototypical response made up of chemokines, innate cytokines such as interleukin-1 (IL-1), IL-25, IL-33, and thymic stromal lymphopoietin (TSLP), as well as the alarmins uric acid, ATP, HMGB1, and S100 proteins. These signals program dendritic cells (DCs) to mount Th2-cell-mediated immunity and in so doing boost ILC2, basophil, and mast cell function. Here we review the general mechanisms of how different stimuli trigger type-2-cell-mediated immunity at mucosal barriers and how this leads to protection or disease.

  6. Gelsolin Induces Colorectal Tumor Cell Invasion via Modulation of the Urokinase-Type Plasminogen Activator Cascade

    PubMed Central

    Zhuo, Jingli; Tan, Ee Hong; Yan, Benedict; Tochhawng, Lalchhandami; Jayapal, Manikandan; Koh, Shiuan; Tay, Hwee Kee; Maciver, Sutherland K.; Hooi, Shing Chuan; Salto-Tellez, Manuel; Kumar, Alan Prem; Goh, Yaw Chong; Lim, Yaw Chyn; Yap, Celestial T.

    2012-01-01

    Gelsolin is a cytoskeletal protein which participates in actin filament dynamics and promotes cell motility and plasticity. Although initially regarded as a tumor suppressor, gelsolin expression in certain tumors correlates with poor prognosis and therapy-resistance. In vitro, gelsolin has anti-apoptotic and pro-migratory functions and is critical for invasion of some types of tumor cells. We found that gelsolin was highly expressed at tumor borders infiltrating into adjacent liver tissues, as examined by immunohistochemistry. Although gelsolin contributes to lamellipodia formation in migrating cells, the mechanisms by which it induces tumor invasion are unclear. Gelsolin’s influence on the invasive activity of colorectal cancer cells was investigated using overexpression and small interfering RNA knockdown. We show that gelsolin is required for invasion of colorectal cancer cells through matrigel. Microarray analysis and quantitative PCR indicate that gelsolin overexpression induces the upregulation of invasion-promoting genes in colorectal cancer cells, including the matrix-degrading urokinase-type plasminogen activator (uPA). Conversely, gelsolin knockdown reduces uPA levels, as well as uPA secretion. The enhanced invasiveness of gelsolin-overexpressing cells was attenuated by treatment with function-blocking antibodies to either uPA or its receptor uPAR, indicating that uPA/uPAR activity is crucial for gelsolin-dependent invasion. In summary, our data reveals novel functions of gelsolin in colorectal tumor cell invasion through its modulation of the uPA/uPAR cascade, with potentially important roles in colorectal tumor dissemination to metastatic sites. PMID:22927998

  7. Human laryngeal ganglia contain both sympathetic and parasympathetic cell types.

    PubMed

    Ibanez, Marta; Valderrama-Canales, Francisco J; Maranillo, Eva; Vazquez, Teresa; Pascual-Font, Arán; McHanwell, Stephen; Sanudo, Jose

    2010-09-01

    The presence of ganglia associated with the laryngeal nerves is well documented. In man, these ganglia have been less well studied than in other species and, in particular, the cell types within these ganglia are less well characterized. Using a panel of antibodies to a variety of markers found in the paraganglion cells of other species, we were able to show the existence of at least two populations of cells within human laryngeal paraganglia. One population contained chromogranin and tyrosine hydroxylase representing a neurosecretory population possibly secreting dopamine. A second population of choline acetyltransferase positive cells would appear to have a putative parasympathetic function. Further work is needed to characterize these cell populations more fully before it will be possible to assign functions to these cell types but our results are consistent with the postulated functions of these ganglia as chemoreceptors, neurosecretory cells, and regulators of laryngeal mucus secretion.

  8. Transcripts of the MHM region on the chicken Z chromosome accumulate as non-coding RNA in the nucleus of female cells adjacent to the DMRT1 locus.

    PubMed

    Teranishi, M; Shimada, Y; Hori, T; Nakabayashi, O; Kikuchi, T; Macleod, T; Pym, R; Sheldon, B; Solovei, I; Macgregor, H; Mizuno, S

    2001-01-01

    The male hypermethylated (MHM) region, located near the middle of the short arm of the Z chromosome of chickens, consists of approximately 210 tandem repeats of a BamHI 2.2-kb sequence unit. Cytosines of the CpG dinucleotides of this region are extensively methylated on the two Z chromosomes in the male but much less methylated on the single Z chromosome in the female. The state of methylation of the MHM region is established after fertilization by about the 1-day embryonic stage. The MHM region is transcribed only in the female from the particular strand into heterogeneous, high molecular-mass, non-coding RNA, which is accumulated at the site of transcription, adjacent to the DMRT1 locus, in the nucleus. The transcriptional silence of the MHM region in the male is most likely caused by the CpG methylation, since treatment of the male embryonic fibroblasts with 5-azacytidine results in hypo-methylation and active transcription of this region. In ZZW triploid chickens, MHM regions are hypomethylated and transcribed on the two Z chromosomes, whereas MHM regions are hypermethylated and transcriptionally inactive on the three Z chromosomes in ZZZ triploid chickens, suggesting a possible role of the W chromosome on the state of the MHM region. PMID:11321370

  9. Murine Coronavirus Cell Type Dependent Interaction with the Type I Interferon Response

    PubMed Central

    Rose, Kristine M.; Weiss, Susan R.

    2009-01-01

    Coronaviruses infect many species of animal including humans, causing acute and chronic diseases of many organ systems. Murine coronavirus, mouse hepatitis virus (MHV) infection of the mouse, provides animal models for the study of central nervous system disease, including encephalitis and demyelinating diseases such as Multiple Sclerosis and for hepatitis. While there are many studies of the adaptive immune response to MHV, there has until recently been scant information on the type I interferon (IFN) response to MHV. The relationship between MHV and the IFN-α/β response is paradoxical. While the type I IFN response is a crucial aspect of host defense against MHV in its natural host, there is little if any induction of IFN following infection of mouse fibroblast cell lines in vitro. Furthermore, MHV is relatively resistant to the antiviral effects of IFN-α/β in mouse fibroblast cell lines and in human 293T cells. MHV can, under some circumstances, compromise the antiviral effects of IFN signaling. The nucleocapsid protein as well as the nsp1 and nsp3 proteins of MHV has been reported to have IFN antagonist activity. However, in primary cell types such as plasmacytoid dendritic cells (pDC) and macrophages, IFN is induced by MHV infection and an antiviral state is established. Other primary cell types such as neurons, astrocytes and hepatocytes fail to produce IFN following infection and, in vivo, likely depend on IFN produced by pDCs and macrophages for protection from MHV. Thus MHV induction of IFN-α/β and the ability to induce an antiviral state in response to interferon is extremely cell type dependent. IFN induced protection from MHV pathogenesis likely requires the orchestrated activities of several cell types, however, the cell types involved in limiting MHV replication may be different in the liver and in the immune privileged CNS. PMID:20221421

  10. Type I natural killer T cells: naturally born for fighting

    PubMed Central

    Tan, Jin-quan; Xiao, Wei; Wang, Lan; He, Yu-ling

    2010-01-01

    Type І natural killer T cells (NKT cells), a subset of CD1d-restricted T cells with invariant Vαβ TCR, are characterized by prompt production of large amounts of Th1 and/or Th2 cytokines upon primary stimulation through the TCR complex. The rapid release of cytokines implies that type І NKT cells may play a critical role in modulating the upcoming immune responses, such as anti-tumor response, protection against infection, and autoimmunity. As a bridge between innate and adaptive immunity, type І NKT cells differentiate and mature upon stimulations to achieve and maintain a homeostasis. Orchestrating with other arms of adaptive immunity, type І NKT cells show strong cytotoxic effects in response to various tumors in a direct and/or indirect manner(s). This review will focus primarily on type І NKT cell development, homeostasis, and effector functions, especially in anti-tumor immunity, and followed by their potential applications in treatment of cancers. PMID:20694020

  11. Distinct type I and type II toxin-antitoxin modules control Salmonella lifestyle inside eukaryotic cells

    PubMed Central

    Lobato-Márquez, Damián; Moreno-Córdoba, Inmaculada; Figueroa, Virginia; Díaz-Orejas, Ramón; García-del Portillo, Francisco

    2015-01-01

    Toxin-antitoxin (TA) modules contribute to the generation of non-growing cells in response to stress. These modules abound in bacterial pathogens although the bases for this profusion remain largely unknown. Using the intracellular bacterial pathogen Salmonella enterica serovar Typhimurium as a model, here we show that a selected group of TA modules impact bacterial fitness inside eukaryotic cells. We characterized in this pathogen twenty-seven TA modules, including type I and type II TA modules encoding antisense RNA and proteinaceous antitoxins, respectively. Proteomic and gene expression analyses revealed that the pathogen produces numerous toxins of TA modules inside eukaryotic cells. Among these, the toxins HokST, LdrAST, and TisBST, encoded by type I TA modules and T4ST and VapC2ST, encoded by type II TA modules, promote bacterial survival inside fibroblasts. In contrast, only VapC2ST shows that positive effect in bacterial fitness when the pathogen infects epithelial cells. These results illustrate how S. Typhimurium uses distinct type I and type II TA modules to regulate its intracellular lifestyle in varied host cell types. This function specialization might explain why the number of TA modules increased in intracellular bacterial pathogens. PMID:25792384

  12. Distinct type I and type II toxin-antitoxin modules control Salmonella lifestyle inside eukaryotic cells.

    PubMed

    Lobato-Márquez, Damián; Moreno-Córdoba, Inmaculada; Figueroa, Virginia; Díaz-Orejas, Ramón; García-del Portillo, Francisco

    2015-01-01

    Toxin-antitoxin (TA) modules contribute to the generation of non-growing cells in response to stress. These modules abound in bacterial pathogens although the bases for this profusion remain largely unknown. Using the intracellular bacterial pathogen Salmonella enterica serovar Typhimurium as a model, here we show that a selected group of TA modules impact bacterial fitness inside eukaryotic cells. We characterized in this pathogen twenty-seven TA modules, including type I and type II TA modules encoding antisense RNA and proteinaceous antitoxins, respectively. Proteomic and gene expression analyses revealed that the pathogen produces numerous toxins of TA modules inside eukaryotic cells. Among these, the toxins HokST, LdrAST, and TisBST, encoded by type I TA modules and T4ST and VapC2ST, encoded by type II TA modules, promote bacterial survival inside fibroblasts. In contrast, only VapC2ST shows that positive effect in bacterial fitness when the pathogen infects epithelial cells. These results illustrate how S. Typhimurium uses distinct type I and type II TA modules to regulate its intracellular lifestyle in varied host cell types. This function specialization might explain why the number of TA modules increased in intracellular bacterial pathogens.

  13. Nitric oxide alters metabolism in isolated alveolar type II cells.

    PubMed

    Miles, P R; Bowman, L; Huffman, L

    1996-07-01

    Alveolar type II cells may be exposed to nitric oxide (.NO) from external sources, and these cells can also generate .NO. Therefore we studied the effects of altering .NO levels on various type II cell metabolic processes. Incubation of cells with the .NO generator, S-nitroso-N-acetylpenicillamine (SNAP; 1 mM), leads to reductions of 60-70% in the synthesis of disaturated phosphatidylcholines (DSPC) and cell ATP levels. Cellular oxygen consumption, an indirect measure of cell ATP synthesis, is also reduced by SNAP. There is no direct effect of SNAP on lung mitochondrial ATP synthesis, suggesting that .NO does not directly inhibit this process. On the other hand, incubation of cells with NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase (NOS), the enzyme responsible for .NO synthesis, results in increases in DSPC synthesis, cell ATP content, and cellular oxygen consumption. The L-NAME effects are reversed by addition of L-arginine, the substrate for NOS. Production of .NO by type II cells is inhibited by L-NAME, a better inhibitor of constitutive NOS (cNOS) than inducible NOS (iNOS), and is reduced in the absence of external calcium. Aminoguanidine, a specific inhibitor of iNOS, has no effect on cell ATP content or on .NO production. These results indicate that alveolar type II cell lipid and energy metabolism can be affected by .NO and suggest that there may be cNOS activity in these cells. PMID:8760128

  14. Regulation of immune cell homeostasis by type I interferons.

    PubMed

    Mattei, Fabrizio; Schiavoni, Giovanna; Tough, David F

    2010-08-01

    Although initially identified and best characterized for their role in innate antiviral defence, type I interferons (IFN-I) are also known to have an important impact on the adaptive immune response. In part, this is linked to another long-recognised property of IFN-I, namely their ability to modify cellular proliferation and survival. Here, we review the influence of IFN-I on immune cell homeostasis, focusing on their effects on T cells and antigen-presenting cells. PMID:20627800

  15. Generation of diverse neural cell types through direct conversion

    PubMed Central

    Petersen, Gayle F; Strappe, Padraig M

    2016-01-01

    A characteristic of neurological disorders is the loss of critical populations of cells that the body is unable to replace, thus there has been much interest in identifying methods of generating clinically relevant numbers of cells to replace those that have been damaged or lost. The process of neural direct conversion, in which cells of one lineage are converted into cells of a neural lineage without first inducing pluripotency, shows great potential, with evidence of the generation of a range of functional neural cell types both in vitro and in vivo, through viral and non-viral delivery of exogenous factors, as well as chemical induction methods. Induced neural cells have been proposed as an attractive alternative to neural cells derived from embryonic or induced pluripotent stem cells, with prospective roles in the investigation of neurological disorders, including neurodegenerative disease modelling, drug screening, and cellular replacement for regenerative medicine applications, however further investigations into improving the efficacy and safety of these methods need to be performed before neural direct conversion becomes a clinically viable option. In this review, we describe the generation of diverse neural cell types via direct conversion of somatic cells, with comparison against stem cell-based approaches, as well as discussion of their potential research and clinical applications. PMID:26981169

  16. Cell-type-resolved quantitative proteomics of murine liver.

    PubMed

    Azimifar, S Babak; Nagaraj, Nagarjuna; Cox, Juergen; Mann, Matthias

    2014-12-01

    Mass spectrometry (MS)-based proteomics provides a powerful approach to globally investigate the biological function of individual cell types in mammalian organs. Here, we applied this technology to the in-depth analysis of purified hepatic cell types from mouse. We quantified 11,520 proteins, making this the most comprehensive proteomic resource of any organ to date. Global protein copy number determination demonstrated that a large proportion of the hepatocyte proteome is dedicated to fatty acid and xenobiotic metabolism. We identified as-yet-unknown components of the TGF-β signaling pathway and extracellular matrix in hepatic stellate cells, uncovering their regulative role in liver physiology. Moreover, our high-resolution proteomic data set enabled us to compare the distinct functional roles of hepatic cell types in cholesterol flux, cellular trafficking, and growth factor receptor signaling. This study provides a comprehensive resource for liver biology and biomedicine.

  17. Development of a laser capture microscope-based single-cell-type proteomics tool for studying proteomes of individual cell layers of plant roots

    PubMed Central

    Zhu, Yingde; Li, Hui; Bhatti, Sarabjit; Zhou, Suping; Yang, Yong; Fish, Tara; Thannhauser, Theodore W

    2016-01-01

    Single-cell-type proteomics provides the capability to revealing the genomic and proteomics information at cell-level resolution. However, the methodology for this type of research has not been well-developed. This paper reports developing a workflow of laser capture microdissection (LCM) followed by gel-liquid chromatography-tandem mass spectrometry (GeLC-MS/MS)-based proteomics analysis for the identification of proteomes contained in individual cell layers of tomato roots. Thin-sections (~10-μm thick, 10 sections per root tip) were prepared for root tips of tomato germinating seedlings. Epidermal and cortical cells (5000–7000 cells per tissue type) were isolated under a LCM microscope. Proteins were isolated and then separated by SDS–polyacrylamide gel electrophoresis followed by in-gel-tryptic digestion. The MS and MS/MS spectra generated using nanoLC-MS/MS analysis of the tryptic peptides were searched against ITAG2.4 tomato protein database to identify proteins contained in each single-cell-type sample. Based on the biological functions, proteins with proven functions in root hair development were identified in epidermal cells but not in the cortical cells. Several of these proteins were found in Al-treated roots only. The results demonstrated that the cell-type-specific proteome is relevant for tissue-specific functions in tomato roots. Increasing the coverage of proteomes and reducing the inevitable cross-contamination from adjacent cell layers, in both vertical and cross directions when cells are isolated from slides prepared using intact root tips, are the major challenges using the technology in proteomics analysis of plant roots. PMID:27280026

  18. Development of a laser capture microscope-based single-cell-type proteomics tool for studying proteomes of individual cell layers of plant roots.

    PubMed

    Zhu, Yingde; Li, Hui; Bhatti, Sarabjit; Zhou, Suping; Yang, Yong; Fish, Tara; Thannhauser, Theodore W

    2016-01-01

    Single-cell-type proteomics provides the capability to revealing the genomic and proteomics information at cell-level resolution. However, the methodology for this type of research has not been well-developed. This paper reports developing a workflow of laser capture microdissection (LCM) followed by gel-liquid chromatography-tandem mass spectrometry (GeLC-MS/MS)-based proteomics analysis for the identification of proteomes contained in individual cell layers of tomato roots. Thin-sections (~10-μm thick, 10 sections per root tip) were prepared for root tips of tomato germinating seedlings. Epidermal and cortical cells (5000-7000 cells per tissue type) were isolated under a LCM microscope. Proteins were isolated and then separated by SDS-polyacrylamide gel electrophoresis followed by in-gel-tryptic digestion. The MS and MS/MS spectra generated using nanoLC-MS/MS analysis of the tryptic peptides were searched against ITAG2.4 tomato protein database to identify proteins contained in each single-cell-type sample. Based on the biological functions, proteins with proven functions in root hair development were identified in epidermal cells but not in the cortical cells. Several of these proteins were found in Al-treated roots only. The results demonstrated that the cell-type-specific proteome is relevant for tissue-specific functions in tomato roots. Increasing the coverage of proteomes and reducing the inevitable cross-contamination from adjacent cell layers, in both vertical and cross directions when cells are isolated from slides prepared using intact root tips, are the major challenges using the technology in proteomics analysis of plant roots.

  19. Development of a laser capture microscope-based single-cell-type proteomics tool for studying proteomes of individual cell layers of plant roots.

    PubMed

    Zhu, Yingde; Li, Hui; Bhatti, Sarabjit; Zhou, Suping; Yang, Yong; Fish, Tara; Thannhauser, Theodore W

    2016-01-01

    Single-cell-type proteomics provides the capability to revealing the genomic and proteomics information at cell-level resolution. However, the methodology for this type of research has not been well-developed. This paper reports developing a workflow of laser capture microdissection (LCM) followed by gel-liquid chromatography-tandem mass spectrometry (GeLC-MS/MS)-based proteomics analysis for the identification of proteomes contained in individual cell layers of tomato roots. Thin-sections (~10-μm thick, 10 sections per root tip) were prepared for root tips of tomato germinating seedlings. Epidermal and cortical cells (5000-7000 cells per tissue type) were isolated under a LCM microscope. Proteins were isolated and then separated by SDS-polyacrylamide gel electrophoresis followed by in-gel-tryptic digestion. The MS and MS/MS spectra generated using nanoLC-MS/MS analysis of the tryptic peptides were searched against ITAG2.4 tomato protein database to identify proteins contained in each single-cell-type sample. Based on the biological functions, proteins with proven functions in root hair development were identified in epidermal cells but not in the cortical cells. Several of these proteins were found in Al-treated roots only. The results demonstrated that the cell-type-specific proteome is relevant for tissue-specific functions in tomato roots. Increasing the coverage of proteomes and reducing the inevitable cross-contamination from adjacent cell layers, in both vertical and cross directions when cells are isolated from slides prepared using intact root tips, are the major challenges using the technology in proteomics analysis of plant roots. PMID:27280026

  20. Type 1 interferons contribute to the clearance of senescent cell.

    PubMed

    Katlinskaya, Yuliya V; Carbone, Christopher J; Yu, Qiujing; Fuchs, Serge Y

    2015-01-01

    The major known function of cytokines that belong to type I interferons (IFN, including IFNα and IFNβ) is to mount the defense against viruses. This function also protects the genetic information of host cells from alterations in the genome elicited by some of these viruses. Furthermore, recent studies demonstrated that IFN also restrict proliferation of damaged cells by inducing cell senescence. Here we investigated the subsequent role of IFN in elimination of the senescent cells. Our studies demonstrate that endogenous IFN produced by already senescent cells contribute to increased expression of the natural killer (NK) receptor ligands, including MIC-A and ULBP2. Furthermore, neutralization of endogenous IFN or genetic ablation of its receptor chain IFNAR1 compromises the recognition of senescent cells and their clearance in vitro and in vivo. We discuss the role of IFN in protecting the multi-cellular host from accumulation of damaged senescent cells and potential significance of this mechanism in human cancers.

  1. Porcine circovirus type 2 displays pluripotency in cell targeting

    SciTech Connect

    Steiner, Esther Balmelli, Carole Herrmann, Brigitte; Summerfield, Artur; McCullough, Kenneth

    2008-09-01

    Porcine circovirus type 2 (PCV2) is the causative agent of a multifactorial disease associated with immunocompromisation and co-infections. In vivo, viral DNA and antigens are found in monocytic, epithelial and endothelial cells. Of these, PCV2 replication has only been studied in monocytic cells, in which little or no replication was identified. Accordingly, PCV2 infection was studied in the endothelial cell line PEDSV.15, aortic endothelial cells, gut epithelial cells, fibrocytes and dendritic cells (DC). In all cells except DC PCV2 replication was detectable, with an increase in the levels of capsid and replicase protein. Variations in endocytic activity, virus binding and uptake did not relate to the replication efficiency in a particular cell. Furthermore, replication did not correlate to cell proliferation, although a close association of viral proteins with chromatin in dividing cells was observed. No alteration in the division rate of PCV2-infected cultures was measurable, relating to replicase expression in only a small minority of the cells. In conclusion, the broad cell targeting of PCV2 offers an explanation for its widespread tissue distribution.

  2. Alveolar epithelial type II cell: defender of the alveolus revisited

    PubMed Central

    Fehrenbach, Heinz

    2001-01-01

    In 1977, Mason and Williams developed the concept of the alveolar epithelial type II (AE2) cell as a defender of the alveolus. It is well known that AE2 cells synthesise, secrete, and recycle all components of the surfactant that regulates alveolar surface tension in mammalian lungs. AE2 cells influence extracellular surfactant transformation by regulating, for example, pH and [Ca2+] of the hypophase. AE2 cells play various roles in alveolar fluid balance, coagulation/fibrinolysis, and host defence. AE2 cells proliferate, differentiate into AE1 cells, and remove apoptotic AE2 cells by phagocytosis, thus contributing to epithelial repair. AE2 cells may act as immunoregulatory cells. AE2 cells interact with resident and mobile cells, either directly by membrane contact or indirectly via cytokines/growth factors and their receptors, thus representing an integrative unit within the alveolus. Although most data support the concept, the controversy about the character of hyperplastic AE2 cells, reported to synthesise profibrotic factors, proscribes drawing a definite conclusion today. PMID:11686863

  3. Is Transforming Stem Cells to Pancreatic Beta Cells Still the Holy Grail for Type 2 Diabetes?

    PubMed

    Kahraman, Sevim; Okawa, Erin R; Kulkarni, Rohit N

    2016-08-01

    Diabetes is a progressive disease affecting millions of people worldwide. There are several medications and treatment options to improve the life quality of people with diabetes. One of the strategies for the treatment of diabetes could be the use of human pluripotent stem cells or induced pluripotent stem cells. The recent advances in differentiation of stem cells into insulin-secreting beta-like cells in vitro make the transplantation of the stem cell-derived beta-like cells an attractive approach for treatment of type 1 and type 2 diabetes. While stem cell-derived beta-like cells provide an unlimited cell source for beta cell replacement therapies, these cells can also be used as a platform for drug screening or modeling diseases.

  4. Regulatory T Cells in Type 1 Autoimmune Pancreatitis

    PubMed Central

    Uchida, Kazushige; Kusuda, Takeo; Koyabu, Masanori; Miyoshi, Hideaki; Fukata, Norimasa; Sumimoto, Kimi; Fukui, Yuri; Sakaguchi, Yutaku; Ikeura, Tsukasa; Shimatani, Masaaki; Fukui, Toshiro; Matsushita, Mitsunobu; Takaoka, Makoto; Nishio, Akiyoshi; Okazaki, Kazuichi

    2012-01-01

    Autoimmune pancreatitis (AIP) is a newly recognized pancreatic disorder. Recently, International Consensus Diagnostic Criteria for AIP (ICDC) was published. In this ICDC, AIP was classified into Type 1 and Type 2. Patients with Type 1 AIP have several immunologic and histologic abnormalities specific to the disease, including increased levels of serum IgG4 and storiform fibrosis with infiltration of lymphocytes and IgG4-positive plasmacytes in the involved organs. Among the involved organs showing extrapancreatic lesions, the bile duct is the most common, exhibiting sclerosing cholangitis (IgG4-SC). However, the role of IgG4 is unclear. Recently, it has been reported that regulatory T cells (Tregs) are involved in both the development of various autoimmune diseases and the shift of B cells toward IgG4, producing plasmacytes. Our study showed that Tregs were increased in the pancreas with Type 1 AIP and IgG4-SC compared with control. In the patients with Type 1 AIP and IgG4-SC, the numbers of infiltrated Tregs were significantly positively correlated with IgG4-positive plasma cells. In Type 1 AIP, inducible costimulatory molecule (ICOS)+ and IL-10+ Tregs significantly increased compared with control groups. Our data suggest that increased quantities of ICOS+ Tregs may influence IgG4 production via IL-10 in Type 1 AIP. PMID:22536257

  5. Permian to Triassic I to S-type magmatic switch in the northeast Sierra Nevada de Santa Marta and adjacent regions, Colombian Caribbean: Tectonic setting and implications within Pangea paleogeography

    NASA Astrophysics Data System (ADS)

    Cardona, A.; Valencia, V.; Garzón, A.; Montes, C.; Ojeda, G.; Ruiz, J.; Weber, M.

    2010-10-01

    The Late Paleozoic to Triassic tectonics of northwestern South America have major implications for the understanding of Laurentia-Gondwana interactions that formed Pangea, and the origin of several tectonostratigraphic terranes dispersed by the break-up of this supercontinent during the formation of the Caribbean. Two mylonitic and orthogneissic granitoid suites have been recognized in the northeastern segment of the Sierra Nevada de Santa Marta, the lower Magdalena basin and the Guajira Serranias, within the Caribbean region of Colombia. For the Santa Marta region U/Pb LAM-ICP-MS analysis yielded zircon crystallization ages of 288.1 ± 4.5 Ma, 276.5 ± 5,1 Ma and 264.9 ± 4.0 Ma, related to the magmatic intrusion. Geochemical and modal variations show a compositional spectrum between diorite and granite, whereas LREE enrichment, Ti and Nb anomalies and geochemical discrimination suggest that this granitoid suite was formed within a magmatic arc setting. Inherited zircons suggest that this Early Permian plutonism was formed with the participation of Neoproterozoic and Grenvillian basement proximal to the South American continent. Evidence of a superimposed Early Triassic (ca. 250 Ma) deformational event in Santa Marta, together with a well defined S-type magmatism in the basement rocks from the adjacent lower Magdalena Valley and Guajira Peninsula regions are related to a major shift in the regional tectonic evolution. It's envisioned that this event records either terrane accretion or strong plate coupling during the final stages of Pangea agglutination. Connections with the main Alleghanian-Ouachitan Pangean orogen are precluded due to their timing differences. The plutons temporally and compositionally correlate with an arc found in the northern Andes and Mexican Gondwana terranes, and represent a broader magmatic event formed at the proto-Pacific margin, outside the nucleus of the Laurentia-Gondwana Alleghanian-Oachitan orogens. Evidence of lower temperature

  6. Adjacent Segment Pathology after Anterior Cervical Fusion.

    PubMed

    Chung, Jae Yoon; Park, Jong-Beom; Seo, Hyoung-Yeon; Kim, Sung Kyu

    2016-06-01

    Anterior cervical fusion has become a standard of care for numerous pathologic conditions of the cervical spine. However, subsequent development of clinically significant disc disease at levels adjacent to fused discs is a serious long-term complication of this procedure. As more patients live longer after surgery, it is foreseeable that adjacent segment pathology (ASP) will develop in increasing numbers of patients. Also, ASP has been studied more intensively with the recent popularity of motion preservation technologies like total disc arthroplasty. The true nature and scope of ASP remains poorly understood. The etiology of ASP is most likely multifactorial. Various factors including altered biomechanical stresses, surgical disruption of soft tissue and the natural history of cervical disc disease contribute to the development of ASP. General factors associated with disc degeneration including gender, age, smoking and sports may play a role in the development of ASP. Postoperative sagittal alignment and type of surgery are also considered potential causes of ASP. Therefore, a spine surgeon must be particularly careful to avoid unnecessary disruption of the musculoligamentous structures, reduced risk of direct injury to the disc during dissection and maintain a safe margin between the plate edge and adjacent vertebrae during anterior cervical fusion.

  7. Adjacent Segment Pathology after Anterior Cervical Fusion

    PubMed Central

    Chung, Jae Yoon; Park, Jong-Beom; Seo, Hyoung-Yeon

    2016-01-01

    Anterior cervical fusion has become a standard of care for numerous pathologic conditions of the cervical spine. However, subsequent development of clinically significant disc disease at levels adjacent to fused discs is a serious long-term complication of this procedure. As more patients live longer after surgery, it is foreseeable that adjacent segment pathology (ASP) will develop in increasing numbers of patients. Also, ASP has been studied more intensively with the recent popularity of motion preservation technologies like total disc arthroplasty. The true nature and scope of ASP remains poorly understood. The etiology of ASP is most likely multifactorial. Various factors including altered biomechanical stresses, surgical disruption of soft tissue and the natural history of cervical disc disease contribute to the development of ASP. General factors associated with disc degeneration including gender, age, smoking and sports may play a role in the development of ASP. Postoperative sagittal alignment and type of surgery are also considered potential causes of ASP. Therefore, a spine surgeon must be particularly careful to avoid unnecessary disruption of the musculoligamentous structures, reduced risk of direct injury to the disc during dissection and maintain a safe margin between the plate edge and adjacent vertebrae during anterior cervical fusion. PMID:27340541

  8. Targeting memory T cells in type 1 diabetes.

    PubMed

    Ehlers, Mario R; Rigby, Mark R

    2015-11-01

    Type 1 diabetes (T1D) is a chronic autoimmune disease that leads to progressive destruction of pancreatic beta cells. Compared to healthy controls, a characteristic feature of patients with T1D is the presence of self-reactive T cells with a memory phenotype. These autoreactive memory T cells in both the CD4(+) and CD8(+) compartments are likely to be long-lived, strongly responsive to antigenic stimulation with less dependence on costimulation for activation and clonal expansion, and comparatively resistant to suppression by regulatory T cells (Tregs) or downregulation by immune-modulating agents. Persistence of autoreactive memory T cells likely contributes to the difficulty in preventing disease progression in new-onset T1D and maintaining allogeneic islet transplants by regular immunosuppressive regimens. The majority of immune interventions that have demonstrated some success in preserving beta cell function in the new-onset period have been shown to deplete or modulate memory T cells. Based on these and other considerations, preservation of residual beta cells early after diagnosis or restoration of beta cell mass by use of stem cell or transplantation technology will require a successful strategy to control the autoreactive memory T cell compartment, which could include depletion, inhibition of homeostatic cytokines, induction of hyporesponsiveness, or a combination of these approaches.

  9. Cell-Type Specific Four-Component Hydrogel

    PubMed Central

    Aberle, Timo; Franke, Katrin; Rist, Elke; Benz, Karin; Schlosshauer, Burkhard

    2014-01-01

    In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin) to generate a blend (technical term: quattroGel), an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appropriate for general cell adhesion, and restricted diffusion. Cell proliferation of endothelial cells, chondrocytes and fibroblasts was essentially unaffected. In contrast, on quattroGels neither endothelial cells formed vascular tubes nor did primary neurons extend neurites in significant amounts. Only chondrocytes differentiated properly as judged by collagen isoform expression. The biophysical quattroGel characteristics appeared to leave distinct cell processes such as mitosis unaffected and favored differentiation of sessile cells, but hampered differentiation of migratory cells. This cell-type selectivity is of interest e.g. during articular cartilage or invertebral disc repair, where pathological innervation and angiogenesis represent adverse events in tissue engineering. PMID:24475174

  10. Cell-type-dependent action potentials and voltage-gated currents in mouse fungiform taste buds.

    PubMed

    Kimura, Kenji; Ohtubo, Yoshitaka; Tateno, Katsumi; Takeuchi, Keita; Kumazawa, Takashi; Yoshii, Kiyonori

    2014-01-01

    Taste receptor cells fire action potentials in response to taste substances to trigger non-exocytotic neurotransmitter release in type II cells and exocytotic release in type III cells. We investigated possible differences between these action potentials fired by mouse taste receptor cells using in situ whole-cell recordings, and subsequently we identified their cell types immunologically with cell-type markers, an IP3 receptor (IP3 R3) for type II cells and a SNARE protein (SNAP-25) for type III cells. Cells not immunoreactive to these antibodies were examined as non-IRCs. Here, we show that type II cells and type III cells fire action potentials using different ionic mechanisms, and that non-IRCs also fire action potentials with either of the ionic mechanisms. The width of action potentials was significantly narrower and their afterhyperpolarization was deeper in type III cells than in type II cells. Na(+) current density was similar in type II cells and type III cells, but it was significantly smaller in non-IRCs than in the others. Although outwardly rectifying current density was similar between type II cells and type III cells, tetraethylammonium (TEA) preferentially suppressed the density in type III cells and the majority of non-IRCs. Our mathematical model revealed that the shape of action potentials depended on the ratio of TEA-sensitive current density and TEA-insensitive current one. The action potentials of type II cells and type III cells under physiological conditions are discussed.

  11. Adult human adipose tissue contains several types of multipotent cells.

    PubMed

    Tallone, Tiziano; Realini, Claudio; Böhmler, Andreas; Kornfeld, Christopher; Vassalli, Giuseppe; Moccetti, Tiziano; Bardelli, Silvana; Soldati, Gianni

    2011-04-01

    Multipotent mesenchymal stromal cells (MSCs) are a type of adult stem cells that can be easily isolated from various tissues and expanded in vitro. Many reports on their pluripotency and possible clinical applications have raised hopes and interest in MSCs. In an attempt to unify the terminology and the criteria to label a cell as MSC, in 2006 the International Society for Cellular Therapy (ISCT) proposed a standard set of rules to define the identity of these cells. However, MSCs are still extracted from different tissues, by diverse isolation protocols, are cultured and expanded in different media and conditions. All these variables may have profound effects on the selection of cell types and the composition of heterogeneous subpopulations, on the selective expansion of specific cell populations with totally different potentials and ergo, on the long-term fate of the cells upon in vitro culture. Therefore, specific molecular and cellular markers that identify MSCs subsets as well as standardization of expansion protocols for these cells are urgently needed. Here, we briefly discuss new useful markers and recent data supporting the rapidly emerging concept that many different types of progenitor cells are found in close association with blood vessels. This knowledge may promote the necessary technical improvements required to reduce variability and promote higher efficacy and safety when isolating and expanding these cells for therapeutic use. In the light of the discussed data, particularly the identification of new markers, and advances in the understanding of fundamental MSC biology, we also suggest a revision of the 2006 ISCT criteria.

  12. Type II cochlear ganglion cells in the chinchilla.

    PubMed

    Ruggero, M A; Santi, P A; Rich, N C

    1982-12-01

    In order to ascertain whether Type II cochlear ganglion cells project to the brain, we have studied the retrograde transport of horseradish peroxidase (HRP) from the cochlear nucleus to the spiral ganglion of the chinchilla. In this animal there exist two types of ganglion neurons, which closely correspond to those previously described in guinea pigs, cats and rats. As in the guinea pig, the majority population (Type I) consists of relatively large, myelinated neurons. The minority population (Type II, 10% of the total population) consists of small, mostly unmyelinated cells, with filamentous cytoplasm and finely grained nuclear chromatin. Type II neurons tend to be clustered toward the peripheral side of Rosenthal's canal, often in close proximity to the intraganglionic spiral bundle. By 24 h after injections of HRP into the cochlear nucleus, incubation of the cochlear ganglion in diaminobenzidine/H2O2 reveals abundant HRP label in both Type I and Type II neurons. Type II neurons, however, tend to be labelled less intensely than Type I neurons. Control experiments, consisting of spillage of HRP solution over the cochlear nucleus, were carried out to determine how much HRP might be picked up by neurons after HRP diffusion. Comparison of cochleae from injected animals and from the control animals suggests that most of the label that was found in ganglion neurons after cochlear nucleus injections represents axonally transported HRP. We conclude, at least tentatively, that Type II neurons project to the brain. The fact that less label is found in Type II neurons that in Type I neurons suggests that the former have thinner axons and/or finer terminals in the cochlear nucleus. PMID:6185462

  13. MicroRNA profiling of diverse endothelial cell types

    PubMed Central

    2011-01-01

    Background MicroRNAs are ~22-nt long regulatory RNAs that serve as critical modulators of post-transcriptional gene regulation. The diversity of miRNAs in endothelial cells (ECs) and the relationship of this diversity to epithelial and hematologic cells is unknown. We investigated the baseline miRNA signature of human ECs cultured from the aorta (HAEC), coronary artery (HCEC), umbilical vein (HUVEC), pulmonary artery (HPAEC), pulmonary microvasculature (HPMVEC), dermal microvasculature (HDMVEC), and brain microvasculature (HBMVEC) to understand the diversity of miRNA expression in ECs. Results We identified 166 expressed miRNAs, of which 3 miRNAs (miR-99b, miR-20b and let-7b) differed significantly between EC types and predicted EC clustering. We confirmed the significance of these miRNAs by RT-PCR analysis and in a second data set by Sylamer analysis. We found wide diversity of miRNAs between endothelial, epithelial and hematologic cells with 99 miRNAs shared across cell types and 31 miRNAs unique to ECs. We show polycistronic miRNA chromosomal clusters have common expression levels within a given cell type. Conclusions EC miRNA expression levels are generally consistent across EC types. Three microRNAs were variable within the dataset indicating potential regulatory changes that could impact on EC phenotypic differences. MiRNA expression in endothelial, epithelial and hematologic cells differentiate these cell types. This data establishes a valuable resource characterizing the diverse miRNA signature of ECs. PMID:22047531

  14. Tuft cells, taste-chemosensory cells, orchestrate parasite type 2 immunity in the gut.

    PubMed

    Howitt, Michael R; Lavoie, Sydney; Michaud, Monia; Blum, Arthur M; Tran, Sara V; Weinstock, Joel V; Gallini, Carey Ann; Redding, Kevin; Margolskee, Robert F; Osborne, Lisa C; Artis, David; Garrett, Wendy S

    2016-03-18

    The intestinal epithelium forms an essential barrier between a host and its microbiota. Protozoa and helminths are members of the gut microbiota of mammals, including humans, yet the many ways that gut epithelial cells orchestrate responses to these eukaryotes remain unclear. Here we show that tuft cells, which are taste-chemosensory epithelial cells, accumulate during parasite colonization and infection. Disruption of chemosensory signaling through the loss of TRMP5 abrogates the expansion of tuft cells, goblet cells, eosinophils, and type 2 innate lymphoid cells during parasite colonization. Tuft cells are the primary source of the parasite-induced cytokine interleukin-25, which indirectly induces tuft cell expansion by promoting interleukin-13 production by innate lymphoid cells. Our results identify intestinal tuft cells as critical sentinels in the gut epithelium that promote type 2 immunity in response to intestinal parasites. PMID:26847546

  15. Tuft cells, taste-chemosensory cells, orchestrate parasite type 2 immunity in the gut.

    PubMed

    Howitt, Michael R; Lavoie, Sydney; Michaud, Monia; Blum, Arthur M; Tran, Sara V; Weinstock, Joel V; Gallini, Carey Ann; Redding, Kevin; Margolskee, Robert F; Osborne, Lisa C; Artis, David; Garrett, Wendy S

    2016-03-18

    The intestinal epithelium forms an essential barrier between a host and its microbiota. Protozoa and helminths are members of the gut microbiota of mammals, including humans, yet the many ways that gut epithelial cells orchestrate responses to these eukaryotes remain unclear. Here we show that tuft cells, which are taste-chemosensory epithelial cells, accumulate during parasite colonization and infection. Disruption of chemosensory signaling through the loss of TRMP5 abrogates the expansion of tuft cells, goblet cells, eosinophils, and type 2 innate lymphoid cells during parasite colonization. Tuft cells are the primary source of the parasite-induced cytokine interleukin-25, which indirectly induces tuft cell expansion by promoting interleukin-13 production by innate lymphoid cells. Our results identify intestinal tuft cells as critical sentinels in the gut epithelium that promote type 2 immunity in response to intestinal parasites.

  16. Lasing within Live Cells Containing Intracellular Optical Microresonators for Barcode-Type Cell Tagging and Tracking.

    PubMed

    Schubert, Marcel; Steude, Anja; Liehm, Philipp; Kronenberg, Nils M; Karl, Markus; Campbell, Elaine C; Powis, Simon J; Gather, Malte C

    2015-08-12

    We report on a laser that is fully embedded within a single live cell. By harnessing natural endocytosis of the cell, we introduce a fluorescent whispering gallery mode (WGM) microresonator into the cell cytoplasm. On pumping with nanojoule light pulses, green laser emission is generated inside the cells. Our approach can be applied to different cell types, and cells with microresonators remain viable for weeks under standard conditions. The characteristics of the lasing spectrum provide each cell with a barcode-type label which enables uniquely identifying and tracking individual migrating cells. Self-sustained lasing from cells paves the way to new forms of cell tracking, intracellular sensing, and adaptive imaging. PMID:26186167

  17. Identifying Essential Cell Types and Circuits in Autism Spectrum Disorders

    PubMed Central

    Maloney, Susan E.; Rieger, Michael A.; Dougherty, Joseph D.

    2014-01-01

    Autism spectrum disorder (ASD) is highly genetic in its etiology, with potentially hundreds of genes contributing to risk. Despite this heterogeneity, these disparate genetic lesions may result in the disruption of a limited number of key cell types or circuits –information which could be leveraged for the design of therapeutic interventions. While hypotheses for cellular disruptions can be identified by postmortem anatomical analysis and expression studies of ASD risk genes, testing these hypotheses requires the use of animal models. In this review, we explore the existing evidence supporting the contribution of different cell types to ASD, specifically focusing on rodent studies disrupting serotonergic, GABAergic, cerebellar and striatal cell types, with particular attention to studies of the sufficiency of specific cellular disruptions to generate ASD-related behavioral abnormalities. This evidence suggests multiple cellular routes can create features of the disorder, though it is currently unclear if these cell types converge on a final common circuit. We hope that in the future, systematic studies of cellular sufficiency and genetic interaction will help to classify patients into groups by type of cellular disruptions which suggest tractable therapeutic targets. PMID:24290383

  18. Chronic lymphocytic leukemia cells with allelic deletions at 13q14 commonly have one intact RB1 gene: Evidence for a role of an adjacent locus

    SciTech Connect

    Leu, Y.; Grander, D.; Linder, S.; Einhorn, S.; Soederhall, S. ); Szekely, L. ); Juliusson, G.; Gahrton, G. )

    1993-09-15

    The authors have previously shown that 30% of patients with B-cell chronic lymphocytic leukemia (B-CLL) have hemizygous deletions of the retinoblastoma (RB1) gene at 13q14. RB1 gene deletions may thus participate in malignant transformation of B-CLL, but is it also possible that a neighboring gene on 13q is the relevant one. To answer this question the remaining RB1 allele of eight clones with hemizygous deletions was studied by reverse transcription-polymerase chain reaction (RT-PCR), single-strand conformation polymorphism (SSCP) analysis, and immunofluorescense techniques. Cells from 10 patients without RB1 gene deletions were also studied by these methods. Lack of RB1 mRNA and RB protein expression was seen in leukemia cells from one of the patients. All other cases were found to be normal with regard to immunofluorescense, RT-PCR, and SSCP analysis, indicating at least one functional RB1 allele and supporting the importance of another gene in the 13q14 deletions. The authors then performed extended Southern blot analysis of the 13q region, using probes for 10 different loci. In 14 of 31 CLL clones (45%), deletions of a region telomeric to the RB1 gene (D13S25) were observed. In 4 of the cases the deletions were homozygous. Hemizygous deletions of the RB1 gene were observed in 11 of these patients and in one of the patients without D13S25 deletions. These data thus indicate that a gene(s) telomeric to RB1 is involved in the malignant transformation of CLL clones and that deletions of this region are a common event in this disease. 20 refs., 3 figs., 3 tabs.

  19. Erythroid differentiation of human induced pluripotent stem cells is independent of donor cell type of origin

    PubMed Central

    Dorn, Isabel; Klich, Katharina; Arauzo-Bravo, Marcos J.; Radstaak, Martina; Santourlidis, Simeon; Ghanjati, Foued; Radke, Teja F.; Psathaki, Olympia E.; Hargus, Gunnar; Kramer, Jan; Einhaus, Martin; Kim, Jeong Beom; Kögler, Gesine; Wernet, Peter; Schöler, Hans R.; Schlenke, Peter; Zaehres, Holm

    2015-01-01

    Epigenetic memory in induced pluripotent stem cells, which is related to the somatic cell type of origin of the stem cells, might lead to variations in the differentiation capacities of the pluripotent stem cells. In this context, induced pluripotent stem cells from human CD34+ hematopoietic stem cells might be more suitable for hematopoietic differentiation than the commonly used fibroblast-derived induced pluripotent stem cells. To investigate the influence of an epigenetic memory on the ex vivo expansion of induced pluripotent stem cells into erythroid cells, we compared induced pluripotent stem cells from human neural stem cells and human cord blood-derived CD34+ hematopoietic stem cells and evaluated their potential for differentiation into hematopoietic progenitor and mature red blood cells. Although genome-wide DNA methylation profiling at all promoter regions demonstrates that the epigenetic memory of induced pluripotent stem cells is influenced by the somatic cell type of origin of the stem cells, we found a similar hematopoietic induction potential and erythroid differentiation pattern of induced pluripotent stem cells of different somatic cell origin. All human induced pluripotent stem cell lines showed terminal maturation into normoblasts and enucleated reticulocytes, producing predominantly fetal hemoglobin. Differences were only observed in the growth rate of erythroid cells, which was slightly higher in the induced pluripotent stem cells derived from CD34+ hematopoietic stem cells. More detailed methylation analysis of the hematopoietic and erythroid promoters identified similar CpG methylation levels in the induced pluripotent stem cell lines derived from CD34+ cells and those derived from neural stem cells, which confirms their comparable erythroid differentiation potential. PMID:25326431

  20. Type I collagen gel protects murine fibrosarcoma L929 cells from TNFα-induced cell death

    SciTech Connect

    Wang, Hong-Ju; He, Wen-Qi; Chen, Ling; Liu, Wei-Wei; Xu, Qian; Xia, Ming-Yu; Hayashi, Toshihiko; Fujisaki, Hitomi; Hattori, Shunji; Tashiro, Shin-ichi; Onodera, Satoshi; Ikejima, Takashi

    2015-02-20

    Murine fibrosarcoma L929 cells have been used to test efficacy of proinflammatory cytokine TNFα. In the present study, we reported on protective effect of type I collagen gel used as L929 cell culture. L929 cell grew and proliferated well on collagen gel. However, the L929 cells exhibited cobblestone-like morphology which was much different from the spread fusiform shape when cultured on conventional cell dishes as well as the cells tended to aggregate. On conventional cell culture dishes, the cells treated with TNFα became round in shape and eventually died in a necroptotic manner. The cells cultured on collagen gel, however, were completely unaffected. TNFα treatment was reported to induce autophagy in L929 cells on the plastic dish, and therefore we investigated the effect of collagen gel on induction of autophagy. The results indicated that autophagy induced by TNFα treatment was much reduced when the cells were cultured on collagen gel. In conclusion, type I collagen gel protected L929 cell from TNFα-induced cell death. - Highlights: • Collagen gel culture changed the morphology of L929 cells. • L929 cell cultured on collagen gel were resistant to TNFα-induced cell death. • Collagen gel culture inhibited TNFα-induced autophagy in L929 cells.

  1. Flow and transport within a coastal aquifer adjacent to a stratified water body

    NASA Astrophysics Data System (ADS)

    Oz, Imri; Yechieli, Yoseph; Eyal, Shalev; Gavrieli, Ittai; Gvirtzman, Haim

    2016-04-01

    The existence of a freshwater-saltwater interface and the circulation flow of saltwater beneath the interface is a well-known phenomenon found at coastal aquifers. This flow is a natural phenomenon that occurs due to density differences between fresh groundwater and the saltwater body. The goals of this research are to use analytical, numerical, and physical models in order to examine the configuration of the freshwater-saltwater interface and the density-driven flow patterns within a coastal aquifer adjacent to long-term stratified saltwater bodies (e.g. meromictic lake). Such hydrological systems are unique, as they consist of three different water types: the regional fresh groundwater, and low and high salinity brines forming the upper and lower water layers of the stratified water body, respectively. This research also aims to examine the influence of such stratification on hydrogeological processes within the coastal aquifer. The coastal aquifer adjacent to the Dead Sea, under its possible future meromictic conditions, serves as an ideal example to examine these processes. The results show that adjacent to a stratified saltwater body three interfaces between three different water bodies are formed, and that a complex flow system, controlled by the density differences, is created, where three circulation cells are developed. These results are significantly different from the classic circulation cell that is found adjacent to non-stratified water bodies (lakes or oceans). In order to obtain a more generalized insight into the groundwater behavior adjacent to a stratified water body, we used the numerical model to perform sensitivity analysis. The hydrological system was found be sensitive to three dimensionless parameters: dimensionless density (i.e. the relative density of the three water bodies'); dimensionless thickness (i.e. the ratio between the relative thickness of the upper layer and the whole thickness of the lake); and dimensionless flux. The results

  2. Harnessing immune cells to enhance β-cell mass in type 1 diabetes.

    PubMed

    Dirice, Ercument; Kulkarni, Rohit N

    2016-01-01

    Type 1 diabetes is characterized by early β-cell loss leading to insulin dependence in virtually all patients with the disease in order to maintain glucose homeostasis. Most studies over the past few decades have focused on limiting the autoimmune attack on the β cells. However, emerging data from patients with long-standing diabetes who continue to harbor functional insulin-producing cells in their diseased pancreas have prompted scientists to examine whether proliferation of existing β cells can be enhanced to promote better glycemic control. In support of this concept, several studies indicate that mononuclear cells that infiltrate the islets have the capacity to trigger proliferation of islet cells including β cells. These observations indicate the exciting possibility of identifying those mononuclear cell types and their soluble factors and harnessing their ability to promote β-cell growth concomitant with autoimmune therapy to prevent the onset and/or halt the progression of the disease. PMID:26755809

  3. Receptosecretory nature of type III cells in the taste bud.

    PubMed

    Yoshie, Sumio

    2009-01-01

    Type III cells in taste buds form chemical synapses with intragemmal afferent nerve fibers and are characterized by the presence of membrane-bound vesicles in the cytoplasm. Although the vesicles differ in shape and size among species, they are primarily categorized into small clear (40 nm in diameter) and large dense-cored (90-200 nm) types. As such vesicles tend to be closely juxtaposed to the synaptic membrane of the cells, it is reasonable to consider that the vesicles include transmitter(s) towards the gustatory nerve. In the guinea-pig taste bud, stimulation with various taste substances (sucrose, sodium chloride, quinine hydrochloride, or monosodium L-glutamate) causes ultrastructural alterations of the type III cells. At the synapse, the presynaptic plasma membrane often displays invaginations of 90 nm in a mean diameter towards the cytoplasm, which indicates the dense-cored vesicles opening into the synaptic cleft by means of exocytosis. The vesicles are also exocytosed at the non-synaptic region into the intercellular space. These findings strongly suggest that the transmitters presumably contained in the vesicles are released to conduct the excitement of the type III cells to the nerves and also to exert their paracrine effects upon the surroundings, such as the Ebner's salivary gland, acting as local hormones. PMID:20224182

  4. The Type III Secretion Translocation Pore Senses Host Cell Contact

    PubMed Central

    Armentrout, Erin I.; Rietsch, Arne

    2016-01-01

    Type III secretion systems (T3SS) are nano-syringes used by a wide range of Gram-negative pathogens to promote infection by directly injecting effector proteins into targeted host cells. Translocation of effectors is triggered by host-cell contact and requires assembly of a pore in the host-cell plasma membrane, which consists of two translocator proteins. Our understanding of the translocation pore, how it is assembled in the host cell membrane and its precise role in effector translocation, is extremely limited. Here we use a genetic technique to identify protein-protein contacts between pore-forming translocator proteins, as well as the T3SS needle-tip, that are critical for translocon function. The data help establish the orientation of the translocator proteins in the host cell membrane. Analysis of translocon function in mutants that break these contacts demonstrates that an interaction between the pore-forming translocator PopD and the needle-tip is required for sensing host cell contact. Moreover, tethering PopD at a dimer interface also specifically prevents host-cell sensing, arguing that the translocation pore is actively involved in detecting host cell contact. The work presented here therefore establishes a signal transduction pathway for sensing host cell contact that is initiated by a conformational change in the translocation pore, and is subsequently transmitted to the base of the apparatus via a specific contact between the pore and the T3SS needle-tip. PMID:27022930

  5. The Enhancer of Split Complex and Adjacent Genes in the 96f Region of Drosophila Melanogaster Are Required for Segregation of Neural and Epidermal Progenitor Cells

    PubMed Central

    Schrons, H.; Knust, E.; Campos-Ortega, J. A.

    1992-01-01

    The Enhancer of split complex [E(spl)-C] of Drosophila melanogaster is located in the 96F region of the third chromosome and comprises at least seven structurally related genes, HLH-mδ, HLH-mγ, HLH-mβ, HLH-m3, HLH-m5, HLH-m7 and E(spl). The functions of these genes are required during early neurogenesis to give neuroectodermal cells access to the epidermal pathway of development. Another gene in the 96F region, namely groucho, is also required for this process. However, groucho is not structurally related to, and appears to act independently of, the genes of the E(spl)-C; the possibility is discussed that groucho acts upstream to the E(spl)-C genes. Indirect evidence suggests that a neighboring transcription unit (m4) may also take part in the process. Of all these genes, only gro is essential; m4 is a dispensable gene, the deletion of which does not produce detectable morphogenetic abnormalities, and the genes of the E(spl)-C are to some extent redundant and can partially substitute for each other. This redundancy is probably due to the fact that the seven genes of the E(spl)-C encode highly conserved putative DNA-binding proteins of the bHLH family. The genes of the complex are interspersed among other genes which appear to be unrelated to the neuroepidermal lineage dichotomy. PMID:1427039

  6. Type II secretory phospholipase A2 binds to ischemic flip-flopped cardiomyocytes and subsequently induces cell death.

    PubMed

    Nijmeijer, R; Willemsen, M; Meijer, C J L M; Visser, C A; Verheijen, R H; Gottlieb, R A; Hack, C E; Niessen, H W M

    2003-11-01

    Type II secretory phospholipase A2 (sPLA2) is a cardiovascular risk factor. We recently found depositions of sPLA2 in the necrotic center of infarcted human myocardium and normally appearing cardiomyocytes adjacent to the border zone. The consequences of binding of sPLA2 to ischemic cardiomyocytes are not known. To explore a potential effect of sPLA2 on ischemic cardiomyocytes at a cellular level we used an in vitro model. The cardiomyocyte cell line H9c2 or adult cardiomyocytes were isolated from rabbits that were incubated with sPLA2 in the presence of metabolic inhibitors to mimic ischemia-reperfusion conditions. Cell viability was established with the use of annexin V and propidium iodide or 7-aminoactinomycin D. Metabolic inhibition induced an increase of the number of flip-flopped cells, including a population that did not stain with propidium iodide and that was caspase-3 negative. sPLA2 bound to the flip-flopped cells, including those negative for caspase-3. sPLA2 binding induced cell death in these latter cells. In addition, sPLA2 potentiated the binding of C-reactive protein (CRP) to these cells. We conclude that by binding to flip-flopped cardiomyocytes, including those that are caspase-3 negative and presumably reversibly injured, sPLA2 may induce cell death and tag these cells with CRP.

  7. A Stromal Cell Niche for Human and Mouse Type 3 Innate Lymphoid Cells.

    PubMed

    Hoorweg, Kerim; Narang, Priyanka; Li, Zhi; Thuery, Anne; Papazian, Natalie; Withers, David R; Coles, Mark C; Cupedo, Tom

    2015-11-01

    Adaptive immunity critically depends on the functional compartmentalization of secondary lymphoid organs. Mesenchymal stromal cells create and maintain specialized niches that support survival, activation, and expansion of T and B cells, and integrated analysis of lymphocytes and their niche has been instrumental in understanding adaptive immunity. Lymphoid organs are also home to type 3 innate lymphoid cells (ILC3), innate effector cells essential for barrier immunity. However, a specialized stromal niche for ILC3 has not been identified. A novel lineage-tracing approach now identifies a subset of murine fetal lymphoid tissue organizer cells that gives rise exclusively to adult marginal reticular cells. Moreover, both cell types are conserved from mice to humans and colocalize with ILC3 in secondary lymphoid tissues throughout life. In sum, we provide evidence that fetal stromal organizers give rise to adult marginal reticular cells and form a dedicated stromal niche for innate ILC3 in adaptive lymphoid organs.

  8. Identifying Cell Types from Spatially Referenced Single-Cell Expression Datasets

    PubMed Central

    Achim, Kaia; Richardson, Sylvia; Azizi, Lamiae; Marioni, John

    2014-01-01

    Complex tissues, such as the brain, are composed of multiple different cell types, each of which have distinct and important roles, for example in neural function. Moreover, it has recently been appreciated that the cells that make up these sub-cell types themselves harbour significant cell-to-cell heterogeneity, in particular at the level of gene expression. The ability to study this heterogeneity has been revolutionised by advances in experimental technology, such as Wholemount in Situ Hybridizations (WiSH) and single-cell RNA-sequencing. Consequently, it is now possible to study gene expression levels in thousands of cells from the same tissue type. After generating such data one of the key goals is to cluster the cells into groups that correspond to both known and putatively novel cell types. Whilst many clustering algorithms exist, they are typically unable to incorporate information about the spatial dependence between cells within the tissue under study. When such information exists it provides important insights that should be directly included in the clustering scheme. To this end we have developed a clustering method that uses a Hidden Markov Random Field (HMRF) model to exploit both quantitative measures of expression and spatial information. To accurately reflect the underlying biology, we extend current HMRF approaches by allowing the degree of spatial coherency to differ between clusters. We demonstrate the utility of our method using simulated data before applying it to cluster single cell gene expression data generated by applying WiSH to study expression patterns in the brain of the marine annelid Platynereis dumereilii. Our approach allows known cell types to be identified as well as revealing new, previously unexplored cell types within the brain of this important model system. PMID:25254363

  9. Inducible human immunodeficiency virus type 1 packaging cell lines.

    PubMed Central

    Yu, H; Rabson, A B; Kaul, M; Ron, Y; Dougherty, J P

    1996-01-01

    Packaging cell lines are important tools for transferring genes into eukaryotic cells. Human immunodeficiency virus type 1 (HIV-1)-based packaging cell lines are difficult to obtain, in part owing to the problem that some HIV-1 proteins are cytotoxic in a variety of cells. To overcome this, we have developed an HIV-1-based packaging cell line which has an inducible expression system. The tetracycline-inducible expression system was utilized to control the expression of the Rev regulatory protein, which in turn controls the expression of the late proteins including Gag, Pol, and Env. Western blotting (immunoblotting) demonstrated that the expression of p24gag and gp120env from the packaging cells peaked on days 6 and 7 postinduction. Reverse transcriptase activity could be detected by day 4 after induction and also peaked on days 6 and 7. Defective vector virus could be propagated, yielding titers as high as 7 x 10(3) CFU/ml, while replication-competent virus was not detectable at any time. Thus, the cell line should enable the transfer of specific genes into CD4+ cells and should be a useful tool for studying the biology of HIV-1. We have also established an inducible HIV-1 Env-expressing cell line which could be used to propagate HIV-1 vectors that require only Env in trans. The env-minus vector virus titer produced from the Env-expressing cells reached 2 x 10(4) CFU/ml. The inducible HIV-1 Env-expressing cell line should be a useful tool for the study of HIV-1 Env as well. PMID:8676479

  10. Glial-cell-derived neuroregulators control type 3 innate lymphoid cells and gut defence.

    PubMed

    Ibiza, Sales; García-Cassani, Bethania; Ribeiro, Hélder; Carvalho, Tânia; Almeida, Luís; Marques, Rute; Misic, Ana M; Bartow-McKenney, Casey; Larson, Denise M; Pavan, William J; Eberl, Gérard; Grice, Elizabeth A; Veiga-Fernandes, Henrique

    2016-07-21

    Group 3 innate lymphoid cells (ILC3) are major regulators of inflammation and infection at mucosal barriers. ILC3 development is thought to be programmed, but how ILC3 perceive, integrate and respond to local environmental signals remains unclear. Here we show that ILC3 in mice sense their environment and control gut defence as part of a glial–ILC3–epithelial cell unit orchestrated by neurotrophic factors. We found that enteric ILC3 express the neuroregulatory receptor RET. ILC3-autonomous Ret ablation led to decreased innate interleukin-22 (IL-22), impaired epithelial reactivity, dysbiosis and increased susceptibility to bowel inflammation and infection. Neurotrophic factors directly controlled innate Il22 downstream of the p38 MAPK/ERK-AKT cascade and STAT3 activation. Notably, ILC3 were adjacent to neurotrophic-factor-expressing glial cells that exhibited stellate-shaped projections into ILC3 aggregates. Glial cells sensed microenvironmental cues in a MYD88-dependent manner to control neurotrophic factors and innate IL-22. Accordingly, glial-intrinsic Myd88 deletion led to impaired production of ILC3-derived IL-22 and a pronounced propensity towards gut inflammation and infection. Our work sheds light on a novel multi-tissue defence unit, revealing that glial cells are central hubs of neuron and innate immune regulation by neurotrophic factor signals. PMID:27409807

  11. Glial-cell-derived neuroregulators control type 3 innate lymphoid cells and gut defence.

    PubMed

    Ibiza, Sales; García-Cassani, Bethania; Ribeiro, Hélder; Carvalho, Tânia; Almeida, Luís; Marques, Rute; Misic, Ana M; Bartow-McKenney, Casey; Larson, Denise M; Pavan, William J; Eberl, Gérard; Grice, Elizabeth A; Veiga-Fernandes, Henrique

    2016-07-21

    Group 3 innate lymphoid cells (ILC3) are major regulators of inflammation and infection at mucosal barriers. ILC3 development is thought to be programmed, but how ILC3 perceive, integrate and respond to local environmental signals remains unclear. Here we show that ILC3 in mice sense their environment and control gut defence as part of a glial–ILC3–epithelial cell unit orchestrated by neurotrophic factors. We found that enteric ILC3 express the neuroregulatory receptor RET. ILC3-autonomous Ret ablation led to decreased innate interleukin-22 (IL-22), impaired epithelial reactivity, dysbiosis and increased susceptibility to bowel inflammation and infection. Neurotrophic factors directly controlled innate Il22 downstream of the p38 MAPK/ERK-AKT cascade and STAT3 activation. Notably, ILC3 were adjacent to neurotrophic-factor-expressing glial cells that exhibited stellate-shaped projections into ILC3 aggregates. Glial cells sensed microenvironmental cues in a MYD88-dependent manner to control neurotrophic factors and innate IL-22. Accordingly, glial-intrinsic Myd88 deletion led to impaired production of ILC3-derived IL-22 and a pronounced propensity towards gut inflammation and infection. Our work sheds light on a novel multi-tissue defence unit, revealing that glial cells are central hubs of neuron and innate immune regulation by neurotrophic factor signals.

  12. Cell type- and brain region-resolved mouse brain proteome.

    PubMed

    Sharma, Kirti; Schmitt, Sebastian; Bergner, Caroline G; Tyanova, Stefka; Kannaiyan, Nirmal; Manrique-Hoyos, Natalia; Kongi, Karina; Cantuti, Ludovico; Hanisch, Uwe-Karsten; Philips, Mari-Anne; Rossner, Moritz J; Mann, Matthias; Simons, Mikael

    2015-12-01

    Brain transcriptome and connectome maps are being generated, but an equivalent effort on the proteome is currently lacking. We performed high-resolution mass spectrometry-based proteomics for in-depth analysis of the mouse brain and its major brain regions and cell types. Comparisons of the 12,934 identified proteins in oligodendrocytes, astrocytes, microglia and cortical neurons with deep sequencing data of the transcriptome indicated deep coverage of the proteome. Cell type-specific proteins defined as tenfold more abundant than average expression represented about a tenth of the proteome, with an overrepresentation of cell surface proteins. To demonstrate the utility of our resource, we focused on this class of proteins and identified Lsamp, an adhesion molecule of the IgLON family, as a negative regulator of myelination. Our findings provide a framework for a system-level understanding of cell-type diversity in the CNS and serves as a rich resource for analyses of brain development and function. PMID:26523646

  13. The statistical geometry of transcriptome divergence in cell-type evolution and cancer.

    PubMed

    Liang, Cong; Forrest, Alistair R R; Wagner, Günter P

    2015-01-01

    In evolution, body plan complexity increases due to an increase in the number of individualized cell types. Yet, there is very little understanding of the mechanisms that produce this form of organismal complexity. One model for the origin of novel cell types is the sister cell-type model. According to this model, each cell type arises together with a sister cell type through specialization from an ancestral cell type. A key prediction of the sister cell-type model is that gene expression profiles of cell types exhibit tree structure. Here we present a statistical model for detecting tree structure in transcriptomic data and apply it to transcriptomes from ENCODE and FANTOM5. We show that transcriptomes of normal cells harbour substantial amounts of hierarchical structure. In contrast, cancer cell lines have less tree structure, suggesting that the emergence of cancer cells follows different principles from that of evolutionary cell-type origination. PMID:25585899

  14. Mesenchymal stem cell-based therapy for type 1 diabetes.

    PubMed

    Wu, Hao; Mahato, Ram I

    2014-03-01

    Diabetes has increasingly become a worldwide health problem, causing huge burden on healthcare system and economy. Type 1 diabetes (T1D), traditionally termed "juvenile diabetes" because of an early onset age, is affecting 5-10% of total diabetic population. Insulin injection, the predominant treatment for T1D, is effective to ameliorate the hyperglycemia but incompetent to relieve the autoimmunity and to regenerate lost islets. Islet transplantation, an experimental treatment for T1D, also suffers from limited supply of human islets and poor immunosuppression. The recent progress in regenerative medicine, especially stem cell therapy, has suggested several novel and potential cures for T1D. Mesenchymal stem cell (MSC) based cell therapy is among one of them. MSCs are a type of adult stem cells residing in bone marrow, adipose tissue, umbilical cord blood, and many other tissues. MSCs, with self-renewal potential and transdifferentiation capability, can be expanded in vitro and directed to various cell lineages with relatively less efforts. MSCs have well-characterized hypoimmunogenicity and immunomodulatory effect. All these features make MSCs attractive for treating T1D. Here, we review the properties of MSCs and some of the recent progress using MSCs as a new therapeutic in the treatment of T1D. We also discuss the strength and limitations of using MSC therapy in human trials.

  15. Type-Specific Cell Line Models for Type-Specific Ovarian Cancer Research

    PubMed Central

    Anglesio, Michael S.; Wiegand, Kimberly C.; Melnyk, Nataliya; Chow, Christine; Salamanca, Clara; Prentice, Leah M.; Senz, Janine; Yang, Winnie; Spillman, Monique A.; Cochrane, Dawn R.; Shumansky, Karey; Shah, Sohrab P.; Kalloger, Steve E.; Huntsman, David G.

    2013-01-01

    Background Ovarian carcinomas consist of at least five distinct diseases: high-grade serous, low-grade serous, clear cell, endometrioid, and mucinous. Biomarker and molecular characterization may represent a more biologically relevant basis for grouping and treating this family of tumors, rather than site of origin. Molecular characteristics have become the new standard for clinical pathology, however development of tailored type-specific therapies is hampered by a failure of basic research to recognize that model systems used to study these diseases must also be stratified. Unrelated model systems do offer value for study of biochemical processes but specific cellular context needs to be applied to assess relevant therapeutic strategies. Methods We have focused on the identification of clear cell carcinoma cell line models. A panel of 32 “ovarian cancer” cell lines has been classified into histotypes using a combination of mutation profiles, IHC mutation-surrogates, and a validated immunohistochemical model. All cell lines were identity verified using STR analysis. Results Many described ovarian clear cell lines have characteristic mutations (including ARID1A and PIK3CA) and an overall molecular/immuno-profile typical of primary tumors. Mutations in TP53 were present in the majority of high-grade serous cell lines. Advanced genomic analysis of bona-fide clear cell carcinoma cell lines also support copy number changes in typical biomarkers such at MET and HNF1B and a lack of any recurrent expressed re-arrangements. Conclusions: As with primary ovarian tumors, mutation status of cancer genes like ARID1A and TP53 and a general immuno-profile serve well for establishing histotype of ovarian cancer cell We describe specific biomarkers and molecular features to re-classify generic “ovarian carcinoma” cell lines into type specific categories. Our data supports the use of prototype clear cell lines, such as TOV21G and JHOC-5, and questions the use of SKOV3 and A

  16. Stem cell therapies for type 1 diabetes mellitus.

    PubMed

    Voltarelli, Júilio C; Couri, Carlos E B; Rodrigues, Maria C; Moraes, Daniela A; Stracieri, Ana-Beatriz P L; Pieroni, Fabiano; Navarro, George; Leal, Angela M O; Simões, Belinda P

    2011-06-01

    The present review discusses the use of autologous hematopoietic stem cell transplantation (HSCT) for the treatment of diabetes mellitus type 1 (DM 1). It has been observed that high dose immunosuppression followed by HSCT shows better results among other immunotherapeutic treatments for the disease as the patients with adequate beta cell reserve achieve insulin independence. However, this response is not maintained and reoccurrence of the disease is major a major challenge to use HSCT in future to prevent or control relapse of DM 1.

  17. Characterization of tissue plasminogen activator binding proteins isolated from endothelial cells and other cell types

    SciTech Connect

    Beebe, D.P.; Wood, L.L.; Moos, M. )

    1990-07-15

    Human tissue plasminogen activator (t-PA) was shown to bind specifically to human osteosarcoma cells (HOS), and human epidermoid carcinoma cells (A-431 cells). Crosslinking studies with DTSSP demonstrated high molecular weight complexes (130,000) between {sup 125}I-t-PA and cell membrane protein on human umbilical vein endothelial cells (HUVEC), HOS, and A-431 cells. A 48-65,000 molecular weight complex was demonstrated after crosslinking t-PA peptide (res. 7-20) to cells. Ligand blotting of cell lysates which had been passed over a t-PA affinity column revealed binding of t-PA to 54,000 and 95,000 molecular weight proteins. Several t-PA binding proteins were identified in immunopurified cell lysates, including tubulin beta chain, plasminogen activator inhibitor type 1 and single chain urokinase.

  18. Immunological applications of stem cells in type 1 diabetes.

    PubMed

    Fiorina, Paolo; Voltarelli, Julio; Zavazava, Nicholas

    2011-12-01

    Current approaches aiming to cure type 1 diabetes (T1D) have made a negligible number of patients insulin-independent. In this review, we revisit the role of stem cell (SC)-based applications in curing T1D. The optimal therapeutic approach for T1D should ideally preserve the remaining β-cells, restore β-cell function, and protect the replaced insulin-producing cells from autoimmunity. SCs possess immunological and regenerative properties that could be harnessed to improve the treatment of T1D; indeed, SCs may reestablish peripheral tolerance toward β-cells through reshaping of the immune response and inhibition of autoreactive T-cell function. Furthermore, SC-derived insulin-producing cells are capable of engrafting and reversing hyperglycemia in mice. Bone marrow mesenchymal SCs display a hypoimmunogenic phenotype as well as a broad range of immunomodulatory capabilities, they have been shown to cure newly diabetic nonobese diabetic (NOD) mice, and they are currently undergoing evaluation in two clinical trials. Cord blood SCs have been shown to facilitate the generation of regulatory T cells, thereby reverting hyperglycemia in NOD mice. T1D patients treated with cord blood SCs also did not show any adverse reaction in the absence of major effects on glycometabolic control. Although hematopoietic SCs rarely revert hyperglycemia in NOD mice, they exhibit profound immunomodulatory properties in humans; newly hyperglycemic T1D patients have been successfully reverted to normoglycemia with autologous nonmyeloablative hematopoietic SC transplantation. Finally, embryonic SCs also offer exciting prospects because they are able to generate glucose-responsive insulin-producing cells. Easy enthusiasm should be mitigated mainly because of the potential oncogenicity of SCs.

  19. Cell therapeutic options in liver diseases: cell types, medical devices and regulatory issues.

    PubMed

    Nussler, Andreas K; Zeilinger, Katrin; Schyschka, Lilianna; Ehnert, Sabrina; Gerlach, Jörg C; Yan, Xueying; Lee, Serene M L; Ilowski, Maren; Thasler, Wolfgang E; Weiss, Thomas S

    2011-05-01

    Although significant progress has been made in the field of orthotopic liver transplantation, cell-based therapies seem to be a promising alternative to whole-organ transplantation. The reasons are manifold but organ shortage is the main cause for this approach. However, many problems such as the question which cell type should be used or which application site is best for transplantation have been raised. In addition, some clinicians have had success by cultivating liver cells in bioreactors for temporary life support. Besides answering the question which cell type, which injection site or even which culture form should be used for liver support recent international harmonization of legal requirements is needed to be addressed by clinicians, scientists and companies dealing with cellular therapies. We here briefly summarize the possible cell types used to partially or temporarily correct liver diseases, the most recent development of bioreactor technology and important regulatory issues.

  20. Molecular comparison of single cell MDA products derived from different cell types.

    PubMed

    Glentis, Stavros; SenGupta, Sioban; Thornhill, Alan; Wang, Rubin; Craft, Ian; Harper, Joyce Catherine

    2009-07-01

    The quality of DNA obtained from single cells for molecular analysis is primarily dependent on cell type and cell lysis. Multiple displacement amplification (MDA) amplifies the DNA isothermally with the use of Phi29 polymerase and random hexamer primers. The efficiency and accuracy of MDA was assessed on different cell types (buccal cells, lymphocytes, fibroblasts) using two multiplex PCR reactions that have been applied in clinical preimplantation genetic diagnosis cases (DM triplex-DM1, APOC2, Dl9S112 and CF triplex-DF508del, IVS8CA, IVS17TA). These results were compared using the DM triplex with MDA products from single blastomeres. Cells were lysed using a modified protocol excluding dithiothreitol in the alkaline lysis buffer. The MDA amplification efficiency for buccal cells was 82.0% (41/50) compared with 96.0% (48/50) for lymphocytes and 100% (20/20) for fibroblasts. The average allele dropout (ADO) rates were 31.0% for buccal cells, 20.8% for lymphocytes and 20.0% for fibroblasts with high inter-locus variation across all cell types (5.0-45.5%). Overall, MDA on single lymphocytes and fibroblasts lysed using the modified protocol produced DNA of sufficient quantity and quality for subsequent molecular analysis by PCR and gave results comparable with MDA products from blastomeres, in contrast to buccal cells. PMID:19573296

  1. Development and Testing of Shingle-type Solar Cell Modules

    NASA Technical Reports Server (NTRS)

    Shepard, N. F., Jr.

    1979-01-01

    The design, development, fabrication and testing of a shingle-type terrestrial solar cell module which produces 98 watts/sq m of exposed module area at 1 kW/sq m insolation and 61 C are reported. These modules make it possible to easily incorporate photovoltaic power generation into the sloping roofs of residential or commercial buildings by simply nailing the modules to the plywood roof sheathing. This design consists of nineteen series-connected 53 mm diameter solar cells arranged in a closely packed hexagon configuration. These cells are individually bonded to the embossed surface of a 3 mm thick thermally tempered hexagon-shaped piece of glass. Polyvinyl butyral is used as the laminating adhesive.

  2. Induction of Human Squamous Cell-Type Carcinomas by Arsenic

    PubMed Central

    Martinez, Victor D.; Becker-Santos, Daiana D.; Vucic, Emily A.; Lam, Stephen; Lam, Wan L.

    2011-01-01

    Arsenic is a potent human carcinogen. Around one hundred million people worldwide have potentially been exposed to this metalloid at concentrations considered unsafe. Exposure occurs generally through drinking water from natural geological sources, making it difficult to control this contamination. Arsenic biotransformation is suspected to have a role in arsenic-related health effects ranging from acute toxicities to development of malignancies associated with chronic exposure. It has been demonstrated that arsenic exhibits preference for induction of squamous cell carcinomas in the human, especially skin and lung cancer. Interestingly, keratins emerge as a relevant factor in this arsenic-related squamous cell-type preference. Additionally, both genomic and epigenomic alterations have been associated with arsenic-driven neoplastic process. Some of these aberrations, as well as changes in other factors such as keratins, could explain the association between arsenic and squamous cell carcinomas in humans. PMID:22175027

  3. Coordination between donor cell type and cell cycle stage improves nuclear cloning efficiency in cattle.

    PubMed

    Wells, D N; Laible, G; Tucker, F C; Miller, A L; Oliver, J E; Xiang, T; Forsyth, J T; Berg, M C; Cockrem, K; L'Huillier, P J; Tervit, H R; Oback, B

    2003-01-01

    Several studies have shown that both quiescent and proliferating somatic donor cells can be fully reprogrammed after nuclear transfer (NT) and result in viable offspring. So far, however, no comparative study has conclusively demonstrated the relative importance of donor cell cycle stage on nuclear cloning efficiency. Here, we compare two different types of bovine fetal fibroblasts (BFFs) that were synchronized in G(0), G(1), and different phases within G(1). We show that for non-transgenic (non-TG) fibroblasts, serum starvation into G(0) results in a significantly higher percentage of viable calves at term than synchronization in early G(1) or late G(1). For transgenic fibroblasts, however, cells selected in G(1) show significantly higher development to calves at term and higher post-natal survival to weaning than cells in G(0). This suggests that it may be necessary to coordinate donor cell type and cell cycle stage to maximize overall cloning efficiency.

  4. Melatonin modulates M4-type ganglion-cell photoreceptors.

    PubMed

    Pack, W; Hill, D D; Wong, K Y

    2015-09-10

    In the retina, melatonin is secreted at night by rod/cone photoreceptors and serves as a dark-adaptive signal. Melatonin receptors have been found in many retinal neurons including melanopsin-containing intrinsically photosensitive retinal ganglion cells (ipRGCs), suggesting it could modulate the physiology of these inner retinal photoreceptors. Here, we investigated whether melatonin modulates the alpha-like M4-type ipRGCs, which are believed to mediate image-forming vision as well as non-image-forming photoresponses. Applying melatonin during daytime (when endogenous melatonin secretion is low) caused whole-cell-recorded M4 cells' rod/cone-driven depolarizing photoresponses to become broader and larger, whereas the associated elevation in spike rate was reduced. Melanopsin-based light responses were not affected significantly. Nighttime application of the melatonin receptor antagonist luzindole also altered M4 cells' rod/cone-driven light responses but in the opposite ways: the duration and amplitude of the graded depolarization were reduced, whereas the accompanying spiking increase was enhanced. These luzindole-induced changes confirmed that M4 cells are modulated by endogenous melatonin. Melatonin could induce the above effects by acting directly on M4 cells because immunohistochemistry detected MT1 receptors in these cells, although it could also act presynaptically. Interestingly, the daytime and nighttime recordings showed significant differences in resting membrane potential, spontaneous spike rate and rod/cone-driven light responses, suggesting that M4 cells are under circadian control. This is the first report of a circadian variation in ipRGCs' resting properties and synaptic input, and of melatoninergic modulation of ipRGCs. PMID:26141846

  5. Differential transcriptional profiling of damaged and intact adjacent dorsal root ganglia neurons in neuropathic pain.

    PubMed

    Reinhold, A K; Batti, L; Bilbao, D; Buness, A; Rittner, H L; Heppenstall, P A

    2015-01-01

    Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG. PMID:25880204

  6. Differential transcriptional profiling of damaged and intact adjacent dorsal root ganglia neurons in neuropathic pain.

    PubMed

    Reinhold, A K; Batti, L; Bilbao, D; Buness, A; Rittner, H L; Heppenstall, P A

    2015-01-01

    Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

  7. Differential Transcriptional Profiling of Damaged and Intact Adjacent Dorsal Root Ganglia Neurons in Neuropathic Pain

    PubMed Central

    Reinhold, A. K.; Batti, L.; Bilbao, D.; Buness, A.; Rittner, H. L.; Heppenstall, P. A.

    2015-01-01

    Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and “bystanders,” thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG. PMID:25880204

  8. Cardiac stem cell therapy: Have we put too much hype in which cell type to use?

    PubMed

    Ye, Jianqin; Yeghiazarians, Yerem

    2015-09-01

    Injection of various stem cells has been tested with the hopes of improving cardiac function after a myocardial infarction (MI). However, there is continued controversy as to which cell type is best for repair. Due to technical differences in cell isolation, processing, delivery, and cardiac functional assessment by various investigators, it has been difficult to directly compare the results of different cells. Using same techniques to evaluate the efficacy of different cell types, we have separately delivered bone marrow cells (BMCs), cardiospheres (CSs), CS-derived Sca-1(+)/CD45(-) cells, human embryonic stem cell-derived cardiomyocytes, and BMC extract into infarcted murine myocardium and found that all of these treatments reduce infarct size and improve cardiac function post-MI similarly without one regimen being superior to another. The beneficial effects appear to be via paracrine influences. Different progenitors lead to improved cardiac function post-MI, but it is premature to hype any specific cell type at this time. PMID:26024953

  9. How subaerial salt extrusions influence water quality in adjacent aquifers

    NASA Astrophysics Data System (ADS)

    Mehdizadeh, Razieh; Zarei, Mehdi; Raeisi, Ezzat

    2015-12-01

    Brines supplied from salt extrusions cause significant groundwater salinization in arid and semi-arid regions where salt rock is exposed to dissolution by episodic rainfalls. Here we focus on 62 of the 122 diapirs of Hormuz salt emergent in the southern Iran. To consider managing the degradation effect that salt extrusions have on the quality of adjoining aquifers, it is first necessary to understand how they influence adjacent water resources. We evaluate here the impacts that these diapirs have on adjacent aquifers based on investigating their geomorphologies, geologies, hydrologies and hydrogeologies. The results indicate that 28/62 (45%) of our sample of salt diapirs have no significant impact on the quality of groundwater in adjoining aquifers (namely Type N), while the remaining 34/62 (55%) degrade nearby groundwater quality. We offer simple conceptual models that account for how brines flowing from each of these types of salt extrusions contaminate adjacent aquifers. We identify three main mechanisms that lead to contamination: surface impact (Type A), subsurface intrusion (Type B) and indirect infiltration (Type C). A combination of all these mechanisms degrades the water quality in nearby aquifers in 19/62 (31%) of the salt diapirs studied. Having characterized the mechanism(s) by which each diapir affects the adjacent aquifer, we suggest a few possible remediation strategies to be considered. For instance, engineering the surface runoff of diapirs Types A and C into nearby evaporation basins would improve groundwater quality.

  10. Type 1 diabetes immunotherapy using polyclonal regulatory T cells.

    PubMed

    Bluestone, Jeffrey A; Buckner, Jane H; Fitch, Mark; Gitelman, Stephen E; Gupta, Shipra; Hellerstein, Marc K; Herold, Kevan C; Lares, Angela; Lee, Michael R; Li, Kelvin; Liu, Weihong; Long, S Alice; Masiello, Lisa M; Nguyen, Vinh; Putnam, Amy L; Rieck, Mary; Sayre, Peter H; Tang, Qizhi

    2015-11-25

    Type 1 diabetes (T1D) is an autoimmune disease that occurs in genetically susceptible individuals. Regulatory T cells (Tregs) have been shown to be defective in the autoimmune disease setting. Thus, efforts to repair or replace Tregs in T1D may reverse autoimmunity and protect the remaining insulin-producing β cells. On the basis of this premise, a robust technique has been developed to isolate and expand Tregs from patients with T1D. The expanded Tregs retained their T cell receptor diversity and demonstrated enhanced functional activity. We report on a phase 1 trial to assess safety of Treg adoptive immunotherapy in T1D. Fourteen adult subjects with T1D, in four dosing cohorts, received ex vivo-expanded autologous CD4(+)CD127(lo/-)CD25(+) polyclonal Tregs (0.05 × 10(8) to 26 × 10(8) cells). A subset of the adoptively transferred Tregs was long-lived, with up to 25% of the peak level remaining in the circulation at 1 year after transfer. Immune studies showed transient increases in Tregs in recipients and retained a broad Treg FOXP3(+)CD4(+)CD25(hi)CD127(lo) phenotype long-term. There were no infusion reactions or cell therapy-related high-grade adverse events. C-peptide levels persisted out to 2+ years after transfer in several individuals. These results support the development of a phase 2 trial to test efficacy of the Treg therapy. PMID:26606968

  11. Type 2 Diabetes Dysregulates Glucose Metabolism in Cardiac Progenitor Cells.

    PubMed

    Salabei, Joshua K; Lorkiewicz, Pawel K; Mehra, Parul; Gibb, Andrew A; Haberzettl, Petra; Hong, Kyung U; Wei, Xiaoli; Zhang, Xiang; Li, Qianhong; Wysoczynski, Marcin; Bolli, Roberto; Bhatnagar, Aruni; Hill, Bradford G

    2016-06-24

    Type 2 diabetes is associated with increased mortality and progression to heart failure. Recent studies suggest that diabetes also impairs reparative responses after cell therapy. In this study, we examined potential mechanisms by which diabetes affects cardiac progenitor cells (CPCs). CPCs isolated from the diabetic heart showed diminished proliferation, a propensity for cell death, and a pro-adipogenic phenotype. The diabetic CPCs were insulin-resistant, and they showed higher energetic reliance on glycolysis, which was associated with up-regulation of the pro-glycolytic enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3). In WT CPCs, expression of a mutant form of PFKFB, which mimics PFKFB3 activity and increases glycolytic rate, was sufficient to phenocopy the mitochondrial and proliferative deficiencies found in diabetic cells. Consistent with activation of phosphofructokinase in diabetic cells, stable isotope carbon tracing in diabetic CPCs showed dysregulation of the pentose phosphate and glycero(phospho)lipid synthesis pathways. We describe diabetes-induced dysregulation of carbon partitioning using stable isotope metabolomics-based coupling quotients, which relate relative flux values between metabolic pathways. These findings suggest that diabetes causes an imbalance in glucose carbon allocation by uncoupling biosynthetic pathway activity, which could diminish the efficacy of CPCs for myocardial repair. PMID:27151219

  12. Inhibition of type I NKT cells by retinoids or following sulfatide-mediated activation of type II NKT cells attenuates alcoholic liver disease

    PubMed Central

    Maricic, Igor; Sheng, Huiming; Marrero, Idania; Seki, Ehikiro; Kisseleva, Tatiana; Chaturvedi, Som; Molle, Natasha; Mathews, K. Stephanie; Gao, Bin; Kumar, Vipin

    2015-01-01

    Innate immune mechanisms leading to liver injury following chronic alcohol ingestion are poorly understood. Natural killer T (NKT) cells, enriched in the liver and comprised of at least two distinct subsets, type I and type II, recognize different lipid antigens presented by CD1d molecules. We have investigated whether differential activation of NKT cell subsets orchestrates inflammatory events leading to alcoholic liver disease (ALD). We found that following chronic plus binge feeding of Lieber-DeCarli liquid diet in male C57BL/6 mice, type I but not type II NKT cells are activated leading to recruitment of inflammatory Gr-1highCD11b+ cells into liver. A central finding is that liver injury following alcohol feeding is dependent upon type I NKT cells. Thus liver injury is significantly inhibited in Jα18−/− mice deficient in type I NKT cells as well as following their inactivation by sulfatide-mediated activation of type II NKT cells. Furthermore we have identified a novel pathway involving all-trans retinoic acid (ATRA) and its receptor RARγ signaling that inhibits type I NKT cells and consequently ALD. A semi-quantitative PCR analysis of hepatic gene expression of some of the key proinflammatory molecules shared in human disease indicated that their upregulation in ALD is dependent upon type I NKT cells. Conclusion Type I but not type II NKT cells become activated following alcohol feeding. Type I NKT cells-induced inflammation and neutrophil recruitment results in liver tissue damage while type II NKT cells protect from injury in ALD. Inhibition of type I NKT cells by retinoids or by sulfatide prevents ALD. Since the CD1d pathway is highly conserved between mice and humans, NKT cell subsets might be targeted for potential therapeutic intervention in ALD. PMID:25477000

  13. Global methylation profiles in DNA from different blood cell types

    PubMed Central

    Wu, Hui-Chen; Delgado-Cruzata, Lissette; Flom, Julie D; Kappil, Maya; Ferris, Jennifer S; Liao, Yuyan; Santella, Regina M

    2011-01-01

    DNA methylation measured in white blood cell DNA is increasingly being used in studies of cancer susceptibility. However, little is known about the correlation between different assays to measure global methylation and whether the source of DNA matters when examining methylation profiles in different blood cell types. Using information from 620 women, 217 and 403 women with DNA available from granulocytes (Gran) and total white blood cells (WBC), respectively, and 48 women with DNA available from four different sources [WBC, Gran, mononuclear (MN) and lymphoblastoid cell lines (LCL)], we compared DNA methylation for three repetitive elements (LINE1, Sat2, Alu) by MethyLight, luminometric methylation assay (LUMA) and [3H]-methyl acceptance assay. For four of the five assays, DNA methylation levels measured in Gran were not correlated with methylation in LCL, MN or WBC; the exception was Sat2. DNA methylation in LCL was correlated with methylation in MN and WBC for the [3H]-methyl acceptance, LINE1 and Alu assays. Methylation in MN was correlated with methylation in WBC for the [3H]-methyl acceptance and LUMA assays. When we compared the five assays to each other by source of DNA, we observed statistically significant correlations ranging from 0.3–0.7 for each cell type with one exception (Sat2 and Alu in MN). Among the 620 women stratified by DNA source, correlations among assays were highest for the three repetitive elements (range 0.39–0.64). Results from the LUMA assay were modestly correlated with LINE1 (0.18–0.20). These results suggest that both assay and source of DNA are critical components in the interpretation of global DNA methylation patterns from WBC. PMID:20890131

  14. Single-Cell mRNA Profiling Reveals Cell-Type-Specific Expression of Neurexin Isoforms.

    PubMed

    Fuccillo, Marc V; Földy, Csaba; Gökce, Özgün; Rothwell, Patrick E; Sun, Gordon L; Malenka, Robert C; Südhof, Thomas C

    2015-07-15

    Neurexins are considered central organizers of synapse architecture that are implicated in neuropsychiatric disorders. Expression of neurexins in hundreds of alternatively spliced isoforms suggested that individual neurons might exhibit a cell-type-specific neurexin expression pattern (a neurexin code). To test this hypothesis, we quantified the single-cell levels of neurexin isoforms and other trans-synaptic cell-adhesion molecules by microfluidics-based RT-PCR. We show that the neurexin repertoire displays pronounced cell-type specificity that is remarkably consistent within each type of neuron. Furthermore, we uncovered region-specific regulation of neurexin transcription and splice-site usage. Finally, we demonstrate that the transcriptional profiles of neurexins can be altered in an experience-dependent fashion by exposure to a drug of abuse. Our data provide evidence of cell-type-specific expression patterns of multiple neurexins at the single-cell level and suggest that expression of synaptic cell-adhesion molecules overlaps with other key features of cellular identity and diversity.

  15. Single-Cell mRNA Profiling Reveals Cell-Type Specific Expression of Neurexin Isoforms

    PubMed Central

    Fuccillo, Marc V.; Földy, Csaba; Gökce, Özgün; Rothwell, Patrick E.; Sun, Gordon L.; Malenka, Robert C.; Südhof, Thomas C.

    2016-01-01

    Summary Neurexins are considered central organizers of synapse architecture that are implicated in neuropsychiatric disorders. Expression of neurexins in hundreds of alternatively spliced isoforms suggested that individual neurons might exhibit a cell type-specific neurexin expression pattern (a neurexin code). To test this hypothesis, we quantified the single-cell levels of neurexin isoforms and other trans-synaptic cell-adhesion molecules by microfluidics-based RT-PCR. We show that the neurexin repertoire displays pronounced cell-type specificity that is remarkably consistent within each type of neuron. Furthermore, we uncovered region-specific regulation of neurexin transcription and splice-site usage. Finally, we demonstrate that the transcriptional profiles of neurexins can be altered in an experience-dependent fashion by exposure to a drug of abuse. Our data provide evidence of cell type-specific expression patterns of multiple neurexins at the single-cell level, and suggest that expression of synaptic cell-adhesion molecules overlaps with other key features of cellular identity and diversity. PMID:26182417

  16. 3-D Reconstruction of Macular Type II Cell Innervation Patterns in Space-Flight and Control Rats

    NASA Technical Reports Server (NTRS)

    Ross, Muriel Dorothy; Montgomery, K.; Linton, S.; Cheng, R.; Tomko, David L. (Technical Monitor)

    1995-01-01

    A semiautomated method for reconstructing objects from serial thin sections has been developed in the Biocomputation Center. The method is being used to completely, for the first time, type II hair cells and their innervations. The purposes are to learn more about the fundamental circuitry of the macula on Earth and to determine whether changes in connectivities occur under space flight conditions. Data captured directly from a transmission electron microscope via a video camera are sent to a graphics workstation. There, the digitized micrographs are mosaicked into sections and contours are traced, registered and displayed by semiautomated methods. Current reconstructions are of type II cells from the medial part of rat maculas collected in-flight on the Space Life Sciences-2 mission, 4.5 hrs post-flight, and from a ground control. Results show that typical type II cells receive processes from tip to six nearby calyces or afferents. Nearly all processes are elongated and have bouton-like enlargements; some have numerous vesicles. Multiple (2 to 4) processes from a single calyx to a type II cell are common, and approximately 1/3 of the processes innervale 2 or 3 type II cells or a neighboring cluster. From 2% to 6% of the cells resemble type I cells morphologically but have demi-calyces. Thus far, increments in synaptic number in type II cells of flight rats are prominent along processes that supply two hair cells. It is clear that reconstruction methods provide insights into details of macular circuitry not obtainable by other techniques. The results demonstrate a morphological basis for interactions between adjacent receptive fields through feed back-feed forward connections, and for dynamic alterations in receptive field range and activity during preprocessing of linear acceleratory information by the maculas. The reconstruction method we have developed will find further applications in the study of the details of neuronal architecture of more complex systems, to

  17. Generation of stem cell-derived β-cells from patients with type 1 diabetes

    PubMed Central

    Millman, Jeffrey R.; Xie, Chunhui; Van Dervort, Alana; Gürtler, Mads; Pagliuca, Felicia W.; Melton, Douglas A.

    2016-01-01

    We recently reported the scalable in vitro production of functional stem cell-derived β-cells (SC-β cells). Here we extend this approach to generate the first SC-β cells from type 1 diabetic patients (T1D). β-cells are destroyed during T1D disease progression, making it difficult to extensively study them in the past. These T1D SC-β cells express β-cell markers, respond to glucose both in vitro and in vivo, prevent alloxan-induced diabetes in mice and respond to anti-diabetic drugs. Furthermore, we use an in vitro disease model to demonstrate the cells respond to different forms of β-cell stress. Using these assays, we find no major differences in T1D SC-β cells compared with SC-β cells derived from non-diabetic patients. These results show that T1D SC-β cells could potentially be used for the treatment of diabetes, drug screening and the study of β-cell biology. PMID:27163171

  18. Liver stem cell-derived β-cell surrogates for treatment of type 1 diabetes☆

    PubMed Central

    Yang, Li-Jun

    2012-01-01

    Consistent with the common embryonic origin of liver and pancreas as well the similar glucose-sensing systems in hepatocytes and pancreatic β-cells, it should not be surprising that liver stem cells/hepatocytes can transdifferentiate into insulin-producing cells under high-glucose culture conditions or by genetic reprogramming. Persistent expression of the pancreatic duodenal homeobox-1 (Pdx1) transcription factor or its super-active form Pdx1-VP16 fusion protein in hepatic cells reprograms these cells into pancreatic β-cell precursors. In vitro culture at elevated glucose concentrations or in vivo exposure to a hyperglycemia are required for further differentiation and maturation of liver-derived pancreatic β-cell precursor into functional insulin-producing pancreatic β-like cells. Under appropriate conditions, multiple pancreatic transcription factors can work in concert to reprogram liver stem/adult liver cells into functional insulin-producing cells. If such autologous liver-derived insulin-producing cells can be made to escape the type 1 diabetes-associated autoimmunity, they may serve as a valuable cell source for future cell replacement therapy without the need for life-long immunosuppression. PMID:16890895

  19. Morphine biosynthesis in opium poppy involves two cell types: sieve elements and laticifers.

    PubMed

    Onoyovwe, Akpevwe; Hagel, Jillian M; Chen, Xue; Khan, Morgan F; Schriemer, David C; Facchini, Peter J

    2013-10-01

    Immunofluorescence labeling and shotgun proteomics were used to establish the cell type-specific localization of morphine biosynthesis in opium poppy (Papaver somniferum). Polyclonal antibodies for each of six enzymes involved in converting (R)-reticuline to morphine detected corresponding antigens in sieve elements of the phloem, as described previously for all upstream enzymes transforming (S)-norcoclaurine to (S)-reticuline. Validated shotgun proteomics performed on whole-stem and latex total protein extracts generated 2031 and 830 distinct protein families, respectively. Proteins corresponding to nine morphine biosynthetic enzymes were represented in the whole stem, whereas only four of the final five pathway enzymes were detected in the latex. Salutaridine synthase was detected in the whole stem, but not in the latex subproteome. The final three enzymes converting thebaine to morphine were among the most abundant active latex proteins despite a limited occurrence in laticifers suggested by immunofluorescence labeling. Multiple charge isoforms of two key O-demethylases in the latex were revealed by two-dimensional immunoblot analysis. Salutaridine biosynthesis appears to occur only in sieve elements, whereas conversion of thebaine to morphine is predominant in adjacent laticifers, which contain morphine-rich latex. Complementary use of immunofluorescence labeling and shotgun proteomics has substantially resolved the cellular localization of morphine biosynthesis in opium poppy.

  20. Concise review: alchemy of biology: generating desired cell types from abundant and accessible cells.

    PubMed

    Pournasr, Behshad; Khaloughi, Keynoush; Salekdeh, Ghasem Hosseini; Totonchi, Mehdi; Shahbazi, Ebrahim; Baharvand, Hossein

    2011-12-01

    A major goal of regenerative medicine is to produce cells to participate in the generation, maintenance, and repair of tissues that are damaged by disease, aging, or trauma, such that function is restored. The establishment of induced pluripotent stem cells, followed by directed differentiation, offers a powerful strategy for producing patient-specific therapies. Given how laborious and lengthy this process can be, the conversion of somatic cells into lineage-specific stem/progenitor cells in one step, without going back to, or through, a pluripotent stage, has opened up tremendous opportunities for regenerative medicine. However, there are a number of obstacles to overcome before these cells can be widely considered for clinical applications. Here, we focus on induced transdifferentiation strategies to convert mature somatic cells to other mature cell types or progenitors, and we summarize the challenges that need to be met if the potential applications of transdifferentiation technology are to be achieved.

  1. Concise review: alchemy of biology: generating desired cell types from abundant and accessible cells.

    PubMed

    Pournasr, Behshad; Khaloughi, Keynoush; Salekdeh, Ghasem Hosseini; Totonchi, Mehdi; Shahbazi, Ebrahim; Baharvand, Hossein

    2011-12-01

    A major goal of regenerative medicine is to produce cells to participate in the generation, maintenance, and repair of tissues that are damaged by disease, aging, or trauma, such that function is restored. The establishment of induced pluripotent stem cells, followed by directed differentiation, offers a powerful strategy for producing patient-specific therapies. Given how laborious and lengthy this process can be, the conversion of somatic cells into lineage-specific stem/progenitor cells in one step, without going back to, or through, a pluripotent stage, has opened up tremendous opportunities for regenerative medicine. However, there are a number of obstacles to overcome before these cells can be widely considered for clinical applications. Here, we focus on induced transdifferentiation strategies to convert mature somatic cells to other mature cell types or progenitors, and we summarize the challenges that need to be met if the potential applications of transdifferentiation technology are to be achieved. PMID:21997905

  2. Clear cell renal cell carcinoma with a syncytial-type multinucleated giant tumor cell component: implications for differential diagnosis.

    PubMed

    Williamson, Sean R; Kum, Jennifer B; Goheen, Michael P; Cheng, Liang; Grignon, David J; Idrees, Muhammad T

    2014-04-01

    A component of syncytial-type multinucleated tumor giant cells is uncommon in clear cell renal cell carcinoma, and the histogenesis, incidence, and clinical implications of this finding are not well understood. We retrieved 13 such tumors from our pathology archives in patients with a median age of 60years, comprising 1.5% of clear cell renal cell carcinomas. Stage was typically pT4 or pT3 (each 38%). Microscopically, all tumors included a component of low-grade clear cell renal cell carcinoma with usual features. Syncytial-type giant tumor cells possessed voluminous cytoplasm, usually granular and eosinophilic, and numerous nuclei similar to those of the mononuclear tumor cells. Transition between areas of mononuclear and multinucleated cells was sometimes abrupt. Other findings included necrosis (77%), hyaline globules (46%), emperipolesis (46%), and intranuclear cytoplasmic invaginations (23%). Immunohistochemical staining typically revealed both mononuclear and multinucleated cells to be positive for carbonic anhydrase IX, CD10, epithelial membrane antigen, vimentin, and cytokeratin AE1/AE3 and negative for β human chorionic gonadotropin, TFE3, cathepsin K, cytokeratin 7, cytokeratin 20, HMB45, CD68, smooth muscle actin, and S100. Most patients with available information (7/9) were alive with metastatic disease at the most recent follow-up. Syncytial-type giant cells are an uncommon finding associated with aggressive clear cell renal cell carcinomas. Despite the unusual appearance of this tumor component, its immunoprofile supports an epithelial lineage and argues against trophoblastic, osteoclast-like, or histiocytic differentiation. Reactivity for typical clear cell renal cell carcinoma antigens facilitates discrimination from giant cells of epithelioid angiomyolipoma or other tumors, particularly in a biopsy specimen or a metastatic tumor. PMID:24499686

  3. Thiol-ene Michael-type formation of gelatin/poly(ethylene glycol) biomatrices for three-dimensional mesenchymal stromal/stem cell administration to cutaneous wounds.

    PubMed

    Xu, Kedi; Cantu, David Antonio; Fu, Yao; Kim, Jaehyup; Zheng, Xiaoxiang; Hematti, Peiman; Kao, W John

    2013-11-01

    Mesenchymal stromal/stem cells (MSCs) are considered promising cellular therapeutics in the fields of tissue engineering and regenerative medicine. MSCs secrete high concentrations of immunomodulatory cytokines and growth factors, which exert paracrine effects on infiltrating immune and resident cells in the wound microenvironment that could favorably promote healing after acute injury. However, better spatial delivery and improved retention at the site of injury are two factors that could improve the clinical application of MSCs. In this study, we utilized thiol-ene Michael-type addition for rapid encapsulation of MSCs within a gelatin/poly(ethylene glycol) biomatrix. This biomatrix was also applied as a provisional dressing to full thickness wounds in Sprague-Dawley rats. The three-way interaction of MSCs, gelatin/poly(ethylene glycol) biomatrices, and host immune cells and adjacent resident cells in the wound microenvironment favorably modulated wound progression and host response. In this model we observed attenuated immune cell infiltration, lack of foreign giant cell (FBGC) formation, accelerated wound closure and re-epithelialization, as well as enhanced neovascularization and granulation tissue formation by 7 days. The MSC entrapped in the gelatin/poly(ethylene glycol) biomatrix localized cell presentation adjacent to the wound microenvironment and thus mediated the early resolution of inflammatory events and facilitated the proliferative phases in wound healing. PMID:23811217

  4. Cell Type-Specific Differences in Spike Timing and Spike Shape in the Rat Parasubiculum and Superficial Medial Entorhinal Cortex.

    PubMed

    Ebbesen, Christian Laut; Reifenstein, Eric Torsten; Tang, Qiusong; Burgalossi, Andrea; Ray, Saikat; Schreiber, Susanne; Kempter, Richard; Brecht, Michael

    2016-07-26

    The medial entorhinal cortex (MEC) and the adjacent parasubiculum are known for their elaborate spatial discharges (grid cells, border cells, etc.) and the precessing of spikes relative to the local field potential. We know little, however, about how spatio-temporal firing patterns map onto cell types. We find that cell type is a major determinant of spatio-temporal discharge properties. Parasubicular neurons and MEC layer 2 (L2) pyramids have shorter spikes, discharge spikes in bursts, and are theta-modulated (rhythmic, locking, skipping), but spikes phase-precess only weakly. MEC L2 stellates and layer 3 (L3) neurons have longer spikes, do not discharge in bursts, and are weakly theta-modulated (non-rhythmic, weakly locking, rarely skipping), but spikes steeply phase-precess. The similarities between MEC L3 neurons and MEC L2 stellates on one hand and parasubicular neurons and MEC L2 pyramids on the other hand suggest two distinct streams of temporal coding in the parahippocampal cortex. PMID:27425616

  5. Curcumin inhibits bovine herpesvirus type 1 entry into MDBK cells.

    PubMed

    Zhu, L; Ding, X; Zhang, D; Yuan, Ch; Wang, J; Ndegwa, E; Zhu, G

    2015-09-01

    The generation of antiviral drugs from herbs and other natural resources with traditionally long-confirmed effects is an efficient approach. So far, no herb or components from herbs that could inhibit bovine herpesvirus type 1 (BoHV-1) replication have been described. In this study, the antiviral effect of curcumin, a natural phenolic constituent of the spice turmeric, on BoHV-1 replication was evaluated in cell culture. We demonstrated that curcumin impairs BoHV-1 viral particles and affects the virus post-binding entry process. Furthermore, curcumin upregulated the proportion of the plasma membrane adopting a lipid raft conformation in MDBK cells, which supported the previous reports that curcumin can modulate the lipid bilayer. Though the antiviral mechanism of curcumin on BoHV-1 needs further study, we identified for the first time a component from herb that could inhibit BoHV-1 replication, in vitro.

  6. Transplantation of mesenchymal stem cells improves type 1 diabetes mellitus.

    PubMed

    Li, Lisha; Li, Furong; Gao, Feng; Yang, Yali; Liu, Yuanyuan; Guo, Pingping; Li, Yulin

    2016-05-01

    Bone-marrow-derived stem cells can regenerate pancreatic tissue in a model of type 1 diabetes mellitus. Mesenchymal stem cells (MSCs) form the main part of bone marrow. We show that the intrapancreatic transplantation of MSCs elevates serum insulin and C-peptide, while decreasing blood glucose. MSCs engrafted into the damaged rat pancreas become distributed into the blood vessels, acini, ducts, and islets. Renascent islets, islet-like clusters, and a small number of MSCs expressing insulin protein have been observed in the pancreas of diabetic rats. Intrapancreatic transplantation of MSCs triggers a series of molecular and cellular events, including differentiation towards the pancreas directly and the provision of a niche to start endogenous pancreatic regeneration, which ameliorates hypoinsulinemia and hyperglycemia caused by streptozotocin. These data establish the many roles of MSCs in the restoration of the function of an injured organ. PMID:26650464

  7. Induction of DNA Double-Strand Breaks and Cellular Migration Through Bystander Effects in Cells Irradiated With the Slit-Type Microplanar Beam of the Spring-8 Synchrotron

    SciTech Connect

    Kashino, Genro Kondoh, Takeshi; Nariyama, Nobuteru; Umetani, Keiji; Ohigashi, Takuji; Shinohara, Kunio; Kurihara, Ai; Fukumoto, Manabu; Tanaka, Hiroki; Maruhashi, Akira; Suzuki, Minoru; Kinashi, Yuko; Liu Yong; Masunaga, Shin-ichiro; Watanabe, Masami; Ono, Koji

    2009-05-01

    Purpose: To determine whether glioma cells irradiated with a microplanar X-ray beam exert bystander effects. Methods and Materials: Microplanar beam irradiation of glioma cells in vitro was done using the SPring-8 synchrotron radiation facility. The amount of DNA double-strand breaks (dsbs) was measured by the fluorescence intensity of phosphorylated H2AX or the number of 53BP1 foci. The dose distribution in a cell population exposed to a single microplanar beam was determined by the amount of phosphorylated H2AX-positive cells. Bystander effects were determined by counting the number of 53BP1 foci in nonirradiated cells treated with conditioned medium from cultures of irradiated cells. Results: More DNA dsbs were detected in cells adjacent to an area irradiated by the single beam than in cells in distant, nonirradiated areas as a result of bystander effects caused by scattered X-rays and DNA dsbs. In support of this, more 53BP1 foci were observed in nonirradiated, conditioned medium-treated cells than in control cells (i.e., cells not treated with irradiation or conditioned medium). These results suggest that DNA dsbs were induced in nonirradiated cells by soluble factors in the culture medium. In addition, we observed cellular migration into areas irradiated with peak doses, suggesting that irradiated cells send signals that cause nonirradiated cells to migrate toward damaged cells. Conclusions: Bystander effects are produced by factors secreted as a result of slit-type microplanar X-ray beam irradiation.

  8. Target Cell Cyclophilins Facilitate Human Papillomavirus Type 16 Infection

    PubMed Central

    Sapp, Martin

    2009-01-01

    Following attachment to primary receptor heparan sulfate proteoglycans (HSPG), human papillomavirus type 16 (HPV16) particles undergo conformational changes affecting the major and minor capsid proteins, L1 and L2, respectively. This results in exposure of the L2 N-terminus, transfer to uptake receptors, and infectious internalization. Here, we report that target cell cyclophilins, peptidyl-prolyl cis/trans isomerases, are required for efficient HPV16 infection. Cell surface cyclophilin B (CyPB) facilitates conformational changes in capsid proteins, resulting in exposure of the L2 N-terminus. Inhibition of CyPB blocked HPV16 infection by inducing noninfectious internalization. Mutation of a putative CyP binding site present in HPV16 L2 yielded exposed L2 N-terminus in the absence of active CyP and bypassed the need for cell surface CyPB. However, this mutant was still sensitive to CyP inhibition and required CyP for completion of infection, probably after internalization. Taken together, these data suggest that CyP is required during two distinct steps of HPV16 infection. Identification of cell surface CyPB will facilitate the study of the complex events preceding internalization and adds a putative drug target for prevention of HPV–induced diseases. PMID:19629175

  9. Single-cell-type proteomics: toward a holistic understanding of plant function.

    PubMed

    Dai, Shaojun; Chen, Sixue

    2012-12-01

    Multicellular organisms such as plants contain different types of cells with specialized functions. Analyzing the protein characteristics of each type of cell will not only reveal specific cell functions, but also enhance understanding of how an organism works. Most plant proteomics studies have focused on using tissues and organs containing a mixture of different cells. Recent single-cell-type proteomics efforts on pollen grains, guard cells, mesophyll cells, root hairs, and trichomes have shown utility. We expect that high resolution proteomic analyses will reveal novel functions in single cells. This review provides an overview of recent developments in plant single-cell-type proteomics. We discuss application of the approach for understanding important cell functions, and we consider the technical challenges of extending the approach to all plant cell types. Finally, we consider the integration of single-cell-type proteomics with transcriptomics and metabolomics with the goal of providing a holistic understanding of plant function.

  10. Cell type specific expression of p11 controls cocaine reward

    PubMed Central

    Arango-Lievano, Margarita; Schwarz, Justin T.; Vernov, Mary; Wilkinson, Matthew B.; Bradbury, Kathryn; Feliz, Akira; Marongiu, Roberta; Gelfand, Yaroslav; Warner-Schmidt, Jennifer; Nestler, Eric J.; Greengard, Paul; Russo, Scott J.; Kaplitt, Michael G.

    2014-01-01

    Background The high rate of comorbidity between depression and cocaine addiction suggests shared molecular mechanisms and anatomical pathways. Limbic structures, such as the Nucleus Accumbens (NAc), play a crucial role in both disorders, yet how different cell types within these structures contribute to the pathogenesis remains elusive. Downregulation of p11 (S100A10) specifically in the NAc elicits depressive like behaviors in mice but its role in drug addiction is unknown. Methods We combine mouse genetics and viral strategies to determine how the titration of p11 levels within the entire NAc affects the rewarding actions of cocaine on behavior (6 to 8 mice per group) and molecular correlates (3 experiments, 5 to 8 mice per group). Finally, the manipulation of p11 expression in distinct NAc dopaminoceptive neuronal subsets distinguished cell type specific effects of p11 on cocaine reward (5 to 8 mice per group). Results We demonstrate that p11 knockout mice have enhanced cocaine conditioned place preference (CPP), which is reproduced by the focal downregulation of p11 in the NAc of wild-type mice. In wild-type mice, cocaine reduced p11 expression in the NAc, while p11 overexpression exclusively in the NAc reduced cocaine CPP. Finally, we identify dopamine receptor-1 (D1) expressing medium spiny neurons (MSNs) as key mediators of p11’s effects on cocaine reward. Conclusions Our data provide evidence that disruption of p11 homeostasis in the NAc particularly in D1 expressing MSNs may underlie pathophysiological mechanisms of cocaine rewarding action. Treatments to counter maladaptation of p11 levels may provide novel therapeutic opportunities for cocaine addiction. PMID:24725970

  11. Islet transplantation versus stem cells for the cell therapy of type 1 diabetes mellitus.

    PubMed

    Basta, G; Montanucci, P; Calafiore, R

    2015-12-01

    Pancreatic islet cell transplantation has represented the mainstay of cell therapy for the potential, final cure of type 1 diabetes mellitus (T1D), along the past two decades. Unfortunately, the restricted availability of cadaveric human donor pancreases coupled with heavy side effects of the recipient's general immunosuppression, have severely crippled progress of this approach into clinical trials. Only a few excellence centers, worldwide, have thus far accrued still quite marginal clinical success. In an attempt to overcome the limits of islet transplantation new technologies for use of several stem cell lineages are being under investigation, with initial experimental evidence of success. Essentially, the actual lines of research involve attempts to either activate native endogenous stem cells that replace diseased/dead cells, by a cell regeneration process, or condition other stem cells to acquire the functional properties of the targeted cells to be substituted (i.e., beta-cell-like elements associated with insulin secretory competence). A wide array of stem cells may fulfill this task, from embryonic (whose use still faces strong ethical barriers), to adult, to induced pluripotent stem cells. Mesenchymal adult stem cells, retrievable from many different sites, including adipose tissue, bone marrow and post-partum umbilical cord Wharton Jelly, seem to couple plastic to immunoregulatory properties that might greatly help progress for the disease cure.

  12. Micropatterned superhydrophobic structures for the simultaneous culture of multiple cell types and the study of cell-cell communication.

    PubMed

    Efremov, Alexander N; Stanganello, Eliana; Welle, Alexander; Scholpp, Steffen; Levkin, Pavel A

    2013-02-01

    The ability to control spatial arrangement and geometry of different cell types while keeping them separated and in close proximity for a long time is crucial to mimic and study variety of biological processes in vitro. Although the existing cell patterning technologies allow co-culturing of different cell types, they are usually limited to relatively simple geometry. The methods used for obtaining complex geometries are usually applicable for patterning only one or two cell types. Here we introduce a convenient method for creating patterns of multiple (up to twenty) different cell types on one substrate. The method virtually allows any complexity of cell pattern geometry. Cell positioning on the substrate is realized by a parallel formation of multiple cell-containing microreservoirs confined to the geometry of highly hydrophilic regions surrounded by superhydrophobic borders built-in a fine nanoporous polymer film. As a case study we showed the cross-talk between two cell populations via Wnt signaling molecules propagation during co-culture in a mutual culture medium. PMID:23228425

  13. Human cell type diversity, evolution, development, and classification with special reference to cells derived from the neural crest.

    PubMed

    Vickaryous, Matthew K; Hall, Brian K

    2006-08-01

    Metazoans are composed of a finite number of recognisable cell types. Similar to the relationship between species and ecosystems, knowledge of cell type diversity contributes to studies of complexity and evolution. However, as with other units of evolution, the cell type often resists definition. This review proposes guidelines for characterising cell types and discusses cell homology and the various developmental pathways by which cell types arise, including germ layers, blastemata (secondary development/neurulation), stem cells, and transdifferentiation. An updated list of cell types is presented for a familiar, albeit overlooked model taxon, adult Homo sapiens, with 411 cell types, including 145 types of neurons, recognised. Two methods for organising these cell types are explored. One is the artificial classification technique, clustering cells using commonly accepted criteria of similarity. The second approach, an empirical method modeled after cladistics, resolves the classification in terms of shared features rather than overall similarity. While the results of each scheme differ, both methods address important questions. The artificial classification provides compelling (and independent) support for the neural crest as the fourth germ layer, while the cladistic approach permits the evaluation of cell type evolution. Using the cladistic approach we observe a correlation between the developmental and evolutionary origin of a cell, suggesting that this method is useful for predicting which cell types share common (multipotential) progenitors. Whereas the current effort is restricted by the availability of phenotypic details for most cell types, the present study demonstrates that a comprehensive cladistic classification is practical, attainable, and warranted. The use of cell types and cell type comparative classification schemes has the potential to offer new and alternative models for therapeutic evaluation. PMID:16790079

  14. FAMA is an essential component for the differentiation of two distinct cell types, myrosin cells and guard cells, in Arabidopsis.

    PubMed

    Shirakawa, Makoto; Ueda, Haruko; Nagano, Atsushi J; Shimada, Tomoo; Kohchi, Takayuki; Hara-Nishimura, Ikuko

    2014-10-01

    Brassicales plants, including Arabidopsis thaliana, have an ingenious two-compartment defense system, which sequesters myrosinase from the substrate glucosinolate and produces a toxic compound when cells are damaged by herbivores. Myrosinase is stored in vacuoles of idioblast myrosin cells. The molecular mechanism that regulates myrosin cell development remains elusive. Here, we identify the basic helix-loop-helix transcription factor FAMA as an essential component for myrosin cell development along Arabidopsis leaf veins. FAMA is known as a regulator of stomatal development. We detected FAMA expression in myrosin cell precursors in leaf primordia in addition to stomatal lineage cells. FAMA deficiency caused defects in myrosin cell development and in the biosynthesis of myrosinases THIOGLUCOSIDE GLUCOHYDROLASE1 (TGG1) and TGG2. Conversely, ectopic FAMA expression conferred myrosin cell characteristics to hypocotyl and root cells, both of which normally lack myrosin cells. The FAMA interactors ICE1/SCREAM and its closest paralog SCREAM2/ICE2 were essential for myrosin cell development. DNA microarray analysis identified 32 candidate genes involved in myrosin cell development under the control of FAMA. This study provides a common regulatory pathway that determines two distinct cell types in leaves: epidermal guard cells and inner-tissue myrosin cells.

  15. 8. Exterior view, showing tank and associated piping adjacent to ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    8. Exterior view, showing tank and associated piping adjacent to Test Cell 6, Systems Integration Laboratory Building (T-28), looking south. - Air Force Plant PJKS, Systems Integration Laboratory, Systems Integration Laboratory Building, Waterton Canyon Road & Colorado Highway 121, Lakewood, Jefferson County, CO

  16. Membrane Targeting of P-type ATPases in Plant Cells

    SciTech Connect

    Jeffrey F. Harper, Ph.D.

    2004-06-30

    How membrane proteins are targeted to specific subcellular locations is a very complex and poorly understood area of research. Our long-term goal is to use P-type ATPases (ion pumps), in a model plant system Arabidopsis, as a paradigm to understand how members of a family of closely related membrane proteins can be targeted to different subcellular locations. The research is divided into two specific aims. The first aim is focused on determining the targeting destination of all 10 ACA-type calcium pumps (Arabidopsis Calcium ATPase) in Arabidopsis. ACAs represent a plant specific-subfamily of plasma membrane-type calcium pumps. In contrast to animals, the plant homologs have been found in multiple membrane systems, including the ER (ACA2), tonoplast (ACA4) and plasma membrane (ACA8). Their high degree of similarity provides a unique opportunity to use a comparative approach to delineate the membrane specific targeting information for each pump. One hypothesis to be tested is that an endomembrane located ACA can be re-directed to the plasma membrane by including targeting information from a plasma membrane isoform, ACA8. Our approach is to engineer domain swaps between pumps and monitor the targeting of chimeric proteins in plant cells using a Green Fluorescence Protein (GFP) as a tag. The second aim is to test the hypothesis that heterologous transporters can be engineered into plants and targeted to the plasma membrane by fusing them to a plasma membrane proton pump. As a test case we are evaluating the targeting properties of fusions made between a yeast sodium/proton exchanger (Sod2) and a proton pump (AHA2). This fusion may potentially lead to a new strategy for engineering salt resistant plants. Together these aims are designed to provide fundamental insights into the biogenesis and function of plant cell membrane systems.

  17. Thiol-ene Michael-type formation of gelatin/poly(ethylene glycol) biomatrices for three-dimensional mesenchymal stromal/stem cell administration to cutaneous wounds

    PubMed Central

    Xu, Kedi; Cantu, David Antonio; Fu, Yao; Kim, Jaehyup; Zheng, Xiaoxiang; Hematti, Peiman; Kao, W. John

    2013-01-01

    Mesenchymal stromal/stem cells (MSCs) are considered promising cellular therapeutics in the fields of tissue engineering and regenerative medicine. MSCs secrete high concentrations of immunomodulatory cytokines and growth factors, which exert paracrine effects on infiltrating immune and resident cells of the wound microenvironment that could favorably promote healing after acute injury. However, better spatial delivery and improved retention at the site of injury are two factors that could improve the clinical application of MSCs. In this study, we utilized thiol-ene Michael-type addition for rapid encapsulation of MSCs within a gelatin/poly(ethylene glycol) biomatrix; this biomatrix was also applied as a provisional dressing to full-thickness wounds in Sprague-Dawley rats. The three-way interaction of MSCs, gelatin/poly(ethylene glycol) biomatrices, and host immune cells and adjacent resident cells of the wound microenvironment favorably modulated wound progression and host response. In this model we observed attenuated immune cell infiltration, lack of foreign giant cell (FBGC) formation, accelerated wound closure and re-epithelialization, as well as enhanced neovascularization and granulation tissue formation by 7 days. The MSC-entrapped gelatin/poly(ethylene glycol) biomatrix localized the presentation of MSCs adjacent to the wound microenvironment and thus, mediated early resolution of inflammatory events and facilitated proliferative phases in wound healing. PMID:23811217

  18. Dissecting engineered cell types and enhancing cell fate conversion via CellNet

    PubMed Central

    Morris, Samantha A.; Cahan, Patrick; Li, Hu; Zhao, Anna M.; San Roman, Adrianna K.; Shivdasani, Ramesh A.; Collins, James J.; Daley, George Q.

    2014-01-01

    SUMMARY Engineering clinically relevant cells in vitro holds promise for regenerative medicine, but most protocols fail to faithfully recapitulate target cell properties. To address this, we developed CellNet, a network biology platform that determines whether engineered cells are equivalent to their target tissues, diagnoses aberrant gene regulatory networks, and prioritizes candidate transcriptional regulators to enhance engineered conversions. Using CellNet, we improved B cell to macrophage conversion, transcriptionally and functionally, by knocking down predicted B cell regulators. Analyzing conversion of fibroblasts to induced hepatocytes (iHeps), CellNet revealed an unexpected intestinal program regulated by the master regulator Cdx2. We observed long-term functional engraftment of mouse colon by iHeps, thereby establishing their broader potential as endoderm progenitors and demonstrating direct conversion of fibroblasts into intestinal epithelium. Our studies illustrate how CellNet can be employed to improve direct conversion and to uncover unappreciated properties of engineered cells. PMID:25126792

  19. Immune cell populations in cutaneous delayed-type hypersensitivity

    PubMed Central

    1983-01-01

    Delayed-type hypersensitivity (DTH) is a prototypic T lymphocyte- mediated response to antigenic challenge. In this study, mononuclear cells infiltrating the skin during cutaneous response to tuberculin in presensitized human subjects (responders) and nonimmune controls were identified using monoclonal antibodies by indirect immunofluorescence. In both responders and controls the infiltrate consisted mainly of T lymphocytes (T11+ and OKT3+) and monocytes (OKM1+, 63D3+, Mo2+) which initially accumulated in proximity to small blood vessels and later infiltrated the interstitial dermis and epidermis. More T lymphocytes reacted with OKT4 than with OKT8. 6 h after tuberculin the ratio of OKT4/OKT8 in tissue from responders exceeded that in blood, whereas in tissues studied at 15-48 h and in all control tissues those ratios in blood and tissue were similar. Evidence of T lymphocyte activation was sought using monoclonal antibodies anti-Tac, OKT9, and OKT10. In responders but not in controls the proportion of infiltrating cells reactive with these antibodies increased during the course of DTH. The presence of activated T lymphocytes in tissue was not associated with a comparable increase in peripheral blood cell populations identified by anti-Tac and OKT10. Studies using anti-B1, Leu-7, and anti-IgD/IgM revealed comparatively few reactive cells. Dual-labeling studies demonstrated that most Leu-7--reactive cells also bound T11 while fewer bound OKM1 or OKT8 and that cells reactive with OKIa1 and T11 constituted largely nonoverlapping populations. Specific patterns of reactivity were not observed when tissues were stained with anti-human C3, or poly C9-MA, a monoclonal antibody reactive with a neoantigen on polymerized C9 of the membrane attack complex of complement. The number of epidermal Langerhans cells identified by OKT6 was similar in responders and controls. Thus, the cutaneous response to tuberculin in sensitized individuals is characterized by early enrichment of

  20. Massively parallel single cell RNA-Seq for marker-free decomposition of tissues into cell types

    PubMed Central

    Jaitin, Diego Adhemar; Kenigsberg, Ephraim; Keren-Shaul, Hadas; Elefant, Naama; Paul, Franziska; Zaretsky, Irina; Mildner, Alexander; Cohen, Nadav; Jung, Steffen; Tanay, Amos; Amit, Ido

    2015-01-01

    In multi-cellular organisms, biological function emerges when heterogeneous cell types form complex organs. Nevertheless dissection of tissues into mixtures of cellular subpopulations is currently challenging. We introduce an automated massively parallel single-cell RNA sequencing approach for analyzing in vivo transcriptional states in thousands of single cells. Combined with unsupervised classification algorithms, this facilitates ab initio cell type characterization of splenic tissues. Modeling single-cell transcriptional states in dendritic cells and additional hematopoietic cell types uncovers rich cell-type heterogeneity and gene-modules activity in steady-state and after pathogen activation. Cellular diversity is thereby approached through inference of variable and dynamic pathway activity rather than a fixed pre-programmed cell-type hierarchy. These data demonstrate single-cell RNA-Seq as an effective tool for comprehensive cellular decomposition of complex tissues. PMID:24531970

  1. The Macrophage Galactose-Type C-Type Lectin (MGL) Modulates Regulatory T Cell Functions

    PubMed Central

    Zizzari, Ilaria Grazia; Martufi, Paola; Battisti, Federico; Rahimi, Hassan; Caponnetto, Salvatore; Bellati, Filippo; Nuti, Marianna

    2015-01-01

    Regulatory T cells (Tregs) are physiologically designed to prevent autoimmune disease and maintain self-tolerance. In tumour microenvironments, their presence is related to a poor prognosis, and they influence the therapeutic outcome due to their capacity to suppress the immune response by cell-cell contact and to release immunosuppressive cytokines. In this study, we demonstrate that Treg immunosuppressive activity can be modulated by the cross-linking between the CD45RA expressed by Tregs and the C-type lectin MGL. This specific interaction strongly decreases the im-munosuppressive activity of Tregs, restoring the proliferative capacity of co-cultured T lymphocytes. This effect can be attributed to changes in CD45RA and TCR signalling through the inhibition of Lck and inactivation of Zap-70, an increase in the Foxp3 methylation status and, ultimately, the reduced production of suppressive cytokines. These results indicate a role of MGL as an immunomodulator within the tumour microenvironment interfering with Treg functions, suggesting its possible use in the design of anticancer vaccines. PMID:26147970

  2. Cannabinoid Type 2 Receptors Mediate a Cell Type-Specific Plasticity in the Hippocampus.

    PubMed

    Stempel, A Vanessa; Stumpf, Alexander; Zhang, Hai-Ying; Özdoğan, Tuğba; Pannasch, Ulrike; Theis, Anne-Kathrin; Otte, David-Marian; Wojtalla, Alexandra; Rácz, Ildikó; Ponomarenko, Alexey; Xi, Zheng-Xiong; Zimmer, Andreas; Schmitz, Dietmar

    2016-05-18

    Endocannabinoids (eCBs) exert major control over neuronal activity by activating cannabinoid receptors (CBRs). The functionality of the eCB system is primarily ascribed to the well-documented retrograde activation of presynaptic CB1Rs. We find that action potential-driven eCB release leads to a long-lasting membrane potential hyperpolarization in hippocampal principal cells that is independent of CB1Rs. The hyperpolarization, which is specific to CA3 and CA2 pyramidal cells (PCs), depends on the activation of neuronal CB2Rs, as shown by a combined pharmacogenetic and immunohistochemical approach. Upon activation, they modulate the activity of the sodium-bicarbonate co-transporter, leading to a hyperpolarization of the neuron. CB2R activation occurred in a purely self-regulatory manner, robustly altered the input/output function of CA3 PCs, and modulated gamma oscillations in vivo. To conclude, we describe a cell type-specific plasticity mechanism in the hippocampus that provides evidence for the neuronal expression of CB2Rs and emphasizes their importance in basic neuronal transmission. PMID:27133464

  3. β-cell replacement sources for type 1 diabetes: a focus on pancreatic ductal cells.

    PubMed

    Corritore, Elisa; Lee, Yong-Syu; Sokal, Etienne M; Lysy, Philippe A

    2016-08-01

    Thorough research on the capacity of human islet transplantation to cure type 1 diabetes led to the achievement of 3- to 5-year-long insulin independence in nearly half of transplanted patients. Yet, translation of this technique to clinical routine is limited by organ shortage and the need for long-term immunosuppression, restricting its use to adults with unstable disease. The production of new bona fide β cells in vitro was thus investigated and finally achieved with human pluripotent stem cells (PSCs). Besides ethical concerns about the use of human embryos, studies are now evaluating the possibility of circumventing the spontaneous tumor formation associated with transplantation of PSCs. These issues fueled the search for cell candidates for β-cell engineering with safe profiles for clinical translation. In vivo studies revealed the regeneration capacity of the exocrine pancreas after injury that depends at least partially on facultative progenitors in the ductal compartment. These stimulated subpopulations of pancreatic ductal cells (PDCs) underwent β-cell transdifferentiation through reactivation of embryonic signaling pathways. In vitro models for expansion and differentiation of purified PDCs toward insulin-producing cells were described using cocktails of growth factors, extracellular-matrix proteins and transcription factor overexpression. In this review, we will describe the latest findings in pancreatic β-cell mass regeneration due to adult ductal progenitor cells. We will further describe recent advances in human PDC transdifferentiation to insulin-producing cells with potential for clinical translational studies.

  4. Casein gene expression in mouse mammary epithelial cell lines: Dependence upon extracellular matrix and cell type

    SciTech Connect

    Medina, D.; Oborn, C.J. ); Li, M.L.; Bissell, M.J. )

    1987-09-01

    The COMMA-D mammary cell line exhibits mammary-specific functional differentiation under appropriate conditions in cell culture. The cytologically heterogeneous COMMA-D parental line and the clonal lines DB-1, TA-5, and FA-1 derived from the COMMA-D parent were examined for similar properties of functional differentiation. In monolayer cell culture, the cell lines DB-1, TA-5, FA-1, and MA-4 were examined for expression of mammary-specific and epithelial-specific proteins by an indirect immunofluorescence assay. The clonal cell lines were relatively homogeneous in their respective staining properties and seemed to represent three subpopulations found in the heterogeneous parental COMMA-D lines. None of the four clonal lines appeared to represent myoepithelial cells. The cell lines were examined for expression of {beta}-casein mRNA in the presence or absence of prolactin. The inducibility of {beta}-casein in the COMMA-D cell line was further enhanced by a reconstituted basement membrane preparation enriched in laminin, collagen IV, and proteoglycans. These results support the hypothesis that the functional response of inducible mammary cell populations is a result of interaction among hormones, multiple extracellular matrix components, and specific cell types.

  5. β-cell replacement sources for type 1 diabetes: a focus on pancreatic ductal cells.

    PubMed

    Corritore, Elisa; Lee, Yong-Syu; Sokal, Etienne M; Lysy, Philippe A

    2016-08-01

    Thorough research on the capacity of human islet transplantation to cure type 1 diabetes led to the achievement of 3- to 5-year-long insulin independence in nearly half of transplanted patients. Yet, translation of this technique to clinical routine is limited by organ shortage and the need for long-term immunosuppression, restricting its use to adults with unstable disease. The production of new bona fide β cells in vitro was thus investigated and finally achieved with human pluripotent stem cells (PSCs). Besides ethical concerns about the use of human embryos, studies are now evaluating the possibility of circumventing the spontaneous tumor formation associated with transplantation of PSCs. These issues fueled the search for cell candidates for β-cell engineering with safe profiles for clinical translation. In vivo studies revealed the regeneration capacity of the exocrine pancreas after injury that depends at least partially on facultative progenitors in the ductal compartment. These stimulated subpopulations of pancreatic ductal cells (PDCs) underwent β-cell transdifferentiation through reactivation of embryonic signaling pathways. In vitro models for expansion and differentiation of purified PDCs toward insulin-producing cells were described using cocktails of growth factors, extracellular-matrix proteins and transcription factor overexpression. In this review, we will describe the latest findings in pancreatic β-cell mass regeneration due to adult ductal progenitor cells. We will further describe recent advances in human PDC transdifferentiation to insulin-producing cells with potential for clinical translational studies. PMID:27540464

  6. β-cell replacement sources for type 1 diabetes: a focus on pancreatic ductal cells

    PubMed Central

    Corritore, Elisa; Lee, Yong-Syu; Sokal, Etienne M.; Lysy, Philippe A.

    2016-01-01

    Thorough research on the capacity of human islet transplantation to cure type 1 diabetes led to the achievement of 3- to 5-year-long insulin independence in nearly half of transplanted patients. Yet, translation of this technique to clinical routine is limited by organ shortage and the need for long-term immunosuppression, restricting its use to adults with unstable disease. The production of new bona fide β cells in vitro was thus investigated and finally achieved with human pluripotent stem cells (PSCs). Besides ethical concerns about the use of human embryos, studies are now evaluating the possibility of circumventing the spontaneous tumor formation associated with transplantation of PSCs. These issues fueled the search for cell candidates for β-cell engineering with safe profiles for clinical translation. In vivo studies revealed the regeneration capacity of the exocrine pancreas after injury that depends at least partially on facultative progenitors in the ductal compartment. These stimulated subpopulations of pancreatic ductal cells (PDCs) underwent β-cell transdifferentiation through reactivation of embryonic signaling pathways. In vitro models for expansion and differentiation of purified PDCs toward insulin-producing cells were described using cocktails of growth factors, extracellular-matrix proteins and transcription factor overexpression. In this review, we will describe the latest findings in pancreatic β-cell mass regeneration due to adult ductal progenitor cells. We will further describe recent advances in human PDC transdifferentiation to insulin-producing cells with potential for clinical translational studies. PMID:27540464

  7. Type I interferon regulation of natural killer cell function in primary and secondary infections.

    PubMed

    Stackaruk, Michele L; Lee, Amanda J; Ashkar, Ali A

    2013-08-01

    The priming of natural killer (NK) cells by type I interferon (IFN) is necessary for protection against primary and secondary viral infections. However, the pathway by which type I IFN activates NK cells to elicit antiviral responses is controversial. There is evidence to suggest that type I IFN priming of NK cells occurs through both direct and indirect pathways. As with many innate mechanisms, type I IFN and NK cells also orchestrate the adaptive immune response and thus aid in protection against secondary infections. Type I IFN can shape CD4(+) T cell, B cell and humoral memory formation. In addition, long-lived NK cells can perform specific and enhanced memory-like protection in secondary infections. This review outlines the different mechanisms underlying type I IFN regulation of NK cells and how type I IFN and NK cells can be used as a therapeutic target in vaccinations.

  8. Stem cell approaches for the treatment of type 1 diabetes mellitus.

    PubMed

    Wagner, Ryan T; Lewis, Jennifer; Cooney, Austin; Chan, Lawrence

    2010-09-01

    Type 1 diabetes is characterized by near total absence of pancreatic b cells. Current treatments consisting of insulin injections and islet transplantation are clinically unsatisfactory. In order to develop a cure for type 1 diabetes, we must find a way to reverse autoimmunity, which underlies b cell destruction, as well as an effective strategy to generate new b cells. This article reviews the different approaches that are being taken to produce new b cells. Much emphasis has been placed on selecting the right non-b cell population, either in vivo or in vitro, as the starting material. Different cell types, including adult stem cells, other types of progenitor cells in situ, and even differentiated cell populations, as well as embryonic stem cells and induced pluripotent stem cells, will require different methods for islet and b cell induction. We discussed the pros and cons of the different strategies that are being used to re-invent the pancreatic b cell.

  9. Singling out Drosophila tendon cells: a dialogue between two distinct cell types.

    PubMed

    Volk, T

    1999-11-01

    The precise match between somatic muscles and their epidermal attachment cells is achieved through a continuous dialogue between these two cell types. Whereas tendon cells direct myotube migration and final patterning, the muscles are essential for the maintenance of the fate of tendon cells. The Drosophila neuregulin-like ligand, Vein, and its receptor, the epidermal growth factor receptor (Egfr), are critical components in the inductive signaling process that takes place between muscles and tendon cells. Additional gene products that relay the Vein-Egfr effect in Drosophila are conserved in the vertebrate neuregulin-mediated cascade. This review describes genetic and molecular aspects of the muscle-tendon inductive processes in Drosophila, and compares them with the relevant mechanisms in the vertebrate embryo.

  10. Cell-type homologies and the origins of the neocortex

    PubMed Central

    Dugas-Ford, Jennifer; Rowell, Joanna J.; Ragsdale, Clifton W.

    2012-01-01

    The six-layered neocortex is a uniquely mammalian structure with evolutionary origins that remain in dispute. One long-standing hypothesis, based on similarities in neuronal connectivity, proposes that homologs of the layer 4 input and layer 5 output neurons of neocortex are present in the avian forebrain, where they contribute to specific nuclei rather than to layers. We devised a molecular test of this hypothesis based on layer-specific gene expression that is shared across rodent and carnivore neocortex. Our findings establish that the layer 4 input and the layer 5 output cell types are conserved across the amniotes, but are organized into very different architectures, forming nuclei in birds, cortical areas in reptiles, and cortical layers in mammals. PMID:23027930

  11. Metal-sulfur type cell having improved positive electrode

    DOEpatents

    Dejonghe, Lutgard C.; Visco, Steven J.; Mailhe, Catherine C.; Armand, Michel B.

    1989-01-01

    An novel metal-sulfur type cell operable at a temperature of 200.degree. C. or less with an energy density of 150 Whrs/Kg or better is disclosed characterized by an organo-sulfur cathode formed from an organic-sulfur compound having the general formula, in its charged state, of (R(S).sub.y).sub.n wherein y=1 to 6; n=2 to 20; and R is one or more different aliphatic or aromatic organic moieties having 1 to 20 carbon atoms, which may include one or more oxygen, sulfur, or nitrogen heteroatoms when R comprisises one of more aromatic rings, or one or more oxygen, sulfur, nitrogen, or fluorine atoms associtated with the chain when R comprises an aliphatic chain, wherein the aliphatic group may be linear or branched, saturated or unsaturated, and wherein either the aliphatic chain or the aromatic ring may have substituted groups thereon.

  12. Metal-sulfur type cell having improved positive electrode

    DOEpatents

    DeJonghe, L.C.; Visco, S.J.; Mailhe, C.C.; Armand, M.B.

    1988-03-31

    A novel metal-sulfur type cell operable at a temperature of 200/degree/C or less with an energy density of 150 Whrs/Kg or better is disclosed characterized by an organo-sulfur cathode formed from an organic-sulfur compound having the general formula, in its charged state, of (R(S)/sub y/)n wherein y = 1 to 6; n = 2 to 20; and R is one or more different aliphatic or aromatic organic moieties having 1 to 20 carbon atoms, which may include one or more oxygen, sulfur, or nitrogen heteroatoms when R comprises one or more aromatic rings, or one or more oxygen, sulfur, nitrogen, or fluorine atoms associated with the chain when R comprises an aliphatic chain, wherein the aliphatic group may be linear or branched, saturated or unsaturated, and wherein either the aliphatic chain or the aromatic ring may have substituted groups thereon. 4 figs.

  13. Metal-sulfur type cell having improved positive electrode

    NASA Astrophysics Data System (ADS)

    Dejonghe, Lutgard C.; Visco, Steven J.; Mailhe, Catherine C.; Armand, Michel B.

    1988-03-01

    A novel metal-sulfur type cell operable at a temperature of 200 C or less with an energy density of 150 Whrs/Kg or better is disclosed characterized by an organo-sulfur cathode formed from an organic-sulfur compound having the general formula, in its charged state, of (R(S) sub y) n wherein y = 1 to 6; n = 2 to 20; and R is one or more different aliphatic or aromatic organic moieties having 1 to 20 carbon atoms, which may include one or more oxygen, sulfur, or nitrogen heteroatoms when R comprises one or more aromatic rings, or one or more oxygen, sulfur, nitrogen, or fluorine atoms associated with the chain when R comprises an aliphatic chain, wherein the aliphatic group may be linear or branched, saturated or unsaturated, and wherein either the aliphatic chain or the aromatic ring may have substituted groups thereon.

  14. Disease-Associated SNPs From non-Coding Regions in Juvenile Idiopathic Arthritis Are Located Within or Adjacent to Functional Genomic Elements of Human Neutrophils and CD4+ T Cells

    PubMed Central

    Jiang, Kaiyu; Zhu, Lisha; Buck, Michael J.; Chen, Yanmin; Carrier, Bradley; Liu, Tao; Jarvis, James N.

    2015-01-01

    Background Juvenile idiopathic arthritis (JIA) is considered a complex trait in which the environment interacts with inherited genes to produce a phenotype that shows broad inter-individual variance. A recently completed genome-wide association study (GWAS) identified 24 regions of genetic risk for JIA, for example. However, as is typical for GWAS, most of the regions of genetic risk for JIA (22 of 24) were in non-coding regions of the genome. The studies reported here were undertaken to identify functional elements (other than genes) that might be located within the regions of genetic risk. Methods We used paired end RNA sequencing to identify non-coding RNAs located within 5 kb of the disease-associated SNPs. In addition, we used chromatin immunoprecipitation-sequencing (ChIP-Seq) to identify epigenetic marks associated with enhancer function (H3K4me1 and H3K27ac) in human neutrophils to determine whether there was enrichment of enhancer-associated histone marks in linkage disequilibrium (LD) blocks that encompassed the 22 GWAS SNPs from the non-coding genome. Results In human neutrophils, we identified H3K4me1 and/or H3K27ac marks in 15 of the 22 regions previously as identified as risk loci for JIA. In CD4+ T cells, 18 regions demonstrate H3K4me1 and/or H3K27ac marks. In addition, we identified non-coding RNA transcripts at the rs4705862 and rs6894249 loci in human neutrophils. Conclusion Much of the genetic risk for JIA lies within or adjacent to regions of neutrophil and CD4+ T cell genomes that carry epigenetic marks associated with enhancer function and/or ncRNA transcripts. These findings are consistent with the hypothesis that JIA is fundamentally a disorder of gene regulation that includes both the innate and adaptive immune system. Elucidating the specific roles of these non-coding elements within leukocyte genomes in JIA pathogenesis will be critical to our understanding disease pathogenesis. PMID:25833190

  15. Silver nanowire interactions with primary human alveolar type-II epithelial cell secretions: contrasting bioreactivity with human alveolar type-I and type-II epithelial cells

    PubMed Central

    Sweeney, Sinbad; Theodorou, Ioannis G.; Zambianchi, Martina; Chen, Shu; Gow, Andrew; Schwander, Stephan; Zhang, Junfeng (Jim); Chung, Kian Fan; Shaffer, Milo S.; Ryan, Mary P.; Porter, Alexandra E.; Tetley, Teresa D.

    2015-01-01

    Inhaled nanoparticles have a high deposition rate in the alveolar units of the deep lung. The alveolar epithelium is composed of type-I and type-II epithelial cells (ATI and ATII respectively) and is bathed in pulmonary surfactant. The effect of native human ATII cell secretions on nanoparticle toxicity is not known. We investigated the cellular uptake and toxicity of silver nanowires (AgNWs; 70 nm diameter, 1.5 μm length) with human ATI-like cells (TT1), in the absence or presence of Curosurf® (a natural porcine pulmonary surfactant with a low amount of protein) or harvested primary human ATII cell secretions (HAS; containing both the complete lipid as well as the full protein complement of human pulmonary surfactant i.e. SP-A, SP-B, SP-C and SP-D). We hypothesised that Curosurf® or HAS would confer improved protection for TT1 cells, limiting the toxicity of AgNWs. In agreement with our hypothesis, HAS reduced the inflammatory and reactive oxygen species (ROS)-generating potential of AgNWs with exposed TT1 cells. For example, IL-8 release and ROS generation was reduced by 38% and 29%, respectively, resulting in similar levels to that of the non-treated controls. However in contrast to our hypothesis, Curosurf® had no effect. We found a significant reduction in AgNW uptake by TT1 cells in the presence of HAS but not Curosurf. Furthermore, we show that the SP-A and SP-D are likely to be involved in this process as they were found to be specifically bound to the AgNWs. While ATI cells appear to be protected by HAS, evidence suggested that ATII cells, despite no uptake, were vulnerable to AgNW exposure (indicated by increased IL-8 release and ROS generation and decreased intracellular SP-A levels one day post-exposure). This study provides unique findings that may be important for the study of lung epithelial-endothelial translocation of nanoparticles in general and associated toxicity within the alveolar unit. PMID:25996248

  16. Evidence for Differential Glycosylation of Trophoblast Cell Types.

    PubMed

    Chen, Qiushi; Pang, Poh-Choo; Cohen, Marie E; Longtine, Mark S; Schust, Danny J; Haslam, Stuart M; Blois, Sandra M; Dell, Anne; Clark, Gary F

    2016-06-01

    Human placental villi are surfaced by the syncytiotrophoblast (STB), with a layer of cytotrophoblasts (CTB) positioned just beneath the STB. STB in normal term pregnancies is exposed to maternal immune cells in the placental intervillous space. Extravillous cytotrophoblasts (EVT) invade the decidua and spiral arteries, where they act in conjunction with natural killer (NK) cells to convert the spiral arteries into flaccid conduits for maternal blood that support a 3-4 fold increase in the rate of maternal blood flow into the placental intervillous space. The functional roles of these distinct trophoblast subtypes during pregnancy suggested that they could be differentially glycosylated. Glycomic analysis of these trophoblasts has revealed the expression of elevated levels of biantennary N-glycans in STB and CTB, with the majority of them bearing a bisecting GlcNAc. N-glycans terminated with polylactosamine extensions were also detected at low levels. A subset of the N-glycans linked to these trophoblasts were sialylated, primarily with terminal NeuAcα2-3Gal sequences. EVT were decorated with the same N-glycans as STB and CTB, except in different proportions. The level of bisecting type N-glycans was reduced, but the level of N-glycans decorated with polylactosamine sequences were substantially elevated compared with the other types of trophoblasts. The level of triantennary and tetraantennary N-glycans was also elevated in EVT. The sialylated N-glycans derived from EVT were completely susceptible to an α2-3 specific neuraminidase (sialidase S). The possibility exists that the N-glycans associated with these different trophoblast subpopulations could act as functional groups. These potential relationships will be considered. PMID:26929217

  17. Polyploidization and localisation of poly(A)+ RNA in the different cell types of the vitellogenic meroistic ovary of the fleshfly, Sarcophaga bullata.

    PubMed

    Cardoen, J; Schoofs, L; Broekaert, D; Van Mellaert, H; Verachtert, B; De Loof, A

    1986-01-01

    The degree of polyploidization, the level of transcriptional activity and the volume of the different cell types present in the meroistic ovary of Sarcophaga bullata were measured during different vitellogenic stages. The nurse cells and the germinal vesicle exhibited very pronounced differences with regard to DNA content and mRNA synthesis, even though they are genetically identical. During the 4C stage (late vitellogenesis), we observed different degrees of polyploidy in follicle cells adjacent to the oocyte and those surrounding the nurse cells. Although the chromatin of the germinal vesicle is condensed into a karyosome, in situ hybridisation revealed the presence of transcriptional activity. The volume of the germinal vesicle, which contains only 4C DNA, is big enough to contain 2048C DNA. The meroistic ovary is a highly polarized differentiating system. Our results are discussed in the light of the fact that the polytrophic ovary is a miniature electrophoresis chamber.

  18. Cell compartmentalisation in planctomycetes: novel types of structural organisation for the bacterial cell.

    PubMed

    Lindsay, M R; Webb, R I; Strous, M; Jetten, M S; Butler, M K; Forde, R J; Fuerst, J A

    2001-06-01

    The organisation of cells of the planctomycete species Pirellula marina, Isosphaera pallida, Gemmata obscuriglobus, Planctomyces maris and "Candidatus Brocadia anammoxidans" was investigated based on ultrastructure derived from thin-sections of cryosubstituted cells, freeze-fracture replicas, and in the case of Gemmata obscuriglobus and Pirellula marina, computer-aided 3-D reconstructions from serial sections of cryosubstituted cells. All planctomycete cells display a peripheral ribosome-free region, termed here the paryphoplasm, surrounding the perimeter of the cell, and an interior region including any nucleoid regions as well as ribosome-like particles, bounded by a single intracytoplasmic membrane (ICM), and termed the pirellulosome in Pirellula species. Immunogold labelling and RNase-gold cytochemistry indicates that in planctomycetes all the cell DNA is contained wholly within the interior region bounded by the ICM, and the paryphoplasm contains no DNA but at least some of the cell's RNA. The ICM in Isosphaera pallida and Planctomyces maris is invaginated such that the paryphoplasm forms a major portion of the cell interior in sections, but in other planctomycetes it remains as a peripheral zone. In the anaerobic ammonium-oxidising ("anammox" process) chemoautotroph "Candidatus Brocadia anammoxidans" the interior region bounded by ICM contains a further internal single-membrane-bounded region, the anammoxosome. In Gemmata obscuriglobus, the interior ICM-bounded region contains the nuclear body, a double-membrane-bounded region containing the cell's nucleoid and all genomic DNA in addition to some RNA. Shared features of cell compartmentalisation in different planctomycetes are consistent with the monophyletic nature of the planctomycetes as a distinct division of the Bacteria. The shared organisational plan for the planctomycete cell constitutes a new type not known in cells of other bacteria. PMID:11491082

  19. Galvanic Cell Type Sensor for Soil Moisture Analysis.

    PubMed

    Gaikwad, Pramod; Devendrachari, Mruthyunjayachari Chattanahalli; Thimmappa, Ravikumar; Paswan, Bhuneshwar; Raja Kottaichamy, Alagar; Makri Nimbegondi Kotresh, Harish; Thotiyl, Musthafa Ottakam

    2015-07-21

    Here we report the first potentiometric sensor for soil moisture analysis by bringing in the concept of Galvanic cells wherein the redox energies of Al and conducting polyaniline are exploited to design a battery type sensor. The sensor consists of only simple architectural components, and as such they are inexpensive and lightweight, making it suitable for on-site analysis. The sensing mechanism is proved to be identical to a battery type discharge reaction wherein polyaniline redox energy changes from the conducting to the nonconducting state with a resulting voltage shift in the presence of soil moisture. Unlike the state of the art soil moisture sensors, a signal derived from the proposed moisture sensor is probe size independent, as it is potentiometric in nature and, hence, can be fabricated in any shape or size and can provide a consistent output signal under the strong aberration conditions often encountered in soil moisture analysis. The sensor is regenerable by treating with 1 M HCl and can be used for multiple analysis with little read out hysteresis. Further, a portable sensor is fabricated which can provide warning signals to the end user when the moisture levels in the soil go below critically low levels, thereby functioning as a smart device. As the sensor is inexpensive, portable, and potentiometric, it opens up avenues for developing effective and energy efficient irrigation strategies, understanding the heat and water transfer at the atmosphere-land interface, understanding soil mechanics, forecasting the risk of natural calamities, and so on.

  20. DNA methylation status predicts cell type-specific enhancer activity

    PubMed Central

    Wiench, Malgorzata; John, Sam; Baek, Songjoon; Johnson, Thomas A; Sung, Myong-Hee; Escobar, Thelma; Simmons, Catherine A; Pearce, Kenneth H; Biddie, Simon C; Sabo, Pete J; Thurman, Robert E; Stamatoyannopoulos, John A; Hager, Gordon L

    2011-01-01

    Cell-selective glucocorticoid receptor (GR) binding to distal regulatory elements is associated with cell type-specific regions of locally accessible chromatin. These regions can either pre-exist in chromatin (pre-programmed) or be induced by the receptor (de novo). Mechanisms that create and maintain these sites are not well understood. We observe a global enrichment of CpG density for pre-programmed elements, and implicate their demethylated state in the maintenance of open chromatin in a tissue-specific manner. In contrast, sites that are actively opened by GR (de novo) are characterized by low CpG density, and form a unique class of enhancers devoid of suppressive effect of agglomerated methyl-cytosines. Furthermore, treatment with glucocorticoids induces rapid changes in methylation levels at selected CpGs within de novo sites. Finally, we identify GR-binding elements with CpGs at critical positions, and show that methylation can affect GR–DNA interactions in vitro. The findings present a unique link between tissue-specific chromatin accessibility, DNA methylation and transcription factor binding and show that DNA methylation can be an integral component of gene regulation by nuclear receptors. PMID:21701563

  1. Silver nanowire interactions with primary human alveolar type-II epithelial cell secretions: contrasting bioreactivity with human alveolar type-I and type-II epithelial cells

    NASA Astrophysics Data System (ADS)

    Sweeney, Sinbad; Theodorou, Ioannis G.; Zambianchi, Marta; Chen, Shu; Gow, Andrew; Schwander, Stephan; Zhang, Junfeng (Jim); Chung, Kian Fan; Shaffer, Milo S. P.; Ryan, Mary P.; Porter, Alexandra E.; Tetley, Teresa D.

    2015-06-01

    Inhaled nanoparticles have a high deposition rate in the alveolar units of the deep lung. The alveolar epithelium is composed of type-I and type-II epithelial cells (ATI and ATII respectively) and is bathed in pulmonary surfactant. The effect of native human ATII cell secretions on nanoparticle toxicity is not known. We investigated the cellular uptake and toxicity of silver nanowires (AgNWs; 70 nm diameter, 1.5 μm length) with human ATI-like cells (TT1), in the absence or presence of Curosurf® (a natural porcine pulmonary surfactant with a low amount of protein) or harvested primary human ATII cell secretions (HAS; containing both the complete lipid as well as the full protein complement of human pulmonary surfactant i.e. SP-A, SP-B, SP-C and SP-D). We hypothesised that Curosurf® or HAS would confer improved protection for TT1 cells, limiting the toxicity of AgNWs. In agreement with our hypothesis, HAS reduced the inflammatory and reactive oxygen species (ROS)-generating potential of AgNWs with exposed TT1 cells. For example, IL-8 release and ROS generation was reduced by 38% and 29%, respectively, resulting in similar levels to that of the non-treated controls. However in contrast to our hypothesis, Curosurf® had no effect. We found a significant reduction in AgNW uptake by TT1 cells in the presence of HAS but not Curosurf. Furthermore, we show that the SP-A and SP-D are likely to be involved in this process as they were found to be specifically bound to the AgNWs. While ATI cells appear to be protected by HAS, evidence suggested that ATII cells, despite no uptake, were vulnerable to AgNW exposure (indicated by increased IL-8 release and ROS generation and decreased intracellular SP-A levels one day post-exposure). This study provides unique findings that may be important for the study of lung epithelial-endothelial translocation of nanoparticles in general and associated toxicity within the alveolar unit.Inhaled nanoparticles have a high deposition rate in

  2. Cell-type-specific responses of RT4 neural cell lines to dibutyryl-cAMP: branch determination versus maturation

    SciTech Connect

    Droms, K.; Sueoka, N.

    1987-03-01

    This report describes the induction of cell-type-specific maturation, by dibutyryl-cAMP and testololactone, of neuronal and glial properties in a family of cell lines derived from a rat peripheral neurotumor, RT4. This maturation allows further understanding of the process of determination because of the close lineage relationship between the cell types of the RT4 family. The RT4 family is characterized by the spontaneous conversion of one of the cell types, RT4-AC (stem-cell type), to any of three derivative cell types, RT4-B, RT4-D, or RT4-E, with a frequency of about 10(-5). The RT4-AC cells express some properties characteristic of both neuronal and glial cells. Of these neural properties expressed by RT4-AC cells, only the neuronal properties are expressed by the RT4-B and RT4-E cells, and only the glial properties are expressed by the RT4-D cells. This in vitro cell-type conversion of RT4-AC to three derivative cell types is a branch point for the coordinate regulation of several properties and seems to resemble determination in vivo. In our standard culture conditions, several other neuronal and glial properties are not expressed by these cell types. However, addition of dibutyryl-cAMP induces expression of additional properties, in a cell-type-specific manner: formation of long cellular processes in the RT4-B8 and RT4-E5 cell lines and expression of high-affinity uptake of gamma-aminobutyric acid, by a glial-cell-specific mechanism, in the RT4-D6-2 cell line. These new properties are maximally expressed 2-3 days after addition of dibutyryl-cAMP.

  3. Phospholipid-transfer activities in cytosols from lung, isolated alveolar type II cells and alveolar type II cell-derived adenomas.

    PubMed Central

    Pool, G L; Bubacz, D G; Lumb, R H; Mason, R J

    1983-01-01

    We have examined phospholipid-transfer activities in cytosols from rat and mouse whole lung, isolated rat alveolar type II cells and alveolar type II cell-derived mouse pulmonary adenomas. We report an enrichment in phosphatidylcholine and phosphatidylglycerol (but not phosphatidylinositol) protein-catalysed transfer in the type II cell and adenoma cytosols compared with the whole-lung cytosols. The activities from these cytosols were resolved using column chromatofocusing, which clearly demonstrated the presence of a phosphatidylcholine-specific transfer protein in each of the four tissues. In addition, two proteins (rat) or three proteins (mouse) catalysing both phosphatidylcholine and phosphatidylglycerol transfer were resolved from whole lung, whereas in both the rat isolated alveolar type II cells and the mouse type II cell-derived adenomas one of these less specific proteins is not present. PMID:6661189

  4. Cell-cell signalling in sexual chemotaxis: a basis for gametic differentiation, mating types and sexes.

    PubMed

    Hadjivasiliou, Zena; Iwasa, Yoh; Pomiankowski, Andrew

    2015-08-01

    While sex requires two parents, there is no obvious need for them to be differentiated into distinct mating types or sexes. Yet this is the predominate state of nature. Here, we argue that mating types could play a decisive role because they prevent the apparent inevitability of self-stimulation during sexual signalling. We rigorously assess this hypothesis by developing a model for signaller-detector dynamics based on chemical diffusion, chemotaxis and cell movement. Our model examines the conditions under which chemotaxis improves partner finding. Varying parameter values within ranges typical of protists and their environments, we show that simultaneous secretion and detection of a single chemoattractant can cause a multifold movement impediment and severely hinder mate finding. Mutually exclusive roles result in faster pair formation, even when cells conferring the same roles cannot pair up. This arrangement also allows the separate mating types to optimize their signalling or detecting roles, which is effectively impossible for cells that are both secretors and detectors. Our findings suggest that asymmetric roles in sexual chemotaxis (and possibly other forms of sexual signalling) are crucial, even without morphological differences, and may underlie the evolution of gametic differentiation among both mating types and sexes.

  5. Basal cell (monomorphic) and minimally pleomorphic adenomas of the salivary glands. Distinction from the solid (anaplastic) type of adenoid cystic carcinoma in fine-needle aspiration.

    PubMed

    Stanley, M W; Horwitz, C A; Rollins, S D; Powers, C N; Bardales, R H; Korourain, S; Stern, S J

    1996-07-01

    Cytologic features of the cell-stroma interface are useful in distinguishing between monomorphic adenomas of the basal cell type and adenoid cystic carcinoma. In basal cell adenomas, the collagenous stroma interdigitates with adjacent cells, whereas in adenoid cystic carcinoma, the two are separated by a sharp smooth border. Furthermore, the stroma of basal cell adenomas can contain rare spindle cells or capillaries, but the cylinders of adenoid cystic carcinoma are acellular. The authors review their experience with five cases of basal cell adenoma, and three cases that were designated "minimally pleomorphic adenomas." The latter group showed the small blue cell pattern of basal cell adenoma at the time of fine-needle aspiration, and histology revealed only small foci of typical pleomorphic adenoma. With the exception of one cystic case, the cell-stroma interface of basal cell adenoma was observed in all eight cases. These cases are contrasted with three adenoid cystic carcinomas with extensive solid (anaplastic) areas. All showed the small blue cell pattern and cell-stroma interface features of basal cell adenoma. Neither showed the smooth-bordered cylinders of adenoid cystic carcinoma. Two of these three were incorrectly interpreted as benign at the time of fine-needle aspiration. The authors suggest that the stroma aspirated from solid adenoid cystic carcinoma represents desmoplastic tumor stroma that mimics the pattern of basal cell adenoma in smear material. Distinction between basal cell adenoma and the solid type of adenoid cystic carcinoma at the time of fine-needle aspiration remains a very difficult problem.

  6. Cell-type-specific modelling of intracellular calcium signalling: a urothelial cell model.

    PubMed

    Appleby, Peter A; Shabir, Saqib; Southgate, Jennifer; Walker, Dawn

    2013-09-01

    Calcium signalling plays a central role in regulating a wide variety of cell processes. A number of calcium signalling models exist in the literature that are capable of reproducing a variety of experimentally observed calcium transients. These models have been used to examine in more detail the mechanisms underlying calcium transients, but very rarely has a model been directly linked to a particular cell type and experimentally verified. It is important to show that this can be achieved within the general theoretical framework adopted by these models. Here, we develop a framework designed specifically for modelling cytosolic calcium transients in urothelial cells. Where possible, we draw upon existing calcium signalling models, integrating descriptions of components known to be important in this cell type from a number of studies in the literature. We then add descriptions of several additional pathways that play a specific role in urothelial cell signalling, including an explicit ionic influx term and an active pumping mechanism that drives the cytosolic calcium concentration to a target equilibrium. The resulting one-pool model of endoplasmic reticulum (ER)-dependent calcium signalling relates the cytosolic, extracellular and ER calcium concentrations and can generate a wide range of calcium transients, including spikes, bursts, oscillations and sustained elevations in the cytosolic calcium concentration. Using single-variate robustness and multivariate sensitivity analyses, we quantify how varying each of the parameters of the model leads to changes in key features of the calcium transient, such as initial peak amplitude and the frequency of bursting or spiking, and in the transitions between bursting- and plateau-dominated modes. We also show that, novel to our urothelial cell model, the ionic and purinergic P2Y pathways make distinct contributions to the calcium transient. We then validate the model using human bladder epithelial cells grown in monolayer cell

  7. Differentially Expressed miRNAs in Tumor, Adjacent, and Normal Tissues of Lung Adenocarcinoma

    PubMed Central

    Tian, Fei; Li, Rui; Chen, Zhenzhu; Shen, Yanting; Lu, Jiafeng; Xie, Xueying; Ge, Qinyu

    2016-01-01

    Lung cancer is the leading cause of cancer deaths. Non-small-cell lung cancer (NSCLC) is the major type of lung cancer. The aim of this study was to characterize the expression profiles of miRNAs in adenocarcinoma (AC), one major subtype of NSCLC. In this study, the miRNAs were detected in normal, adjacent, and tumor tissues by next-generation sequencing. Then the expression levels of differential miRNAs were quantified by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). In the results, 259, 401, and 389 miRNAs were detected in tumor, adjacent, and normal tissues of pooled AC samples, respectively. In addition, for the first time we have found that miR-21-5p and miR-196a-5p were gradually upregulated from normal to adjacent to tumor tissues; miR-218-5p was gradually downregulated with 2-fold or greater change in AC tissues. These 3 miRNAs were validated by qRT-PCR. Lastly, we predicted target genes of these 3 miRNAs and enriched the potential functions and regulatory pathways. The aberrant miR-21-5p, miR-196a-5p, and miR-218-5p may become biomarkers for diagnosis and prognosis of lung adenocarcinoma. This research may be useful for lung adenocarcinoma diagnosis and the study of pathology in lung cancer. PMID:27247934

  8. Molecular disorganization of axons adjacent to human lacunar infarcts.

    PubMed

    Hinman, Jason D; Lee, Monica D; Tung, Spencer; Vinters, Harry V; Carmichael, S Thomas

    2015-03-01

    Cerebral microvascular disease predominantly affects brain white matter and deep grey matter, resulting in ischaemic damage that ranges from lacunar infarcts to white matter hyperintensities seen on magnetic resonance imaging. These lesions are common and result in both clinical stroke syndromes and accumulate over time, resulting in cognitive deficits and dementia. Magnetic resonance imaging studies suggest that these lesions progress over time, accumulate adjacent to prior lesions and have a penumbral region susceptible to further injury. The pathological correlates of this adjacent injury in surviving myelinated axons have not been previously defined. In this study, we sought to determine the molecular organization of axons in tissue adjacent to lacunar infarcts and in the regions surrounding microinfarcts, by determining critical elements in axonal function: the morphology and length of node of Ranvier segments and adjacent paranodal segments. We examined post-mortem brain tissue from six patients with lacunar infarcts and tissue from two patients with autosomal dominant retinal vasculopathy and cerebral leukoencephalopathy (previously known as hereditary endotheliopathy with retinopathy, nephropathy and stroke) who accumulate progressive white matter ischaemic lesions in the form of lacunar and microinfarcts. In axons adjacent to lacunar infarcts yet extending up to 150% of the infarct diameter away, both nodal and paranodal length increase by ∼20% and 80%, respectively, reflecting a loss of normal cell-cell adhesion and signalling between axons and oligodendrocytes. Using premorbid magnetic resonance images, brain regions from patients with retinal vasculopathy and cerebral leukoencephalopathy that harboured periventricular white matter hyperintensities were selected and the molecular organization of axons was determined within these regions. As in regions adjacent to lacunar infarcts, nodal and paranodal length in white matter of these patients is

  9. Molecular disorganization of axons adjacent to human lacunar infarcts

    PubMed Central

    Lee, Monica D.; Tung, Spencer; Vinters, Harry V.; Carmichael, S. Thomas

    2015-01-01

    Cerebral microvascular disease predominantly affects brain white matter and deep grey matter, resulting in ischaemic damage that ranges from lacunar infarcts to white matter hyperintensities seen on magnetic resonance imaging. These lesions are common and result in both clinical stroke syndromes and accumulate over time, resulting in cognitive deficits and dementia. Magnetic resonance imaging studies suggest that these lesions progress over time, accumulate adjacent to prior lesions and have a penumbral region susceptible to further injury. The pathological correlates of this adjacent injury in surviving myelinated axons have not been previously defined. In this study, we sought to determine the molecular organization of axons in tissue adjacent to lacunar infarcts and in the regions surrounding microinfarcts, by determining critical elements in axonal function: the morphology and length of node of Ranvier segments and adjacent paranodal segments. We examined post-mortem brain tissue from six patients with lacunar infarcts and tissue from two patients with autosomal dominant retinal vasculopathy and cerebral leukoencephalopathy (previously known as hereditary endotheliopathy with retinopathy, nephropathy and stroke) who accumulate progressive white matter ischaemic lesions in the form of lacunar and microinfarcts. In axons adjacent to lacunar infarcts yet extending up to 150% of the infarct diameter away, both nodal and paranodal length increase by ∼20% and 80%, respectively, reflecting a loss of normal cell-cell adhesion and signalling between axons and oligodendrocytes. Using premorbid magnetic resonance images, brain regions from patients with retinal vasculopathy and cerebral leukoencephalopathy that harboured periventricular white matter hyperintensities were selected and the molecular organization of axons was determined within these regions. As in regions adjacent to lacunar infarcts, nodal and paranodal length in white matter of these patients is

  10. Haptoglobin directly affects T cells and suppresses T helper cell type 2 cytokine release

    PubMed Central

    Arredouani, M; Matthijs, P; Van Hoeyveld, E; Kasran, A; Baumann, H; Ceuppens, J L; Stevens, E

    2003-01-01

    T helper cell type 1 (Th1) and type 2 (Th2) immune responses are characterized by a different pattern of cytokine expression following T-cell activation. Alterations of the ratio of Th1 to Th2 cells are important determinants of susceptibility to viral and parasitic infections, allergies, anti-tumour responses, and autoimmunity. In this work we bring new evidence for an effect of haptoglobin (Hp), a positive acute-phase protein, on T-lymphocyte functions. We show that Hp specifically interacts with both resting and activated CD4+ and CD8+ T cells. This specific binding results in a strong suppression of induced T-cell proliferation. In addition, Hp exhibits a strong in vitro inhibitory effect on Th2 cytokine release, while the production of interferon-γ (IFN-γ) and interleukin-2 (IL-2) is only slightly inhibited at high Hp doses. As a result, the presence of Hp promotes Th1 activation over Th2 activation in vivo as evidenced in Hp-deficient mice. Anti-CD3 monoclonal antibody injection indeed resulted in predominant IL-4 production in Hp−/− mice, in contrast to predominant IFN-γ production in Hp+/+ mice. We conclude that Hp plays a modulating role on the Th1/Th2 balance by promoting a dominant Th1 cellular response. This points to a role of acute-phase proteins in balancing immune responses. PMID:12562322

  11. Decipher the Molecular Response of Plant Single Cell Types to Environmental Stresses

    SciTech Connect

    Nourbakhsh-Rey, Mehrnoush; Libault, Marc

    2016-01-01

    The analysis of the molecular response of entire plants or organs to environmental stresses suffers from the cellular complexity of the samples used. Specifically, this cellular complexity masks cell-specific responses to environmental stresses and logically leads to the dilution of the molecular changes occurring in each cell type composing the tissue/organ/plant in response to the stress. Therefore, to generate a more accurate picture of these responses, scientists are focusing on plant single cell type approaches. Several cell types are now considered as models such as the pollen, the trichomes, the cotton fiber, various root cell types including the root hair cell, and the guard cell of stomata. Among them, several have been used to characterize plant response to abiotic and biotic stresses. Lastly, in this review, we are describing the various -omic studies performed on these different plant single cell type models to better understand plant cell response to biotic and abiotic stresses.

  12. Decipher the Molecular Response of Plant Single Cell Types to Environmental Stresses

    PubMed Central

    2016-01-01

    The analysis of the molecular response of entire plants or organs to environmental stresses suffers from the cellular complexity of the samples used. Specifically, this cellular complexity masks cell-specific responses to environmental stresses and logically leads to the dilution of the molecular changes occurring in each cell type composing the tissue/organ/plant in response to the stress. Therefore, to generate a more accurate picture of these responses, scientists are focusing on plant single cell type approaches. Several cell types are now considered as models such as the pollen, the trichomes, the cotton fiber, various root cell types including the root hair cell, and the guard cell of stomata. Among them, several have been used to characterize plant response to abiotic and biotic stresses. In this review, we are describing the various -omic studies performed on these different plant single cell type models to better understand plant cell response to biotic and abiotic stresses. PMID:27088086

  13. Decipher the Molecular Response of Plant Single Cell Types to Environmental Stresses.

    PubMed

    Nourbakhsh-Rey, Mehrnoush; Libault, Marc

    2016-01-01

    The analysis of the molecular response of entire plants or organs to environmental stresses suffers from the cellular complexity of the samples used. Specifically, this cellular complexity masks cell-specific responses to environmental stresses and logically leads to the dilution of the molecular changes occurring in each cell type composing the tissue/organ/plant in response to the stress. Therefore, to generate a more accurate picture of these responses, scientists are focusing on plant single cell type approaches. Several cell types are now considered as models such as the pollen, the trichomes, the cotton fiber, various root cell types including the root hair cell, and the guard cell of stomata. Among them, several have been used to characterize plant response to abiotic and biotic stresses. In this review, we are describing the various -omic studies performed on these different plant single cell type models to better understand plant cell response to biotic and abiotic stresses. PMID:27088086

  14. Phase I Trial of Adoptive Cell Transfer with Mixed-Profile Type-I/Type-II Allogeneic T Cells for Metastatic Breast Cancer

    PubMed Central

    Hardy, Nancy M.; Mossoba, Miriam E.; Steinberg, Seth M.; Fellowes, Vicki; Yan, Xiao-Yi; Hakim, Frances T.; Babb, Rebecca R.; Avila, Daniele; Gea-Banacloche, Juan; Sportès, Claude; Levine, Bruce L.; June, Carl H.; Khuu, Hahn M.; Carpenter, Ashley E.; Krumlauf, Michael C.; Dwyer, Andrew J.; Gress, Ronald E.; Fowler, Daniel H.; Bishop, Michael R.

    2011-01-01

    PURPOSE Metastatic breast cancer (MBC) response to allogeneic lymphocytes requires donor T-cell engraftment and is limited by graft-versus-host disease (GVHD). In mice, Type-II-polarized T cells promote engraftment and modulate GVHD whereas Type-I-polarized T cells mediate more potent graft-versus-tumor (GVT) effects. This Phase-I translational study evaluated adoptive transfer of ex-vivo-costimulated Type-I/Type-II (T1/T2) donor T cells with T-cell-depleted (TCD) allogeneic stem-cell transplantation (AlloSCT) for MBC. EXPERIMENTAL DESIGN Patients had received anthracycline, taxane and antibody therapies, been treated for metastatic disease and an HLA-identical-sibling donor. Donor lymphocytes were costimulated ex vivo with anti-CD3/anti-CD28 antibody-coated magnetic beads in IL-2/IL-4-supplemented media. Patients received reduced-intensity conditioning, donor stem cells and T1/T2 cells, and monitoring for toxicity, engraftment, GVHD and tumor response; results were compared with historical controls, identically treated except for T1/T2-product infusions. RESULTS Mixed Type-I/Type-II CD4+-T cells predominated in T1/T2 products. Nine patients received T1/T2 cells at Dose-Level 1 (5×106 cells/kg). T-cell donor chimerism reached 100% by a median of 28 days. Seven (78%) developed acute GVHD. At Day +28, five patients had partial responses (56%) and none had MBC progression; thereafter, two patients had continued responses. Donor-T-cell engraftment and tumor responses appeared faster than in historical controls, but GVHD rates were similar and responders progressed early, often following treatment of acute GVHD. CONCLUSION Allogeneic T1/T2 cells were safely infused with TCD-AlloSCT, appeared to promote donor engraftment, and may have contributed to transient early tumor responses. PMID:21948234

  15. Replication of parainfluenza (Sendai) virus in isolated rat pulmonary type II alveolar epithelial cells.

    PubMed Central

    Castleman, W. L.; Northrop, P. J.; McAllister, P. K.

    1989-01-01

    The major objectives of this study were to determine whether alveolar type II epithelial cells isolated from rat lung and maintained in tissue culture would support productive replication of parainfluenza type 1 (Sendai) virus and to determine whether isolated type II cells from neonatal (5-day-old) rats that are more susceptible to viral-induced alveolar dysplasia supported viral replication to a greater extent than those from weanling (25-day-old) rats. Isolated and cultured type II cells from neonatal and weanling rats that were inoculated with Sendai virus supported productive replication as indicated by ultrastructural identification of budding virions and viral nucleocapsids in type II cells and by demonstration of rising titers of infectious virus from inoculated type II cell cultures. Alveolar macrophages from neonatal and weanling rats also supported viral replication, although infectious viral titers in macrophage cultures were lower than those from type II cell cultures. Only minor differences were detected between viral titers from neonatal and weanling type II epithelial cell cultures. Higher densities of viral nucleocapsids were observed in neonatal type II cells than in those from weanling rats. The results indicate that isolated type II alveolar epithelial cells support productive replication of parainfluenza virus and that type II cells are probably more efficient in supporting productive viral replication than are alveolar macrophages. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:2541612

  16. Rabbit somatic cell cloning: effects of donor cell type, histone acetylation status and chimeric embryo complementation.

    PubMed

    Yang, Feikun; Hao, Ru; Kessler, Barbara; Brem, Gottfried; Wolf, Eckhard; Zakhartchenko, Valeri

    2007-01-01

    The epigenetic status of a donor nucleus has an important effect on the developmental potential of embryos produced by somatic cell nuclear transfer (SCNT). In this study, we transferred cultured rabbit cumulus cells (RCC) and fetal fibroblasts (RFF) from genetically marked rabbits (Alicia/Basilea) into metaphase II oocytes and analyzed the levels of histone H3-lysine 9-lysine 14 acetylation (acH3K9/14) in donor cells and cloned embryos. We also assessed the correlation between the histone acetylation status of donor cells and cloned embryos and their developmental potential. To test whether alteration of the histone acetylation status affects development of cloned embryos, we treated donor cells with sodium butyrate (NaBu), a histone deacetylase inhibitor. Further, we tried to improve cloning efficiency by chimeric complementation of cloned embryos with blastomeres from in vivo fertilized or parthenogenetic embryos. The levels of acH3K9/14 were higher in RCCs than in RFFs (P<0.05). Although the type of donor cells did not affect development to blastocyst, after transfer into recipients, RCC cloned embryos induced a higher initial pregnancy rate as compared to RFF cloned embryos (40 vs 20%). However, almost all pregnancies with either type of cloned embryos were lost by the middle of gestation and only one fully developed, live RCC-derived rabbit was obtained. Treatment of RFFs with NaBu significantly increased the level of acH3K9/14 and the proportion of nuclear transfer embryos developing to blastocyst (49 vs 33% with non-treated RFF, P<0.05). The distribution of acH3K9/14 in either group of cloned embryos did not resemble that in in vivo fertilized embryos suggesting that reprogramming of this epigenetic mark is aberrant in cloned rabbit embryos and cannot be corrected by treatment of donor cells with NaBu. Aggregation of embryos cloned from NaBu-treated RFFs with blastomeres from in vivo derived embryos improved development to blastocyst, but no cloned

  17. Brain structure. Cell types in the mouse cortex and hippocampus revealed by single-cell RNA-seq.

    PubMed

    Zeisel, Amit; Muñoz-Manchado, Ana B; Codeluppi, Simone; Lönnerberg, Peter; La Manno, Gioele; Juréus, Anna; Marques, Sueli; Munguba, Hermany; He, Liqun; Betsholtz, Christer; Rolny, Charlotte; Castelo-Branco, Gonçalo; Hjerling-Leffler, Jens; Linnarsson, Sten

    2015-03-01

    The mammalian cerebral cortex supports cognitive functions such as sensorimotor integration, memory, and social behaviors. Normal brain function relies on a diverse set of differentiated cell types, including neurons, glia, and vasculature. Here, we have used large-scale single-cell RNA sequencing (RNA-seq) to classify cells in the mouse somatosensory cortex and hippocampal CA1 region. We found 47 molecularly distinct subclasses, comprising all known major cell types in the cortex. We identified numerous marker genes, which allowed alignment with known cell types, morphology, and location. We found a layer I interneuron expressing Pax6 and a distinct postmitotic oligodendrocyte subclass marked by Itpr2. Across the diversity of cortical cell types, transcription factors formed a complex, layered regulatory code, suggesting a mechanism for the maintenance of adult cell type identity.

  18. Aberrant Methylation Inactivates Somatostatin and Somatostatin Receptor Type 1 in Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Misawa, Kiyoshi; Misawa, Yuki; Kondo, Haruki; Mochizuki, Daiki; Imai, Atsushi; Fukushima, Hirofumi; Uehara, Takayuki; Kanazawa, Takeharu; Mineta, Hiroyuki

    2015-01-01

    Purpose The aim of this study was to define somatostatin (SST) and somatostatin receptor type 1 (SSTR1) methylation profiles for head and neck squamous cell carcinoma (HNSCC) tumors at diagnosis and follow up and to evaluate their prognostic significance and value as a biomarker. Methods Gene expression was measured by quantitative RT-PCR. Promoter methylation status was determined by quantitative methylation-specific PCR (Q-MSP) in HNSCC. Results Methylation was associated with transcription inhibition. SST methylation in 81% of HNSCC tumor specimens significantly correlated with tumor size (P = 0.043), stage (P = 0.008), galanin receptor type 2 (GALR2) methylation (P = 0.041), and tachykinin-1 (TAC1) (P = 0.040). SSTR1 hypermethylation in 64% of cases was correlated with tumor size (P = 0.037), stage (P = 0.037), SST methylation (P < 0.001), and expression of galanin (P = 0.03), GALR2 (P = 0.014), TAC1 (P = 0.023), and tachykinin receptor type 1 (TACR1) (P = 0.003). SST and SSTR1 promoter hypermethylation showed highly discriminating receiver operator characteristic curve profiles, which clearly distinguished HNSCC from adjacent normal mucosal tissues. Concurrent hypermethylation of galanin and SSTR1 promoters correlated with reduced disease-free survival (log-rank test, P = 0.0001). Among patients with oral cavity and oropharynx cancer, methylation of both SST and SSTR1 promoters correlated with reduced disease-free survival (log-rank test, P = 0.028). In multivariate logistic-regression analysis, concomitant methylation of galanin and SSTR1 was associated with an odds ratio for recurrence of 12.53 (95% CI, 2.62 to 59.8; P = 0.002). Conclusions CpG hypermethylation is a likely mechanism of SST and SSTR1 gene inactivation, supporting the hypothesis that SST and SSTR1 play a role in the tumorigenesis of HNSCC and that this hypermethylation may serve as an important biomarker. PMID:25734919

  19. Type II pneumocytes in mixed cell culture of human lung: a light and electron microscopic study.

    PubMed Central

    Bingle, L; Bull, T B; Fox, B; Guz, A; Richards, R J; Tetley, T D

    1990-01-01

    Alveolar Type II epithelial cells dedifferentiate rapidly in vitro. Studies with animal tissue suggest that cell-cell and extracellular matrix-cell interactions are important in the retention of Type II cell morphology in vitro. Thus, in this study with human tissue, alveolar Type II cells, alveolar macrophages, and spindle cells were prepared from the same sample of lung (obtained following lobectomy for cancer, n = 3), cocultured on glass cover slips or tissue culture plastic, and studied by light microscopy with scanning (SEM) and transmission (TEM) electron microscopy for 8 days. The primary cell isolates contained approximately 45% Type II cells; the remainder were macrophages or unidentifiable cells. Clusters, made up of a single layer of cuboidal Type II cells around a central core of connective tissue (largely collagen and some elastic tissue), formed above a monolayer of spindle cells. The Type II cells were morphologically similar to those seen in vivo. The cells were still cuboidal at 8 days but had lost their lamellar bodies, which were released into the medium via the apical surface. The clusters increased in size with time (area, microns 2: day 1, 29(5-143) x 10(2); day 8, 63(10-311) x 10(2); mean(range); p less than 0.02) without changing in number per culture, suggesting Type II cell proliferation. This may have been due to factors produced by the other cells and adherence to the extracellular matrix (ECM); (free collagen fibers, present in the original preparation, spindle cells, and/or Type II cells could be responsible for presence of ECM). We propose this as a useful model for the study of human Type II epithelial cells in vitro. Images FIGURE 1. a FIGURE 1. b FIGURE 1. c FIGURE 1. d FIGURE 1. e FIGURE 1. f FIGURE 2. a FIGURE 2. b FIGURE 2. c FIGURE 2. d FIGURE 2. e FIGURE 2. f FIGURE 2. g FIGURE 3. PMID:2384069

  20. Cell type-specific responses of peripheral blood mononuclear cells to silver nanoparticles.

    PubMed

    Greulich, C; Diendorf, J; Gessmann, J; Simon, T; Habijan, T; Eggeler, G; Schildhauer, T A; Epple, M; Köller, M

    2011-09-01

    Silver nanoparticles (Ag-NP) are increasingly used in biomedical applications because of their remarkable antimicrobial activity. In biomedicine, Ag-NP are coated onto or embedded in wound dressings, surgical instruments and bone substitute biomaterials, such as silver-containing calcium phosphate cements. Free Ag-NP and silver ions are released from these coatings or after the degradation of a biomaterial, and may come into close contact with blood cells. Despite the widespread use of Ag-NP as an antimicrobial agent, there is a serious lack of information on the biological effects of Ag-NP on human blood cells. In this study, the uptake of Ag-NP by peripheral monocytes and lymphocytes (T-cells) was analyzed, and the influence of nanosilver on cell biological functions (proliferation, the expression of adhesion molecules, cytokine release and the generation of reactive oxygen species) was studied. After cell culture in the presence of monodispersed Ag-NP (5-30μgml(-1) silver concentration), agglomerates of nanoparticles were detected within monocytes (CD14+) but not in T-cells (CD3+) by light microscopy, flow cytometry and combined focused ion beam/scanning electron microscopy. The uptake rate of nanoparticles was concentration dependent, and the silver agglomerates were typically found in the cytoplasm. Furthermore, a concentration-dependent activation (e.g. an increased expression of adhesion molecule CD54) of monocytes at Ag-NP concentrations of 10-15μgml(-1) was observed, and cytotoxicity of Ag-NP-treated monocytes was observed at Ag-NP levels of 25μgml(-1) and higher. However, no modulation of T-cell proliferation was observed in the presence of Ag-NP. Taken together, our results provide the first evidence for a cell-type-specific uptake of Ag-NP by peripheral blood mononuclear cells (PBMC) and the resultant cellular responses after exposure.

  1. Steviol Glycosides Modulate Glucose Transport in Different Cell Types

    PubMed Central

    Rizzo, Benedetta; Zambonin, Laura; Leoncini, Emanuela; Vieceli Dalla Sega, Francesco; Prata, Cecilia; Fiorentini, Diana; Hrelia, Silvana

    2013-01-01

    Extracts from Stevia rebaudiana Bertoni, a plant native to Central and South America, have been used as a sweetener since ancient times. Currently, Stevia extracts are largely used as a noncaloric high-potency biosweetener alternative to sugar, due to the growing incidence of type 2 diabetes mellitus, obesity, and metabolic disorders worldwide. Despite the large number of studies on Stevia and steviol glycosides in vivo, little is reported concerning the cellular and molecular mechanisms underpinning the beneficial effects on human health. The effect of four commercial Stevia extracts on glucose transport activity was evaluated in HL-60 human leukaemia and in SH-SY5Y human neuroblastoma cells. The extracts were able to enhance glucose uptake in both cellular lines, as efficiently as insulin. Our data suggest that steviol glycosides could act by modulating GLUT translocation through the PI3K/Akt pathway since treatments with both insulin and Stevia extracts increased the phosphorylation of PI3K and Akt. Furthermore, Stevia extracts were able to revert the effect of the reduction of glucose uptake caused by methylglyoxal, an inhibitor of the insulin receptor/PI3K/Akt pathway. These results corroborate the hypothesis that Stevia extracts could mimic insulin effects modulating PI3K/Akt pathway. PMID:24327825

  2. Genistein increases glycosaminoglycan levels in mucopolysaccharidosis type I cell models.

    PubMed

    Kingma, Sandra D K; Wagemans, Tom; IJlst, Lodewijk; Wijburg, Frits A; van Vlies, Naomi

    2014-09-01

    Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder characterized by diminished degradation of the glycosaminoglycans (GAGs) heparan sulfate and dermatan sulfate, which results in the accumulation of these GAGs and subsequent cellular dysfunction. Patients present with a variety of symptoms, including severe skeletal disease. Genistein has been shown previously to inhibit GAG synthesis in MPS fibroblasts, presumably through inhibition of tyrosine kinase activity of the epidermal growth factor receptor (EGFR). To determine the potentials of genistein for the treatment of skeletal disease, MPS I fibroblasts were induced into chondrocytes and osteoblasts and treated with genistein. Surprisingly, whereas tyrosine phosphorylation levels (as a measure for tyrosine kinase inhibition) were decreased in all treated cell lines, there was a 1.3 and 1.6 fold increase in GAG levels in MPS I chondrocytes and fibroblast, respectively (p < 0.05). Sulfate incorporation in treated MPS I fibroblasts was 2.6 fold increased (p < 0.05), indicating increased GAG synthesis despite tyrosine kinase inhibition. This suggests that GAG synthesis is not exclusively regulated through the tyrosine kinase activity of the EGFR. We hypothesize that the differences in outcomes between studies on the effect of genistein in MPS are caused by the different effects of genistein on different growth factor signaling pathways, which regulate GAG synthesis. More studies are needed to elucidate the precise signaling pathways which are affected by genistein and alter GAG metabolism in order to evaluate the therapeutic potential of genistein for MPS patients. PMID:24699889

  3. Types of HLA in the bladder transitional cell carcinoma (TCC).

    PubMed

    Yılmaz, Erkan; Uğur Özalp, Ali; Cekmen, Arman; Eren, Bülent; Onal, Bülent; Akkuş, Emre; Erdoğan, Ergun

    2013-02-01

    HLA plays a complementary role in the interaction between tumor and body immunology. The aim of this study was to determine the existence of the association between the HLA system and transitional cell carcinoma (TCC). Using standard micro-lymphocytotoxic method of Terasaki, HLA-A, B, DR and DQ antigen types of 30 patients with TCC of the bladder were compared with the control group (30 healthy people). In the TCC patient group, HLA -DQ6(1) and HLA -DQ7(3) antigens were detected with a significantly higher frequency than in the control group (p=0.018 and p=0.038, respectively), whereas HLA-A10, B4, DR53 and DQ1 antigens were detected with significantly higher frequency in the control group (p less 0.05 in all). It suggests that patients who had the antigens detected were at higher risk of TCC, and the ones who had the antigens displaying protective features as were detected in the control group, were at lesser risk.

  4. Geochronology of cave deposits at Liang Bua and of adjacent river terraces in the Wae Racang valley, western Flores, Indonesia: a synthesis of age estimates for the type locality of Homo floresiensis.

    PubMed

    Roberts, R G; Westaway, K E; Zhao, J-x; Turney, C S M; Bird, M I; Rink, W J; Fifield, L K

    2009-11-01

    A robust timeframe for the extant cave deposits at Liang Bua, and for the river terraces in the adjoining Wae Racang valley, is essential to constrain the period of existence and time of extinction of Homo floresiensis and other biota that have been excavated at this hominin type locality. Reliable age control is also required for the variety of artifacts excavated from these deposits, and to assist in environmental reconstructions for this river valley and for the region more broadly. In this paper, we summarize the available geochronological information for Liang Bua and its immediate environs, obtained using seven numerical-age methods: radiocarbon, thermoluminescence, optically- and infrared-stimulated luminescence (collectively known as optical dating), uranium-series, electron spin resonance, and coupled electron spin resonance/uranium-series. We synthesize the large number of numerical age determinations reported previously and present additional age estimates germane to questions of hominin evolution and extinction.

  5. HCMM imagery for the discrimination of rock types, the detection of geothermal energy sources and the assessment of soil moisture content in western Queensland and adjacent parts of New South Wales and South Australia

    NASA Technical Reports Server (NTRS)

    Cole, M. M. (Principal Investigator)

    1980-01-01

    The author has identified the following significant results. Day-visible and day-IR imagery of northwest Queensland show that large scale geological features like the Mitakoodi anticlinorium, which involves rocks of contrasting lithological type, can be delineated. North of Cloncurry, the contrasting lithological units of the Knapdale quartzite and bedded argillaceous limestones within the Proterozoic Corella sequence are clearly delineated in the area of the Dugald River Lode. Major structural features in the Mount Isa area are revealed on the day-visible cover. Which provides similar but less detailed information than the LANDSAT imagery. The day-IR cover provides less additional information for areas of outcropping bedrock than had been expected. Initial studies of the day-IR and night-IR cover for parts of South Australia suggest that they contain additional information on geology compared with day-visible cover.

  6. Oxygen sensing in neuroendocrine cells and other cell types: pheochromocytoma (PC12) cells as an experimental model.

    PubMed

    Spicer, Zachary; Millhorn, David E

    2003-01-01

    A steady supply of oxygen is an absolute requirement for mammalian cells to maintain normal cellular functions. To answer the challenge that oxygen deprivation represents, mammals have evolved specialized cell types that can sense changes in oxygen tension and alter gene expression to enhance oxygen delivery to hypoxic areas. These oxygensensing cells are rare and difficult to study in vivo. As a result, pheochromocytoma (PC12) cells have become a vital in vitro model system for deciphering the molecular events that confer the hypoxia-resistant and oxygen-sensing phenotypes. Research over the last few years has revealed that the hypoxia response in PC12 cells involves the interactions of several signal transduction pathways (Ca2+/calmodulin-dependent kinases, Akt, SAPKs, and MAPKs) and transcription factors (HIFs, CREB, and c-fos/junB). This review summarizes the current understanding of the role these signal transduction pathways and transcription factors play in determining the hypoxic response. PMID:14739486

  7. Development of both human connective tissue-type and mucosal-type mast cells in mice from hematopoietic stem cells with identical distribution pattern to human body.

    PubMed

    Kambe, Naotomo; Hiramatsu, Hidefumi; Shimonaka, Mika; Fujino, Hisanori; Nishikomori, Ryuta; Heike, Toshio; Ito, Mamoru; Kobayashi, Kimio; Ueyama, Yoshito; Matsuyoshi, Norihisa; Miyachi, Yoshiki; Nakahata, Tatsutoshi

    2004-02-01

    The transplantation of primitive human cells into sublethally irradiated immune-deficient mice is the well-established in vivo system for the investigation of human hematopoietic stem cell function. Although mast cells are the progeny of hematopoietic stem cells, human mast cell development in mice that underwent human hematopoietic stem cell transplantation has not been reported. Here we report on human mast cell development after xenotransplantation of human hematopoietic stem cells into nonobese diabetic severe combined immunodeficient (NOD/SCID)/gamma(c)(null) (NOG) mice with severe combined immunodeficiency and interleukin 2 (IL-2) receptor gamma-chain allelic mutation. Supported by the murine environment, human mast cell clusters developed in mouse dermis, but they required more time than other forms of human cell reconstitution. In lung and gastric tract, mucosal-type mast cells containing tryptase but lacking chymase located on gastric mucosa and in alveoli, whereas connective tissue-type mast cells containing both tryptase and chymase located on gastric submucosa and around major airways, as in the human body. Mast cell development was also observed in lymph nodes, spleen, and peritoneal cavity but not in the peripheral blood. Xenotransplantation of human hematopoietic stem cells into NOG mice can be expected to result in a highly effective model for the investigation of human mast cell development and function in vivo.

  8. Placental phagocytic cells infected with herpes simplex type 2 and echovirus type 19: virological and ultrastructural aspects.

    PubMed

    Oliveira, L H; Fonseca, M E; De Bonis, M

    1992-01-01

    Placental macrophage cells were kept in a short-term culture and infected with herpes simplex type 2 virus and echovirus type 19. These were observed under optical and electron microscopy. Immunofluorescence, virus titration and autoradiographic technique were used to determine if the virus was replicating in the system. The results showed that the placental phagocytic cells do not allow virus growth and that the virus particles are destroyed right after virus uptake, within 4 h post-infection. The increase of lipid bodies and other cellular alterations suggested the intensive action of these cells against viruses.

  9. General approach for in vivo recovery of cell type-specific effector gene sets.

    PubMed

    Barsi, Julius C; Tu, Qiang; Davidson, Eric H

    2014-05-01

    Differentially expressed, cell type-specific effector gene sets hold the key to multiple important problems in biology, from theoretical aspects of developmental gene regulatory networks (GRNs) to various practical applications. Although individual cell types of interest have been recovered by various methods and analyzed, systematic recovery of multiple cell type-specific gene sets from whole developing organisms has remained problematic. Here we describe a general methodology using the sea urchin embryo, a material of choice because of the large-scale GRNs already solved for this model system. This method utilizes the regulatory states expressed by given cells of the embryo to define cell type and includes a fluorescence activated cell sorting (FACS) procedure that results in no perturbation of transcript representation. We have extensively validated the method by spatial and qualitative analyses of the transcriptome expressed in isolated embryonic skeletogenic cells and as a consequence, generated a prototypical cell type-specific transcriptome database.

  10. General approach for in vivo recovery of cell type-specific effector gene sets

    PubMed Central

    Barsi, Julius C.; Tu, Qiang; Davidson, Eric H.

    2014-01-01

    Differentially expressed, cell type-specific effector gene sets hold the key to multiple important problems in biology, from theoretical aspects of developmental gene regulatory networks (GRNs) to various practical applications. Although individual cell types of interest have been recovered by various methods and analyzed, systematic recovery of multiple cell type-specific gene sets from whole developing organisms has remained problematic. Here we describe a general methodology using the sea urchin embryo, a material of choice because of the large-scale GRNs already solved for this model system. This method utilizes the regulatory states expressed by given cells of the embryo to define cell type and includes a fluorescence activated cell sorting (FACS) procedure that results in no perturbation of transcript representation. We have extensively validated the method by spatial and qualitative analyses of the transcriptome expressed in isolated embryonic skeletogenic cells and as a consequence, generated a prototypical cell type-specific transcriptome database. PMID:24604781

  11. BK-Type K(Ca) channels in two parasympathetic cell types: differences in kinetic properties and developmental expression.

    PubMed

    Cameron, J S; Dryer, S E

    2000-12-01

    The intrinsic electrical properties of identified choroid and ciliary neurons of the chick ciliary ganglion were examined by patch-clamp recording methods. These neurons are derived from a common pool of mesencephalic neural crest precursor cells but innervate different target tissues and have markedly different action potential waveforms and intrinsic patterns of repetitive spike discharge. Therefore it is important to determine whether these cell types express different types of plasma membrane ionic channels, and to ascertain the developmental stages at which these cell types begin to diverge. This study has focused on large-conductance Ca(2+)-activated K(+) channels (K(Ca)), which are known to regulate spike waveform and repetitive firing in many cell types. Both ciliary ganglion cell types, identified on the basis of size and somatostatin immunoreactivity, express a robust macroscopic K(Ca) carried by a kinetically homogeneous population of large-conductance (BK-type) K(Ca) channels. However, the kinetic properties of these channels are different in the two cell types. Steady-state fluctuation analyses of macroscopic K(Ca) produced power spectra that could be fitted with a single Lorentzian curve in both cell types. However, the resulting corner frequency was significantly lower in choroid neurons than in ciliary neurons, suggesting that the underlying K(Ca) channels have a longer mean open-time in choroid neurons. Consistent with fluctuation analyses, significantly slower gating of K(Ca) channels in choroid neurons was also observed during macroscopic activation and deactivation at membrane potentials positive to -30 mV. Differences in the kinetic properties of K(Ca) channels could also be observed directly in single-channel recordings from identified embryonic day 13 choroid and ciliary neurons. The mean open-time of large-conductance K(Ca) channels was significantly greater in choroid neurons than in ciliary neurons in excised inside-out patches. The

  12. Cell type-specific, presynaptic LTP of inhibitory synapses on fast-spiking GABAergic neurons in the mouse visual cortex.

    PubMed

    Sarihi, Abdolrahman; Mirnajafi-Zadeh, Javad; Jiang, Bin; Sohya, Kazuhiro; Safari, Mir-Shahram; Arami, Masoumeh Kourosh; Yanagawa, Yuchio; Tsumoto, Tadaharu

    2012-09-19

    Properties and plasticity of inhibitory synapses on fast-spiking (FS) GABAergic (FS-GABA) interneurons in layer II/III of the mouse visual cortex were examined in cortical slices by whole-cell recordings of IPSCs or IPSPs evoked by activation of presynaptic FS or non-FS GABAergic interneurons. Unitary IPSCs (uIPSCs) evoked by action potentials of FS-GABA neurons have shorter onset latency, faster rising slope, higher peak amplitude, and faster decay time than those evoked by action potentials of non-FS-GABA neurons. Tetanic activation of presynaptic FS-GABA neurons induced long-term potentiation (LTP) of uIPSCs, whereas that of presynaptic non-FS-GABA neurons did not induce LTP, indicating that long-term plasticity of inhibitory synapses on FS-GABA neurons is pathway specific. For further analysis of inhibitory synaptic plasticity, IPSPs evoked by electrical stimulation of an adjacent site in the cortex were recorded from FS-GABA neurons. Theta burst stimulation induced LTP of IPSPs in 12 of 14 FS-GABA neurons. The paired-pulse stimulation protocol and coefficient of variation analysis indicated that this form of LTP may be presynaptic in origin. Filling postsynaptic cells with a Ca(2+) chelator did not block the induction of LTP, suggesting no involvement of postsynaptic Ca(2+) rise. Also, this form of LTP was dependent neither on metabotropic glutamate receptors nor voltage-gated Ca(2+) channels of the L and T types. Further pharmacological analysis indicated that voltage-gated Ca(2+) channels other than the P/Q type, such as N and R types, were not involved in LTP, suggesting that P/Q-type channels are a candidate for factors inducing LTP of inhibitory synapses between FS-GABA neurons. PMID:22993435

  13. Alveolar epithelial type I cells contain transport proteins and transport sodium, supporting an active role for type I cells in regulation of lung liquid homeostasis

    PubMed Central

    Johnson, Meshell D.; Widdicombe, Jonathan H.; Allen, Lennell; Barbry, Pascal; Dobbs, Leland G.

    2002-01-01

    Transport of lung liquid is essential for both normal pulmonary physiologic processes and for resolution of pathologic processes. The large internal surface area of the lung is lined by alveolar epithelial type I (TI) and type II (TII) cells; TI cells line >95% of this surface, TII cells <5%. Fluid transport is regulated by ion transport, with water movement following passively. Current concepts are that TII cells are the main sites of ion transport in the lung. TI cells have been thought to provide only passive barrier, rather than active, functions. Because TI cells line most of the internal surface area of the lung, we hypothesized that TI cells could be important in the regulation of lung liquid homeostasis. We measured both Na+ and K+ (Rb+) transport in TI cells isolated from adult rat lungs and compared the results to those of concomitant experiments with isolated TII cells. TI cells take up Na+ in an amiloride-inhibitable fashion, suggesting the presence of Na+ channels; TI cell Na+ uptake, per microgram of protein, is ≈2.5 times that of TII cells. Rb+ uptake in TI cells was ≈3 times that in TII cells and was inhibited by 10−4 M ouabain, the latter observation suggesting that TI cells exhibit Na+-, K+-ATPase activity. By immunocytochemical methods, TI cells contain all three subunits (α, β, and γ) of the epithelial sodium channel ENaC and two subunits of Na+-, K+-ATPase. By Western blot analysis, TI cells contain ≈3 times the amount of αENaC/μg protein of TII cells. Taken together, these studies demonstrate that TI cells not only contain molecular machinery necessary for active ion transport, but also transport ions. These results modify some basic concepts about lung liquid transport, suggesting that TI cells may contribute significantly in maintaining alveolar fluid balance and in resolving airspace edema. PMID:11842214

  14. Type I and type II interferon responses in two human liver cell lines (Huh-7 and HuH6)

    PubMed Central

    Grünvogel, Oliver; Esser-Nobis, Katharina; Windisch, Marc P.; Frese, Michael; Trippler, Martin; Bartenschlager, Ralf; Lohmann, Volker; Binder, Marco

    2015-01-01

    Most studies investigating the biology of Hepatitis C virus (HCV) have used the human hepatoma cell line Huh-7 or subclones thereof, as these are the most permissive cell lines for HCV infection and replication. Other cell lines also support replication of HCV, most notably the human hepatoblastoma cell line HuH6. HCV replication in cell culture is generally highly sensitive to interferons (IFNs) and differences in the IFN-mediated inhibition of virus replication may reflect alterations in the IFN-induced antiviral response inherent to different host cells. For example, HCV replication is highly sensitive to IFN-γ treatment in Huh-7, but not in HuH6 cells. In this study, we used microarray-based gene expression profiling to compare the response of Huh-7 and HuH6 cells to stimulation with IFN-α and IFN-γ. Furthermore, we determined whether the resistance of HCV replication in HuH6 cells can be linked to differences in the expression profile of IFN-regulated genes. Although both cells lines responded to IFNs with rapid changes in gene expression, thereby demonstrating functional type I and type II signaling pathways, differences were observed for a number of genes. Raw and normalized expression data have been deposited in GEO under accession number GSE68927. PMID:26981398

  15. 20. Interior view of fuel storage pit or vault adjacent ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    20. Interior view of fuel storage pit or vault adjacent to Test Cell 9 in Component Test Laboratory (T-27), looking west. Photograph shows upgraded instrumentation, piping, tanks, and technological modifications installed in 1997-99 to accommodate component testing requirements for the Atlas V missile. - Air Force Plant PJKS, Systems Integration Laboratory, Components Test Laboratory, Waterton Canyon Road & Colorado Highway 121, Lakewood, Jefferson County, CO

  16. Beta-cell function and mass in type 2 diabetes.

    PubMed

    Larsen, Marianne O

    2009-08-01

    The aim of the work described here was to improve our understanding of beta-cell function (BCF) and beta-cell mass (BCM) and their relationship in vivo using the minipig as a model for some of the aspects of human type 2 diabetes (T2DM). More specifically, the aim was to evaluate the following questions: How is BCF, especially high frequency pulsatile insulin secretion, affected by a primary reduction in BCM or by primary obesity or a combination of the two in the minipig? Can evaluation of BCF in vivo be used as a surrogate measure to predict BCM in minipigs over a range of BCM and body weight? We first developed a minipig model of reduced BCM and mild diabetes using administration of a combination of streptozotocin (STZ) and nicotinamide (NIA) as a tool to study effects of a primary reduction of BCM on BCF. The model was characterized using a mixed-meal oral glucose tolerance test and intravenous stimulation with glucose and arginine as well as by histology of the pancreas after euthanasia. It was shown that stable, moderate diabetes can be induced and that the model is characterized by fasting and postprandial hyperglycemia, reduced insulin secretion and reduced BCM. Several defects in insulin secretion are well documented in human T2DM; however, the role in the pathogenesis and the possible clinical relevance of high frequency (rapid) pulsatile insulin secretion is still debated. We therefore investigated this phenomenon in normal minipigs and found easily detectable pulses in peripheral vein plasma samples that were shown to be correlated with pulses found in portal vein plasma. Furthermore, the rapid kinetics of insulin in the minipig strongly facilitates pulse detection. These characteristics make the minipig particularly suitable for studying the occurrence of disturbed pulsatility in relation to T2DM. Disturbances of rapid pulsatile insulin secretion have been reported to be a very early event in the development of T2DM and include disorderliness of pulses

  17. Beta-cell function and mass in type 2 diabetes.

    PubMed

    Larsen, Marianne O

    2009-08-01

    The aim of the work described here was to improve our understanding of beta-cell function (BCF) and beta-cell mass (BCM) and their relationship in vivo using the minipig as a model for some of the aspects of human type 2 diabetes (T2DM). More specifically, the aim was to evaluate the following questions: How is BCF, especially high frequency pulsatile insulin secretion, affected by a primary reduction in BCM or by primary obesity or a combination of the two in the minipig? Can evaluation of BCF in vivo be used as a surrogate measure to predict BCM in minipigs over a range of BCM and body weight? We first developed a minipig model of reduced BCM and mild diabetes using administration of a combination of streptozotocin (STZ) and nicotinamide (NIA) as a tool to study effects of a primary reduction of BCM on BCF. The model was characterized using a mixed-meal oral glucose tolerance test and intravenous stimulation with glucose and arginine as well as by histology of the pancreas after euthanasia. It was shown that stable, moderate diabetes can be induced and that the model is characterized by fasting and postprandial hyperglycemia, reduced insulin secretion and reduced BCM. Several defects in insulin secretion are well documented in human T2DM; however, the role in the pathogenesis and the possible clinical relevance of high frequency (rapid) pulsatile insulin secretion is still debated. We therefore investigated this phenomenon in normal minipigs and found easily detectable pulses in peripheral vein plasma samples that were shown to be correlated with pulses found in portal vein plasma. Furthermore, the rapid kinetics of insulin in the minipig strongly facilitates pulse detection. These characteristics make the minipig particularly suitable for studying the occurrence of disturbed pulsatility in relation to T2DM. Disturbances of rapid pulsatile insulin secretion have been reported to be a very early event in the development of T2DM and include disorderliness of pulses

  18. The 3 major types of innate and adaptive cell-mediated effector immunity.

    PubMed

    Annunziato, Francesco; Romagnani, Chiara; Romagnani, Sergio

    2015-03-01

    The immune system has tailored its effector functions to optimally respond to distinct species of microbes. Based on emerging knowledge on the different effector T-cell and innate lymphoid cell (ILC) lineages, it is clear that the innate and adaptive immune systems converge into 3 major kinds of cell-mediated effector immunity, which we propose to categorize as type 1, type 2, and type 3. Type 1 immunity consists of T-bet(+) IFN-γ-producing group 1 ILCs (ILC1 and natural killer cells), CD8(+) cytotoxic T cells (TC1), and CD4(+) TH1 cells, which protect against intracellular microbes through activation of mononuclear phagocytes. Type 2 immunity consists of GATA-3(+) ILC2s, TC2 cells, and TH2 cells producing IL-4, IL-5, and IL-13, which induce mast cell, basophil, and eosinophil activation, as well as IgE antibody production, thus protecting against helminthes and venoms. Type 3 immunity is mediated by retinoic acid-related orphan receptor γt(+) ILC3s, TC17 cells, and TH17 cells producing IL-17, IL-22, or both, which activate mononuclear phagocytes but also recruit neutrophils and induce epithelial antimicrobial responses, thus protecting against extracellular bacteria and fungi. On the other hand, type 1 and 3 immunity mediate autoimmune diseases, whereas type 2 responses can cause allergic diseases.

  19. Coordinating cell proliferation and differentiation: Antagonism between cell cycle regulators and cell type-specific gene expression

    PubMed Central

    Ruijtenberg, Suzan; van den Heuvel, Sander

    2016-01-01

    ABSTRACT Cell proliferation and differentiation show a remarkable inverse relationship. Precursor cells continue division before acquiring a fully differentiated state, while terminal differentiation usually coincides with proliferation arrest and permanent exit from the division cycle. Mechanistic insight in the temporal coordination between cell cycle exit and differentiation has come from studies of cells in culture and genetic animal models. As initially described for skeletal muscle differentiation, temporal coordination involves mutual antagonism between cyclin-dependent kinases that promote cell cycle entry and transcription factors that induce tissue-specific gene expression. Recent insights highlight the contribution of chromatin-regulating complexes that act in conjunction with the transcription factors and determine their activity. In particular SWI/SNF chromatin remodelers contribute to dual regulation of cell cycle and tissue-specific gene expression during terminal differentiation. We review the concerted regulation of the cell cycle and cell type-specific transcription, and discuss common mutations in human cancer that emphasize the clinical importance of proliferation versus differentiation control. PMID:26825227

  20. Advances in understanding the cell types and approaches used for generating induced pluripotent stem cells

    PubMed Central

    2014-01-01

    Successfully reprogramming somatic cells to a pluripotent state generates induced pluripotent stem (iPS) cells (or iPSCs), which have extensive self-renewal capacity like embryonic stem cells (ESCs). iPSCs can also generate daughter cells that can further undergo differentiation into various lineages or terminally differentiate to reach their final functional state. The discovery of how to produce iPSCs opened a new field of stem cell research with both intellectual and therapeutic benefits. The huge potential implications of disease-specific or patient-specific iPSCs have impelled scientists to solve problems hindering their applications in clinical medicine, especially the issues of convenience and safety. To determine the range of tissue types amenable to reprogramming as well as their particular characteristics, cells from three embryonic germ layers have been assessed, and the advantages that some tissue origins have over fibroblast origins concerning efficiency and accessibility have been elucidated. To provide safe iPSCs in an efficient and convenient way, the delivery systems and combinations of inducing factors as well as the chemicals used to generate iPSCs have also been significantly improved in addition to the efforts on finding better donor cells. Currently, iPSCs can be generated without c-Myc and Klf4 oncogenes, and non-viral delivery integration-free chemically mediated reprogramming methods have been successfully employed with relatively satisfactory efficiency. This paper will review recent advances in iPS technology by highlighting tissue origin and generation of iPSCs. The obstacles that need to be overcome for clinical applications of iPSCs are also discussed. PMID:25037625

  1. Advances in understanding the cell types and approaches used for generating induced pluripotent stem cells.

    PubMed

    Li, Jun; Song, Wei; Pan, Guangjin; Zhou, Jun

    2014-01-01

    Successfully reprogramming somatic cells to a pluripotent state generates induced pluripotent stem (iPS) cells (or iPSCs), which have extensive self-renewal capacity like embryonic stem cells (ESCs). iPSCs can also generate daughter cells that can further undergo differentiation into various lineages or terminally differentiate to reach their final functional state. The discovery of how to produce iPSCs opened a new field of stem cell research with both intellectual and therapeutic benefits. The huge potential implications of disease-specific or patient-specific iPSCs have impelled scientists to solve problems hindering their applications in clinical medicine, especially the issues of convenience and safety. To determine the range of tissue types amenable to reprogramming as well as their particular characteristics, cells from three embryonic germ layers have been assessed, and the advantages that some tissue origins have over fibroblast origins concerning efficiency and accessibility have been elucidated. To provide safe iPSCs in an efficient and convenient way, the delivery systems and combinations of inducing factors as well as the chemicals used to generate iPSCs have also been significantly improved in addition to the efforts on finding better donor cells. Currently, iPSCs can be generated without c-Myc and Klf4 oncogenes, and non-viral delivery integration-free chemically mediated reprogramming methods have been successfully employed with relatively satisfactory efficiency. This paper will review recent advances in iPS technology by highlighting tissue origin and generation of iPSCs. The obstacles that need to be overcome for clinical applications of iPSCs are also discussed.

  2. Alternative gene expression in type I and type II cells may enable further nuclear changes during conjugation of Blepharisma japonicum.

    PubMed

    Sugiura, Mayumi; Tanaka, Yuri; Suzaki, Toshinobu; Harumoto, Terue

    2012-03-01

    In contrast to most ciliates, meiosis and successive nuclear changes during conjugation occur only in heterotypic pairs in Blepharisma. It has been suggested that homotypic pairs are ready for conjugation, but lack a trigger to initiate the nuclear changes, and the conjugation process is arrested before the onset of meiosis. To explore the possible nature of the trigger, we previously identified the genes BjCdk1 (homologous to cdk1/cdc2), Bj4HPPD (4-hydroxy-phenylpyruvate dioxygenase) and BjCks (cyclin dependent kinase regulatory subunit) whose expression is up-regulated in gamone1-treated type II cells. In this study, we investigated the molecular structures of these three genes, and compared their expression patterns in homotypic and heterotypic pairs, finding remarkable differences. BjCdk1, Bj4HPPD and BjCks were expressed specifically in gamone1-treated type II cells, but not in gamone2-treated type I cells. In heterotypic pairs, the expression of these genes stayed at the same level or gradually decreased throughout the entire process of conjugation, but it rapidly decreased and ceased after 10hours in homotypic pairs. These results indicate that some genes are expressed in a mating-type specific manner. Alternative gene expression in mating type I and type II cells and merging of individual factors in a heterotypic pair may induce nuclear changes including meiosis.

  3. Cell-Type-Specific Genome-wide Expression Profiling after Laser Capture Microdissection of Living Tissue

    SciTech Connect

    Marchetti, F; Manohar, C F

    2005-02-09

    The purpose of this technical feasibility study was to develop and evaluate robust microgenomic tools for investigations of genome-wide expression of very small numbers of cells isolated from whole tissue sections. Tissues contain large numbers of cell-types that play varied roles in organ function and responses to endogenous and exogenous toxicants whether bacterial, viral, chemical or radiation. Expression studies of whole tissue biopsy are severely limited because heterogeneous cell-types result in an averaging of molecular signals masking subtle but important changes in gene expression in any one cell type(s) or group of cells. Accurate gene expression analysis requires the study of specific cell types in their tissue environment but without contamination from surrounding cells. Laser capture microdissection (LCM) is a new technology to isolate morphologically distinct cells from tissue sections. Alternative methods are available for isolating single cells but not yet for their reliable genome-wide expression analyses. The tasks of this feasibility project were to: (1) Develop efficient protocols for laser capture microdissection of cells from tissues identified by antibody label, or morphological stain. (2) Develop reproducible gene-transcript analyses techniques for single cell-types and determine the numbers of cells needed for reliable genome-wide analyses. (3) Validate the technology for epithelial and endothelial cells isolated from the gastrointestinal tract of mice.

  4. Photovoltaic Cell Having A P-Type Polycrystalline Layer With Large Crystals

    DOEpatents

    Albright, Scot P.; Chamberlin, Rhodes R.

    1996-03-26

    A photovoltaic cell has an n-type polycrystalline layer and a p-type polycrystalline layer adjoining the n-type polycrystalline layer to form a photovoltaic junction. The p-type polycrystalline layer comprises a substantially planar layer portion having relatively large crystals adjoining the n-type polycrystalline layer. The planar layer portion includes oxidized impurities which contribute to obtainment of p-type electrical properties in the planar layer portion.

  5. Stimulation of DNA synthesis in cultured rat alveolar type II cells

    SciTech Connect

    Leslie, C.C.; McCormick-Shannon, K.; Robinson, P.C.; Mason, R.J.

    1985-01-01

    Restoration of the alveolar epithelium after injury is thought to be dependent on the proliferation of alveolar type II cells. To understand the factors that may be involved in promoting type II cell proliferation in vivo, we determined the effect of potential mitogens and culture substrata on DNA synthesis in rat alveolar type II cells in primary culture. Type II cells cultured in basal medium containing 10% fetal bovine serum (FBS) exhibited essentially no DNA synthesis. Factors that stimulated /sup 3/H-thymidine incorporation included cholera toxin, epidermal growth factor, and rat serum. The greatest degree of stimulation was achieved by plating type II cells on an extracellular matrix prepared from bovine corneal endothelial cells and then by culturing the pneumocytes in medium containing rat serum, cholera toxin, insulin, and epidermal growth factor. Under conditions of stimulation of /sup 3/H-thymidine incorporation there was an increased DNA content per culture dish but no increase in cell number. The ability of various culture conditions to promote DNA synthesis in type II cells was verified by autoradiography. Type II cells were identified by the presence of cytoplasmic inclusions, which were visualized by tannic acid staining before autoradiography. These results demonstrate the importance of soluble factors and culture substratum in stimulating DNA synthesis in rat alveolar type II cells in primary culture.

  6. Calcium signaling and T-type calcium channels in cancer cell cycling

    PubMed Central

    Taylor, James T; Zeng, Xiang-Bin; Pottle, Jonathan E; Lee, Kevin; Wang, Alun R; Yi, Stephenie G; Scruggs, Jennifer A S; Sikka, Suresh S; Li, Ming

    2008-01-01

    Regulation of intracellular calcium is an important signaling mechanism for cell proliferation in both normal and cancerous cells. In normal epithelial cells, free calcium concentration is essential for cells to enter and accomplish the S phase and the M phase of the cell cycle. In contrast, cancerous cells can pass these phases of the cell cycle with much lower cytoplasmic free calcium concentrations, indicating an alternative mechanism has developed for fulfilling the intracellular calcium requirement for an increased rate of DNA synthesis and mitosis of fast replicating cancerous cells. The detailed mechanism underlying the altered calcium loading pathway remains unclear; however, there is a growing body of evidence that suggests the T-type Ca2+ channel is abnormally expressed in cancerous cells and that blockade of these channels may reduce cell proliferation in addition to inducing apoptosis. Recent studies also show that the expression of T-type Ca2+ channels in breast cancer cells is proliferation state dependent, i.e. the channels are expressed at higher levels during the fast-replication period, and once the cells are in a non-proliferation state, expression of this channel is minimal. Therefore, selectively blocking calcium entry into cancerous cells may be a valuable approach for preventing tumor growth. Since T-type Ca2+ channels are not expressed in epithelial cells, selective T-type Ca2+ channel blockers may be useful in the treatment of certain types of cancers. PMID:18763278

  7. Rat alveolar type I cells proliferate, express OCT-4, and exhibit phenotypic plasticity in vitro.

    PubMed

    Gonzalez, Robert F; Allen, Lennell; Dobbs, Leland G

    2009-12-01

    Alveolar type I (TI) cells are large, squamous cells that cover 95-99% of the internal surface area of the lung. Although TI cells are believed to be terminally differentiated, incapable of either proliferation or phenotypic plasticity, TI cells in vitro both proliferate and express phenotypic markers of other differentiated cell types. Rat TI cells isolated in purities of >99% proliferate in culture, with a sixfold increase in cell number before the cells reach confluence; >50% of the cultured TI cells are Ki67+. At cell densities of 1-2 cells/well, approximately 50% of the cells had the capacity to form colonies. Under the same conditions, type II cells do not proliferate. Cultured TI cells express RTI40 and aquaporin 5, phenotypic markers of the TI cell phenotype. By immunofluorescence, Western blotting, and Q-PCR, TI cells express OCT-4A (POU5F1), a transcription factor associated with maintenance of the pluripotent state in stem cells. Based on the expression patterns of various marker proteins, TI cells are distinct from either of two recently described putative pulmonary multipotent cell populations, the bronchoalveolar stem cell or the OCT-4+ stem/progenitor cell. Although TI cells in adult rat lung tissue do not express either surfactant protein C (SP-C) or CC10, respective markers of the TII and Clara cell phenotypes, in culture TI cells can be induced to express both SP-C and CC10. Together, the findings that TI cells proliferate and exhibit phenotypic plasticity in vitro raise the possibility that TI cells may have similar properties in vivo. PMID:19717550

  8. Differences in the expression of chromosome 1 genes between lung telocytes and other cells: mesenchymal stem cells, fibroblasts, alveolar type II cells, airway epithelial cells and lymphocytes

    PubMed Central

    Sun, Xiaoru; Zheng, Minghuan; Zhang, Miaomiao; Qian, Mengjia; Zheng, Yonghua; Li, Meiyi; Cretoiu, Dragos; Chen, Chengshui; Chen, Luonan; Popescu, Laurentiu M; Wang, Xiangdong

    2014-01-01

    Telocytes (TCs) are a unique type of interstitial cells with specific, extremely long prolongations named telopodes (Tps). Our previous study showed that TCs are distinct from fibroblasts (Fbs) and mesenchymal stem cells (MSCs) as concerns gene expression and proteomics. The present study explores patterns of mouse TC-specific gene profiles on chromosome 1. We investigated the network of main genes and the potential functional correlations. We compared gene expression profiles of mouse pulmonary TCs, MSCs, Fbs, alveolar type II cells (ATII), airway basal cells (ABCs), proximal airway cells (PACs), CD8+ T cells from bronchial lymph nodes (T-BL) and CD8+ T cells from lungs (T-LL). The functional and feature networks were identified and compared by bioinformatics tools. Our data showed that on TC chromosome 1, there are about 25% up-regulated and 70% down-regulated genes (more than onefold) as compared with the other cells respectively. Capn2, Fhl2 and Qsox1 were over-expressed in TCs compared to the other cells, indicating that biological functions of TCs are mainly associated with morphogenesis and local tissue homoeostasis. TCs seem to have important roles in the prevention of tissue inflammation and fibrogenesis development in lung inflammatory diseases and as modulators of immune cell response. In conclusion, TCs are distinct from the other cell types. PMID:24826900

  9. Anti-tumour necrosis factor treatment increases circulating T helper type 17 cells similarly in different types of inflammatory arthritis

    PubMed Central

    Hull, D N; Williams, R O; Pathan, E; Alzabin, S; Abraham, S; Taylor, P C

    2015-01-01

    We investigated changes in circulating T helper type 17 (Th17) cells following anti-tumour necrosis factor (TNF) in rheumatoid arthritis (RA), ankylosing spondylitis (AS) and psoriatic arthritis (PsA) patients. Peripheral blood mononuclear cells (PBMC) were isolated from 25 RA, 15 AS and eight PsA patients at baseline 4 and 12 weeks after treatment, and Th17 cell frequencies were analysed using interleukin (IL)-17 enzyme-linked immunospot (ELISPOT) and flow cytometry. A significant increase in IL-17-producing cells was observed by ELISPOT in RA and AS patients at 12 weeks. Flow cytometry confirmed significant increases in CD4+IL-17+ cells at 12 weeks in RA and AS and 4 weeks in PsA patients. Anti-TNF treatment increases circulating Th17 cells in three different diseases. PMID:25766640

  10. Development of a shingle-type solar cell module

    NASA Technical Reports Server (NTRS)

    Shepard, N. F., Jr.; Sanchez, L. E.

    1978-01-01

    The development of a solar cell module, which is suitable for use in place of shingles on the sloping roofs of residental or commercial buildings, is reported. The design consists of nineteen series-connected 53 mm diameter solar cells arranged in a closely packed hexagon configuration. The shingle solar cell module consists of two basic functional parts: an exposed rigid portion which contains the solar cell assembly, and a semi-flexible portion which is overlapped by the higher courses of the roof installation. Consideration is given to the semi-flexible substrate configuration and solar cell and module-to-module interconnectors. The results of an electrical performance analysis are given and it is noted that high specific power output can be attributed to the efficient packing of the circular cells within the hexagon shape. The shingle should function for at least 15 years, with a specific power output of 98 W/sq w.

  11. A "hotspot" for autoimmune T cells in type 1 diabetes.

    PubMed

    Stadinski, Brian D; Obst, Reinhard; Huseby, Eric S

    2016-06-01

    The ability of a single T cell antigen receptor (TCR) to cross-react with multiple antigens allows the finite number of T cells within an organism to respond to the compendium of pathogen challenges faced during a lifetime. Effective immune surveillance, however, comes at a price. TCR cross-reactivity can allow molecular mimics to spuriously activate autoimmune T cells; it also underlies T cell rejection of organ transplants and drives graft-versus-host disease. In this issue of the JCI, Cole and colleagues provide insight into how an insulin-reactive T cell cross-reacts with pathogen-derived antigens by focusing on a limited portion of the peptides to provide a hotspot for binding. These findings dovetail with recent studies of alloreactive and autoimmune TCRs and suggest that the biochemical principles that govern conventional protein-protein interactions may allow the specificity and cross-reactivity profiles of T cells to be predicted. PMID:27183386

  12. Adenovirus type 2 expresses fiber in monkey-human hybrids and reconstructed cells

    SciTech Connect

    Zorn, G.A.; Anderson, C.W.

    1981-02-01

    Adenovirus type 2 protein expression was measured by indirect immunofluorescence in monkey-human hybrids and in cells reconstructed from monkey and human cell karyoplasts and cytoplasts. Monkey-human hybrid clones infected with adenovirus type 2 expressed fiber protein, whereas infected monkey cells alone did not. Hybrids constructed after the parental monkey cells were infected with adenovirus type 2 demonstrated that fiber synthesis in these cells could be rescued by fusion to uninfected human cells. Thus, human cells contain a dominant factor that acts in trans and overcomes the inability of monkey cells to synthesize fiber. These results are consistent with the hypothesis that the block to adenovirus replication in monkey cells involves a nuclear event that prevents the formation of functional mRNA for some late viral proteins including fiber polypeptide.

  13. Immune polarization by hookworms: taking cues from T helper type 2, type 2 innate lymphoid cells and alternatively activated macrophages.

    PubMed

    Nair, Meera G; Herbert, De'Broski R

    2016-06-01

    Cellular and molecular investigation of parasitic helminth infections has greatly accelerated the understanding of type 2 immune responses. However, there remains considerable debate regarding the specific leucocytes that kill parasites and whether these mechanisms are distinct from those responsible for tissue repair. Herein, we chronicle discoveries over the past decade highlighting current paradigms in type 2 immunity with a particular emphasis upon how CD4(+) T helper type 2 cells, type 2 innate lymphoid cells and alternatively activated macrophages coordinately control helminth-induced parasitism. Primarily, this review will draw from studies of the murine nematode parasite Nippostrongylus brasiliensis, which bears important similarities to the human hookworms Ancylostoma duodenale and Necator americanus. Given that one or more hookworm species currently infect millions of individuals across the globe, we propose that vaccine and/or pharmaceutical-based cure strategies targeting these affected human populations should incorporate the conceptual advances outlined herein. PMID:26928141

  14. On the time-course of adjacent and non-adjacent transposed-letter priming

    PubMed Central

    Ktori, Maria; Kingma, Brechtsje; Hannagan, Thomas; Holcomb, Phillip J.; Grainger, Jonathan

    2014-01-01

    We compared effects of adjacent (e.g., atricle-ARTICLE) and non-adjacent (e.g., actirle-ARTICLE) transposed-letter (TL) primes in an ERP study using the sandwich priming technique. TL priming was measured relative to the standard double-substitution condition. We found significantly stronger priming effects for adjacent transpositions than non-adjacent transpositions (with 2 intervening letters) in behavioral responses (lexical decision latencies), and the adjacent priming effects emerged earlier in the ERP signal, at around 200 ms post-target onset. Non-adjacent priming effects emerged about 50 ms later and were short-lived, being significant only in the 250-300 ms time-window. Adjacent transpositions on the other hand continued to produce priming in the N400 time-window (300-500 ms post-target onset). This qualitatively different pattern of priming effects for adjacent and non-adjacent transpositions is discussed in the light of different accounts of letter transposition effects, and the utility of drawing a distinction between positional flexibility and positional noise. PMID:25364497

  15. Cell-type specific gene expression profiles of leukocytes in human peripheral blood

    PubMed Central

    Palmer, Chana; Diehn, Maximilian; Alizadeh, Ash A; Brown, Patrick O

    2006-01-01

    Background Blood is a complex tissue comprising numerous cell types with distinct functions and corresponding gene expression profiles. We attempted to define the cell type specific gene expression patterns for the major constituent cells of blood, including B-cells, CD4+ T-cells, CD8+ T-cells, lymphocytes and granulocytes. We did this by comparing the global gene expression profiles of purified B-cells, CD4+ T-cells, CD8+ T-cells, granulocytes, and lymphocytes using cDNA microarrays. Results Unsupervised clustering analysis showed that similar cell populations from different donors share common gene expression profiles. Supervised analyses identified gene expression signatures for B-cells (427 genes), T-cells (222 genes), CD8+ T-cells (23 genes), granulocytes (411 genes), and lymphocytes (67 genes). No statistically significant gene expression signature was identified for CD4+ cells. Genes encoding cell surface proteins were disproportionately represented among the genes that distinguished among the lymphocyte subpopulations. Lymphocytes were distinguishable from granulocytes based on their higher levels of expression of genes encoding ribosomal proteins, while granulocytes exhibited characteristic expression of various cell surface and inflammatory proteins. Conclusion The genes comprising the cell-type specific signatures encompassed many of the genes already known to be involved in cell-type specific processes, and provided clues that may prove useful in discovering the functions of many still unannotated genes. The most prominent feature of the cell type signature genes was the enrichment of genes encoding cell surface proteins, perhaps reflecting the importance of specialized systems for sensing the environment to the physiology of resting leukocytes. PMID:16704732

  16. Cell type-specific affinity purification of nuclei for chromatin profiling in whole animals.

    PubMed

    Steiner, Florian A; Henikoff, Steven

    2015-01-01

    Analyzing cell differentiation during development in a complex organism requires the analysis of expression and chromatin profiles in individual cell types. Our laboratory has developed a simple and generally applicable strategy to purify specific cell types from whole organisms for simultaneous analysis of chromatin and expression. The method, termed INTACT for Isolation of Nuclei TAgged in specific Cell Types, depends on the expression of an affinity-tagged nuclear envelope protein in the cell type of interest. These nuclei can be affinity-purified from the total pool of nuclei and used as a source for RNA and chromatin. The method serves as a simple and scalable alternative to FACS sorting or laser capture microscopy to circumvent the need for expensive equipment and specialized skills. This chapter provides detailed protocols for the cell-type specific purification of nuclei from Caenorhabditis elegans.

  17. Heterogeneity of stromal cells in the human splenic white pulp. Fibroblastic reticulum cells, follicular dendritic cells and a third superficial stromal cell type

    PubMed Central

    Steiniger, Birte S; Wilhelmi, Verena; Seiler, Anja; Lampp, Katrin; Stachniss, Vitus

    2014-01-01

    At least three phenotypically and morphologically distinguishable types of branched stromal cells are revealed in the human splenic white pulp by subtractive immunohistological double-staining. CD271 is expressed in fibroblastic reticulum cells of T-cell zones and in follicular dendritic cells of follicles. In addition, there is a third CD271− and CD271+/− stromal cell population surrounding T-cell zones and follicles. At the surface of follicles the third population consists of individually variable partially overlapping shells of stromal cells exhibiting CD90 (Thy-1), MAdCAM-1, CD105 (endoglin), CD141 (thrombomodulin) and smooth muscle α-actin (SMA) with expression of CD90 characterizing the broadest shell and SMA the smallest. In addition, CXCL12, CXCL13 and CCL21 are also present in third-population stromal cells and/or along fibres. Not only CD27+ and switched B lymphocytes, but also scattered IgD++ B lymphocytes and variable numbers of CD4+ T lymphocytes often occur close to the third stromal cell population or one of its subpopulations at the surface of the follicles. In contrast to human lymph nodes, neither podoplanin nor RANKL (CD254) were detected in adult human splenic white pulp stromal cells. The superficial stromal cells of the human splenic white pulp belong to a widespread cell type, which is also found at the surface of red pulp arterioles surrounded by a mixed T-cell/B-cell population. Superficial white pulp stromal cells differ from fibroblastic reticulum cells and follicular dendritic cells not only in humans, but apparently also in mice and perhaps in rats. However, the phenotype of white pulp stromal cells is species-specific and more heterogeneous than described so far. PMID:24890772

  18. β-cell-specific CD8 T cell phenotype in type 1 diabetes reflects chronic autoantigen exposure

    PubMed Central

    McLaren, James E.; Dolton, Garry; Matthews, Katherine K.; Gostick, Emma; Kronenberg-Versteeg, Deborah; Eichmann, Martin; Knight, Robin R.; Heck, Susanne; Powrie, Jake; Bingley, Polly J.; Dayan, Colin M.; Miles, John J.; Sewell, Andrew K.

    2015-01-01

    Autoreactive CD8 T cells play a central role in the destruction of pancreatic islet β-cells that leads to type 1 diabetes, yet the key features of this immune-mediated process remain poorly defined. In this study, we combined high definition polychromatic flow cytometry with ultrasensitive peptide-human leukocyte antigen class I (pHLAI) tetramer staining to quantify and characterize β-cell-specific CD8 T cell populations in patients with recent onset type 1 diabetes and healthy controls. Remarkably, we found that β-cell-specific CD8 T cell frequencies in peripheral blood were similar between subject groups. In contrast to healthy controls, however, patients with newly diagnosed type 1 diabetes displayed hallmarks of antigen-driven expansion uniquely within the β-cell-specific CD8 T cell compartment. Molecular analysis of selected β-cell-specific CD8 T cell populations further revealed highly skewed oligoclonal T cell receptor (TCR) repertoires comprising exclusively private clonotypes. Collectively, these data identify novel and distinctive features of disease-relevant CD8 T cells that inform the immunopathogenesis of type 1 diabetes. PMID:25249579

  19. β-cell-specific CD8 T cell phenotype in type 1 diabetes reflects chronic autoantigen exposure.

    PubMed

    Skowera, Ania; Ladell, Kristin; McLaren, James E; Dolton, Garry; Matthews, Katherine K; Gostick, Emma; Kronenberg-Versteeg, Deborah; Eichmann, Martin; Knight, Robin R; Heck, Susanne; Powrie, Jake; Bingley, Polly J; Dayan, Colin M; Miles, John J; Sewell, Andrew K; Price, David A; Peakman, Mark

    2015-03-01

    Autoreactive CD8 T cells play a central role in the destruction of pancreatic islet β-cells that leads to type 1 diabetes, yet the key features of this immune-mediated process remain poorly defined. In this study, we combined high-definition polychromatic flow cytometry with ultrasensitive peptide-human leukocyte antigen class I tetramer staining to quantify and characterize β-cell-specific CD8 T cell populations in patients with recent-onset type 1 diabetes and healthy control subjects. Remarkably, we found that β-cell-specific CD8 T cell frequencies in peripheral blood were similar between subject groups. In contrast to healthy control subjects, however, patients with newly diagnosed type 1 diabetes displayed hallmarks of antigen-driven expansion uniquely within the β-cell-specific CD8 T cell compartment. Molecular analysis of selected β-cell-specific CD8 T cell populations further revealed highly skewed oligoclonal T cell receptor repertoires comprising exclusively private clonotypes. Collectively, these data identify novel and distinctive features of disease-relevant CD8 T cells that inform the immunopathogenesis of type 1 diabetes. PMID:25249579

  20. β-cell-specific CD8 T cell phenotype in type 1 diabetes reflects chronic autoantigen exposure.

    PubMed

    Skowera, Ania; Ladell, Kristin; McLaren, James E; Dolton, Garry; Matthews, Katherine K; Gostick, Emma; Kronenberg-Versteeg, Deborah; Eichmann, Martin; Knight, Robin R; Heck, Susanne; Powrie, Jake; Bingley, Polly J; Dayan, Colin M; Miles, John J; Sewell, Andrew K; Price, David A; Peakman, Mark

    2015-03-01

    Autoreactive CD8 T cells play a central role in the destruction of pancreatic islet β-cells that leads to type 1 diabetes, yet the key features of this immune-mediated process remain poorly defined. In this study, we combined high-definition polychromatic flow cytometry with ultrasensitive peptide-human leukocyte antigen class I tetramer staining to quantify and characterize β-cell-specific CD8 T cell populations in patients with recent-onset type 1 diabetes and healthy control subjects. Remarkably, we found that β-cell-specific CD8 T cell frequencies in peripheral blood were similar between subject groups. In contrast to healthy control subjects, however, patients with newly diagnosed type 1 diabetes displayed hallmarks of antigen-driven expansion uniquely within the β-cell-specific CD8 T cell compartment. Molecular analysis of selected β-cell-specific CD8 T cell populations further revealed highly skewed oligoclonal T cell receptor repertoires comprising exclusively private clonotypes. Collectively, these data identify novel and distinctive features of disease-relevant CD8 T cells that inform the immunopathogenesis of type 1 diabetes.

  1. Perlecan Diversely Regulates the Migration and Proliferation of Distinct Cell Types in vitro.

    PubMed

    Nakamura, Ryosuke; Nakamura, Fumio; Fukunaga, Shigeharu

    2015-01-01

    Perlecan is a multifunctional component of the extracellular matrix. It shows different effects on distinct cell types, and therefore it is thought to show potential for therapies targeting multiple cell types. However, the full range of multifunctionality of perlecan remains to be elucidated. We cultured various cell types, which were derived from epithelial/endothelial, connective and muscle tissues, in the presence of either antiserum against perlecan or exogenous perlecan, and examined the effects of perlecan on cell migration and proliferation. Cell migration was determined using a scratch assay. Blocking of perlecan by anti-perlecan antiserum inhibited the migration of vascular endothelial cells (VECs) and bone marrow-derived mesenchymal stem cells, and exogenous perlecan added to the culture medium promoted the migration of these cell types. The migration of other cell types was inhibited or was not promoted by exogenous perlecan. Cell proliferation was measured using a water-soluble tetrazolium dye. When cells were cultured at low densities, perlecan blocking inhibited the proliferation of VECs, and exogenous perlecan promoted the proliferation of keratinocytes. In contrast, the proliferation of fibroblasts, pre-adipocytes and vascular smooth muscle cells cultured at low densities was inhibited by exogenous perlecan. When cells were cultured at high densities, perlecan blocking promoted the proliferation of most cell types, with the exception of skeletal system-derived cells (chondrocytes and osteoblasts), which were inhibited by exogenous perlecan. Our results provide an overview of the multiple functions of perlecan in various cell types, and implicate a potential role of perlecan to inhibit undesirable activities, such as fibrosis, obesity and intimal hyperplasia.

  2. Effects of Cell Type and Culture Media on Interleukin-6 Secretion in Response to Environmental Particles

    PubMed Central

    Veranth, John M.; Cutler, N. Shane; Kaser, Erin G.; Reilly, Christopher A.; Yost, Garold S.

    2008-01-01

    Cultured lung cells provide an alternative to animal exposures for comparing the effects of different types of air pollution particles. Studies of particulate matter in vitro have reported proinflammatory cytokine signaling in response to many types of environmental particles, but there have been few studies comparing identical treatments in multiple cell types or identical cells with alternative cell culture protocols. We compared soil-derived, diesel, coal fly ash, titanium dioxide, and kaolin particles along with soluble vanadium and lipopolysaccharide, applied to airway-derived cells grown in submerged culture. Cell types included A549, BEAS-2B, RAW 264.7, and primary macrophages. The cell culture models (specific combinations of cell types and culture conditions) were reproducibly different in the cytokine signaling responses to the suite of treatments. Further, Interleukin-6 (IL-6) response to the treatments changed when the same cells, BEAS-2B, were grown in KGM versus LHC-9 media or in media containing bovine serum. The effect of changing media composition was reversible over multiple changes of media type. Other variables tested included culture well size and degree of confluence. The observation that sensitivity of a cell type to environmental agonists can be manipulated by modifying culture conditions suggests a novel approach for studying biochemical mechanisms of particle toxicity. PMID:18178371

  3. Separation of two phenotypically similar cell types via a single common marker in microfluidic channels.

    PubMed

    Vickers, Dwayne A L; Chory, Emma J; Murthy, Shashi K

    2012-09-21

    To isolate clinically and biologically relevant cell types from a heterogeneous population, fluorescent or magnetic tagging together with knowledge of surface biomarker profiles represents the state of the art. To date, it remains exceedingly difficult to separate phenotypically and physically similar cell types from a mixed population. We report a microfluidic platform engineered to separate two highly similar cell types using a single antibody by taking advantage of subtle variations in surface receptor density and cell size. This platform utilizes antibody-conjugated surfaces in microfluidic channels together with precise modulation of fluid shear stresses to accomplish selective fractionation in a continuous flow process. Antibody conjugation density variation on the adhesive surfaces is achieved by covalently immobilizing an antibody in the presence of poly(ethylene glycol). This platform is used to demonstrate separation of two CD31 positive cell types, human umbilical vein endothelial cells and human micro vascular endothelial cells. PMID:22782544

  4. Comparison of Standard and Heart-pacer Type 3rd Electrodes in Design Variable Cells

    NASA Technical Reports Server (NTRS)

    Morrow, G. W.

    1984-01-01

    Nine packs of sealed aerospace nickel cadmium cells were put on life test in February 1979. Each 5 cell pack contained one cell with a standard sensor signal electrode and one cell with a new heart pacer sensor signal electrode. Testing was discontinued in May 1983 and the signal electrode performance data was studied. It was found that the heart pacer electrode generally provided a greater voltage swing over a cycle; that both types of electrodes lost significant sensitivity during life, and that both types of electrodes show great signal variation from cell to cell.

  5. Divergent viral presentation among human tumors and adjacent normal tissues

    PubMed Central

    Cao, Song; Wendl, Michael C.; Wyczalkowski, Matthew A.; Wylie, Kristine; Ye, Kai; Jayasinghe, Reyka; Xie, Mingchao; Wu, Song; Niu, Beifang; Grubb, Robert; Johnson, Kimberly J.; Gay, Hiram; Chen, Ken; Rader, Janet S.; Dipersio, John F.; Chen, Feng; Ding, Li

    2016-01-01

    We applied a newly developed bioinformatics system called VirusScan to investigate the viral basis of 6,813 human tumors and 559 adjacent normal samples across 23 cancer types and identified 505 virus positive samples with distinctive, organ system- and cancer type-specific distributions. We found that herpes viruses (e.g., subtypes HHV4, HHV5, and HHV6) that are highly prevalent across cancers of the digestive tract showed significantly higher abundances in tumor versus adjacent normal samples, supporting their association with these cancers. We also found three HPV16-positive samples in brain lower grade glioma (LGG). Further, recurrent HBV integration at the KMT2B locus is present in three liver tumors, but absent in their matched adjacent normal samples, indicating that viral integration induced host driver genetic alterations are required on top of viral oncogene expression for initiation and progression of liver hepatocellular carcinoma. Notably, viral integrations were found in many genes, including novel recurrent HPV integrations at PTPN13 in cervical cancer. Finally, we observed a set of HHV4 and HBV variants strongly associated with ethnic groups, likely due to viral sequence evolution under environmental influences. These findings provide important new insights into viral roles of tumor initiation and progression and potential new therapeutic targets. PMID:27339696

  6. Transcription factors interfering with dedifferentiation induce cell type-specific transcriptional profiles

    PubMed Central

    Hikichi, Takafusa; Matoba, Ryo; Ikeda, Takashi; Watanabe, Akira; Yamamoto, Takuya; Yoshitake, Satoko; Tamura-Nakano, Miwa; Kimura, Takayuki; Kamon, Masayoshi; Shimura, Mari; Kawakami, Koichi; Okuda, Akihiko; Okochi, Hitoshi; Inoue, Takafumi; Suzuki, Atsushi; Masui, Shinji

    2013-01-01

    Transcription factors (TFs) are able to regulate differentiation-related processes, including dedifferentiation and direct conversion, through the regulation of cell type-specific transcriptional profiles. However, the functional interactions between the TFs regulating different transcriptional profiles are not well understood. Here, we show that the TFs capable of inducing cell type-specific transcriptional profiles prevent the dedifferentiation induced by TFs for pluripotency. Of the large number of TFs expressed in a neural-lineage cell line, we identified a subset of TFs that, when overexpressed, strongly interfered with the dedifferentiation triggered by the procedure to generate induced pluripotent stem cells. This interference occurred through a maintenance mechanism of the cell type-specific transcriptional profile. Strikingly, the maintenance activity of the interfering TF set was strong enough to induce the cell line-specific transcriptional profile when overexpressed in a heterologous cell type. In addition, the TFs that interfered with dedifferentiation in hepatic-lineage cells involved TFs with known induction activity for hepatic-lineage cells. Our results suggest that dedifferentiation suppresses a cell type-specific transcriptional profile, which is primarily maintained by a small subset of TFs capable of inducing direct conversion. We anticipate that this functional correlation might be applicable in various cell types and might facilitate the identification of TFs with induction activity in efforts to understand differentiation. PMID:23550161

  7. Single cell-type comparative metabolomics of epidermal bladder cells from the halophyte Mesembryanthemum crystallinum

    PubMed Central

    Barkla, Bronwyn J.; Vera-Estrella, Rosario

    2015-01-01

    One of the remarkable adaptive features of the halophyte Mesembryanthemum crystallinum are the specialized modified trichomes called epidermal bladder cells (EBC) which cover the leaves, stems, and peduncle of the plant. They are present from an early developmental stage but upon salt stress rapidly expand due to the accumulation of water and sodium. This particular plant feature makes it an attractive system for single cell type studies, with recent proteomics and transcriptomics studies of the EBC establishing that these cells are metabolically active and have roles other than sodium sequestration. To continue our investigation into the function of these unusual cells we carried out a comprehensive global analysis of the metabolites present in the EBC extract by gas chromatography Time-of-Flight mass spectrometry (GC-TOF) and identified 194 known and 722 total molecular features. Statistical analysis of the metabolic changes between control and salt-treated samples identified 352 significantly differing metabolites (268 after correction for FDR). Principal components analysis provided an unbiased evaluation of the data variance structure. Biochemical pathway enrichment analysis suggested significant perturbations in 13 biochemical pathways as defined in KEGG. More than 50% of the metabolites that show significant changes in the EBC, can be classified as compatible solutes and include sugars, sugar alcohols, protein and non-protein amino acids, and organic acids, highlighting the need to maintain osmotic homeostasis to balance the accumulation of Na+ and Cl− ions. Overall, the comparison of metabolic changes in salt treated relative to control samples suggests large alterations in M. crystallinum epidermal bladder cells. PMID:26113856

  8. Intestinal epithelial tuft cells initiate type 2 mucosal immunity to helminth parasites.

    PubMed

    Gerbe, François; Sidot, Emmanuelle; Smyth, Danielle J; Ohmoto, Makoto; Matsumoto, Ichiro; Dardalhon, Valérie; Cesses, Pierre; Garnier, Laure; Pouzolles, Marie; Brulin, Bénédicte; Bruschi, Marco; Harcus, Yvonne; Zimmermann, Valérie S; Taylor, Naomi; Maizels, Rick M; Jay, Philippe

    2016-01-14

    Helminth parasitic infections are a major global health and social burden. The host defence against helminths such as Nippostrongylus brasiliensis is orchestrated by type 2 cell-mediated immunity. Induction of type 2 cytokines, including interleukins (IL) IL-4 and IL-13, induce goblet cell hyperplasia with mucus production, ultimately resulting in worm expulsion. However, the mechanisms underlying the initiation of type 2 responses remain incompletely understood. Here we show that tuft cells, a rare epithelial cell type in the steady-state intestinal epithelium, are responsible for initiating type 2 responses to parasites by a cytokine-mediated cellular relay. Tuft cells have a Th2-related gene expression signature and we demonstrate that they undergo a rapid and extensive IL-4Rα-dependent amplification following infection with helminth parasites, owing to direct differentiation of epithelial crypt progenitor cells. We find that the Pou2f3 gene is essential for tuft cell specification. Pou2f3(-/-) mice lack intestinal tuft cells and have defective mucosal type 2 responses to helminth infection; goblet cell hyperplasia is abrogated and worm expulsion is compromised. Notably, IL-4Rα signalling is sufficient to induce expansion of the tuft cell lineage, and ectopic stimulation of this signalling cascade obviates the need for tuft cells in the epithelial cell remodelling of the intestine. Moreover, tuft cells secrete IL-25, thereby regulating type 2 immune responses. Our data reveal a novel function of intestinal epithelial tuft cells and demonstrate a cellular relay required for initiating mucosal type 2 immunity to helminth infection. PMID:26762460

  9. New adjacent Bis-tetrahydrofuran Annonaceous acetogenins from Annona muricata.

    PubMed

    Chang, Fang-Rong; Liaw, Chih-Chuang; Lin, Chih-Yuan; Chou, Chi-Jung; Chiu, Hui-Fen; Wu, Yang-Chang

    2003-03-01

    Bioactivity-guided fractionation led to the isolation of two new Annonaceous acetogenins, annocatacin A ( 1). and annocatacin B ( 2). from the seeds and the leaves, respectively, of Annona muricata. Compounds 1 and 2 are the first examples where the adjacent bis-tetrahydrofuran ring system is located at C-15. The new structures were elucidated and characterized by spectral and chemical methods. Both Annonaceous acetogenins 1 and 2 showed significant in vitro cytotoxicity toward the human hepatoma cell lines, Hep G2 and 2,2,15, and were compared with the known adjacent bis-tetrahydrofuran acetogenins, neoannonin ( 3). desacetyluvaricin ( 4). bullatacin ( 5). asimicin ( 6). annoglaucin ( 7). squamocin ( 8). and rollimusin ( 9).

  10. Human amniotic membrane-derived stromal cells (hAMSC) interact depending on breast cancer cell type through secreted molecules.

    PubMed

    Kim, Sun-Hee; Bang, So Hee; Kang, So Yeong; Park, Ki Dae; Eom, Jun Ho; Oh, Il Ung; Yoo, Si Hyung; Kim, Chan-Wha; Baek, Sun Young

    2015-02-01

    Human amniotic membrane-derived stromal cells (hAMSC) are candidates for cell-based therapies. We examined the characteristics of hAMSC including the interaction between hAMSC and breast cancer cells, MCF-7, and MDA-MB-231. Human amniotic membrane-derived stromal cells showed typical MSC properties, including fibroblast-like morphology, surface antigen expression, and mesodermal differentiation. To investigate cell-cell interaction via secreted molecules, we cultured breast cancer cells in hAMSC-conditioned medium (hAMSC-CM) and analyzed their proliferation, migration, and secretome profiles. MCF-7 and MDA-MB-231 cells exposed to hAMSC-CM showed increased proliferation and migration. However, in hAMSC-CM, MCF-7 cells proliferated significantly faster than MDA-MB-231 cells. When cultured in hAMSC-CM, MCF-7 cells migrated faster than MDA-MB-231 cells. Two cell types showed different profiles of secreted factors. MCF-7 cells expressed much amounts of IL-8, GRO, and MCP-1 in hAMSC-CM. Human amniotic membrane-derived stromal cells interact with breast cancer cells through secreted molecules. Factors secreted by hAMSCs promote the proliferation and migration of MCF-7 breast cancer cells. For much safe cell-based therapies using hAMSC, it is necessary to study carefully about interaction between hAMSC and cancer cells.

  11. Stem cell-based treatments for Type 1 diabetes mellitus: bone marrow, embryonic, hepatic, pancreatic and induced pluripotent stem cells.

    PubMed

    Godfrey, K J; Mathew, B; Bulman, J C; Shah, O; Clement, S; Gallicano, G I

    2012-01-01

    Type 1 diabetes mellitus--characterized by the permanent destruction of insulin-secreting β-cells--is responsive to cell-based treatments that replace lost β-cell populations. The current gold standard of pancreas transplantation provides only temporary independence from exogenous insulin and is fraught with complications, including increased mortality. Stem cells offer a number of theoretical advantages over current therapies. Our review will focus on the development of treatments involving tissue stem cells from bone marrow, liver and pancreatic cells, as well as the potential use of embryonic and induced pluripotent stem cells for Type 1 diabetes therapy. While the body of research involving stem cells is at once promising and inconsistent, bone marrow-derived mesenchymal stem cell transplantation seems to offer the most compelling evidence of efficacy. These cells have been demonstrated to increase endogenous insulin production, while partially mitigating the autoimmune destruction of newly formed β-cells. However, recently successful experiments involving induced pluripotent stem cells could quickly move them into the foreground of therapeutic research. We address the limitations encountered by present research and look toward the future of stem cell treatments for Type 1 diabetes.

  12. Anti-neutrophil cell antibodies in newly diagnosed patients with type-1-diabetes.

    PubMed

    Parlapiano, C; Marangi, M; Campana, E; Giovanniello, T; Pantone, P; Suraci, C; Sanguigni, S

    1999-01-01

    Both anti neutrophil cell antibodies and anti endothelial cell antibodies were found in 7 out of 30 newly-diagnosed type-1 diabetic patients. This confirms the abnormal activation of the immunological system in the early stage of type-1 diabetes mellitus. PMID:10482047

  13. Cell types differ in global coordination of splicing and proportion of highly expressed genes.

    PubMed

    Trakhtenberg, Ephraim F; Pho, Nam; Holton, Kristina M; Chittenden, Thomas W; Goldberg, Jeffrey L; Dong, Lingsheng

    2016-01-01

    Balance in the transcriptome is regulated by coordinated synthesis and degradation of RNA molecules. Here we investigated whether mammalian cell types intrinsically differ in global coordination of gene splicing and expression levels. We analyzed RNA-seq transcriptome profiles of 8 different purified mouse cell types. We found that different cell types vary in proportion of highly expressed genes and the number of alternatively spliced transcripts expressed per gene, and that the cell types that express more variants of alternatively spliced transcripts per gene are those that have higher proportion of highly expressed genes. Cell types segregated into two clusters based on high or low proportion of highly expressed genes. Biological functions involved in negative regulation of gene expression were enriched in the group of cell types with low proportion of highly expressed genes, and biological functions involved in regulation of transcription and RNA splicing were enriched in the group of cell types with high proportion of highly expressed genes. Our findings show that cell types differ in proportion of highly expressed genes and the number of alternatively spliced transcripts expressed per gene, which represent distinct properties of the transcriptome and may reflect intrinsic differences in global coordination of synthesis, splicing, and degradation of RNA molecules. PMID:27577089

  14. Cell types differ in global coordination of splicing and proportion of highly expressed genes

    PubMed Central

    Trakhtenberg, Ephraim F.; Pho, Nam; Holton, Kristina M.; Chittenden, Thomas W.; Goldberg, Jeffrey L.; Dong, Lingsheng

    2016-01-01

    Balance in the transcriptome is regulated by coordinated synthesis and degradation of RNA molecules. Here we investigated whether mammalian cell types intrinsically differ in global coordination of gene splicing and expression levels. We analyzed RNA-seq transcriptome profiles of 8 different purified mouse cell types. We found that different cell types vary in proportion of highly expressed genes and the number of alternatively spliced transcripts expressed per gene, and that the cell types that express more variants of alternatively spliced transcripts per gene are those that have higher proportion of highly expressed genes. Cell types segregated into two clusters based on high or low proportion of highly expressed genes. Biological functions involved in negative regulation of gene expression were enriched in the group of cell types with low proportion of highly expressed genes, and biological functions involved in regulation of transcription and RNA splicing were enriched in the group of cell types with high proportion of highly expressed genes. Our findings show that cell types differ in proportion of highly expressed genes and the number of alternatively spliced transcripts expressed per gene, which represent distinct properties of the transcriptome and may reflect intrinsic differences in global coordination of synthesis, splicing, and degradation of RNA molecules. PMID:27577089

  15. [Xenogeneic cell therapeutics: Treatment of type 1 diabetes using porcine pancreatic islets and islet cells].

    PubMed

    Godehardt, Antonia W; Schilling-Leiß, Dagmar; Sanzenbacher, Ralf; Tönjes, Ralf R

    2015-11-01

    In view of the existing shortage of human donor organs and tissues, xenogeneic cell therapeutics (xCT) offer an alternative for adequate treatment. In particular, porcine pancreatic islets and islet cells have already entered the field of experimental therapy for type-1 diabetes mellitus (T1DM) patients. Thereby, xCT depict challenging products with a glance on medical, ethical, and regulatory questions. With cross-species transplantation (xenotransplantation), the risk of immunological graft rejection as well as the risk of infectious transmission of microbial and viral pathogens must be considered. This includes the bidirectional transmission of microorganisms from graft to host as well as from host to graft. Crossing the border of species requires a critical risk-benefit evaluation as well as a thorough longtime surveillance of transplant recipients after treatment. The international legal and regulatory requirements for xCT are inter alia based on the World Health Organization criteria summarized in the Changsha Communiqué (2008). In the European Union, they were reflected by the European Medicines Agency (EMA) Guideline on Xenogeneic Cell-based Medicinal Products following the implementation of the Regulation on Advanced Therapies (ATMP). On the basis of this regulation, the first non-clinical and clinical experiences were obtained for porcine islets. The results suggest that supportive treatment of T1DM risk patients with xCT may be an alternative to established allogeneic organ transplantation in the future.

  16. Secondary prevention of type 1 diabetes mellitus: stopping immune destruction and promoting beta-cell regeneration.

    PubMed

    Couri, C E B; Foss, M C; Voltarelli, J C

    2006-10-01

    Type 1 diabetes mellitus results from a cell-mediated autoimmune attack against pancreatic beta-cells. Traditional treatments involve numerous daily insulin dosages/injections and rigorous glucose control. Many efforts toward the identification of beta-cell precursors have been made not only with the aim of understanding the physiology of islet regeneration, but also as an alternative way to produce beta-cells to be used in protocols of islet transplantation. In this review, we summarize the most recent studies related to precursor cells implicated in the regeneration process. These include embryonic stem cells, pancreas-derived multipotent precursors, pancreatic ductal cells, hematopoietic stem cells, mesenchymal stem cells, hepatic oval cells, and mature beta-cells. There is controversial evidence of the potential of these cell sources to regenerate beta-cell mass in diabetic patients. However, clinical trials using embryonic stem cells, umbilical cord blood or adult bone marrow stem cells are under way. The results of various immunosuppressive regimens aiming at blocking autoimmunity against pancreatic beta-cells and promoting beta-cell preservation are also analyzed. Most of these regimens provide transient and partial effect on insulin requirements, but new regimens are beginning to be tested. Our own clinical trial combines a high dose immunosuppression with mobilized peripheral blood hematopoietic stem cell transplantation in early-onset type 1 diabetes mellitus.

  17. Theoretical and experimental investigation of 'grating' type photovoltaic cells

    NASA Technical Reports Server (NTRS)

    Loferski, J. J.; Crisman, E. E.; Armitage, W.; Chen, L. Y.

    1974-01-01

    The fabrication procedure and properties of 'grating' cells made by forming a fine grating pattern of aluminum alloyed into n-silicon wafers are described. The finest grating lines achieved in the cells described were 5 microns; the smallest spacing was about 15 microns. The best temperature for alloying was found to be about 600 C, a bit above the Si-Al eutectic temperature (576 C). The short-circuit current obtained from the best of these cells exposed to 100 mW/sq cm of (simulated air mass zero) illumination was at least equal to that obtained from conventional diffused cells, but their open-circuit voltage was lower. Their quantum yield was strongly blue-shifted; it was flat from 4000 to 8500 A.

  18. Polypeptide composition and gag gene-coded products of type-D oncovirus from HEp-2 cells.

    PubMed

    Morozov, V A

    1982-01-01

    The protein composition of type-D oncovirus HEp-2, isolated from cell-free medium of continuous human HEp-2 cell line, has been investigated using electrophoresis on gradient polyacrylamide gels with sodium dodecyl sulfate (SDS). Labeling with 14C-amino acids revealed five viral polypeptides with molecular weights of 70 000 (gp70), 27 000 (p27), 19 000 (p19), 15 000 (p15), 12 000-10 000 (p12-10). The 70 000 dalton protein was shown to be the only glycoprotein by incorporation of radioactive glucosamine. A polypeptide with molecular weight of 78 000 has been specifically precipitated from pulse-labeled type-D oncovirus producing HEp-2 cells with goat anti Mason-Pfizer p27 serum. This protein was shown to be gag gene-coded polyprotein precursor (Pr78gag) of the major virus polypeptide p27. Pulse-labeled HEp-2 and Mason-Pfizer infected Tu 197 cells were rinsed, lysed, clarified and precipitated with goat anti Mason-Pfizer p27 serum. In both cases Pr78gag was detected.

  19. Update on islet cell transplantation for type 1 diabetes.

    PubMed

    Agarwal, Avinash; Brayman, Kenneth L

    2012-06-01

    Despite modern medical breakthroughs, diabetes mellitus is a worldwide leading cause of morbidity and mortality. Definitive surgical treatment of diabetes mellitus was established with the advent and refinement of clinical pancreas transplantation in the 1960s. During the following decades, critical discoveries involving islet isolation and engraftment took place. Clinical islet cell transplantation represents the potential for reduced insulin requirements and debilitating hypoglycemic episodes without the morbidity of surgery. Unfortunately, islet cell transplantation was unable to achieve comparable results with solid organ transplantation. This was until the Edmonton protocol (steroid-free immunosuppression) was described, which demonstrated that islet cell transplantation could be a viable alternative to pancreas transplantation. Significant advances in islet purification techniques and novel immunomodulatory agents have since renewed interest in islet cell transplantation. Yet the field is still challenged by a limited supply of islet cells, inadequate engraftment, and the deleterious effects of chronic immunosuppression. This article discusses the history and the current status of clinical islet cell transplantation. PMID:23729978

  20. Cell type-specific response to high intracellular loading of polyacrylic acid-coated magnetic nanoparticles

    PubMed Central

    Lojk, Jasna; Bregar, Vladimir B; Rajh, Maruša; Miš, Katarina; Kreft, Mateja Erdani; Pirkmajer, Sergej; Veranič, Peter; Pavlin, Mojca

    2015-01-01

    Magnetic nanoparticles (NPs) are a special type of NP with a ferromagnetic, electron-dense core that enables several applications such as cell tracking, hyperthermia, and magnetic separation, as well as multimodality. So far, superparamagnetic iron oxide NPs (SPIONs) are the only clinically approved type of metal oxide NPs, but cobalt ferrite NPs have properties suitable for biomedical applications as well. In this study, we analyzed the cellular responses to magnetic cobalt ferrite NPs coated with polyacrylic acid (PAA) in three cell types: Chinese Hamster Ovary (CHO), mouse melanoma (B16) cell line, and primary human myoblasts (MYO). We compared the internalization pathway, intracellular trafficking, and intracellular fate of our NPs using fluorescence and transmission electron microscopy (TEM) as well as quantified NP uptake and analyzed uptake dynamics. We determined cell viability after 24 or 96 hours’ exposure to increasing concentrations of NPs, and quantified the generation of reactive oxygen species (ROS) upon 24 and 48 hours’ exposure. Our NPs have been shown to readily enter and accumulate in cells in high quantities using the same two endocytic pathways; mostly by macropinocytosis and partially by clathrin-mediated endocytosis. The cell types differed in their uptake rate, the dynamics of intracellular trafficking, and the uptake capacity, as well as in their response to higher concentrations of internalized NPs. The observed differences in cell responses stress the importance of evaluation of NP–cell interactions on several different cell types for better prediction of possible toxic effects on different cell and tissue types in vivo. PMID:25733835

  1. Eight types of stem cells in the life cycle of the moss Physcomitrella patens.

    PubMed

    Kofuji, Rumiko; Hasebe, Mitsuyasu

    2014-02-01

    Stem cells self-renew and produce cells that differentiate to become the source of the plant body. The moss Physcomitrella patens forms eight types of stem cells during its life cycle and serves as a useful model in which to explore the evolution of such cells. The common ancestor of land plants is inferred to have been haplontic and to have formed stem cells only in the gametophyte generation. A single stem cell would have been maintained in the ancestral gametophyte meristem, as occurs in extant basal land plants. During land plant evolution, stem cells diverged in the gametophyte generation to form different types of body parts, including the protonema and rhizoid filaments, leafy-shoot and thalloid gametophores, and gametangia formed in moss. A simplex meristem with a single stem cell was acquired in the sporophyte generation early in land plant evolution. Subsequently, sporophyte stem cells became multiple in the meristem and were elaborated further in seed plant lineages, although the evolutionary origin of niche cells, which maintain stem cells is unknown. Comparisons of gene regulatory networks are expected to give insights into the general mechanisms of stem cell formation and maintenance in land plants and provide information about their evolution. P. patens develops at least seven types of simplex meristem in the gametophyte and at least one type in the sporophyte generation and is a good material for regulatory network comparisons. In this review, we summarize recently revealed molecular mechanisms of stem cell initiation and maintenance in the moss.

  2. Functional Proteomics Screen Enables Enrichment of Distinct Cell Types from Human Pancreatic Islets

    PubMed Central

    Sharivkin, Revital; Walker, Michael D.; Soen, Yoav

    2015-01-01

    The current world-wide epidemic of diabetes has prompted attempts to generate new sources of insulin-producing cells for cell replacement therapy. An inherent challenge in many of these strategies is the lack of cell-surface markers permitting isolation and characterization of specific cell types from differentiating stem cell populations. Here we introduce an iterative proteomics procedure allowing tag-free isolation of cell types based on their function. Our method detects and associates specific cell-surface markers with particular cell functionality by coupling cell capture on antibody arrays with immunofluorescent labeling. Using this approach in an iterative manner, we discovered marker combinations capable of enriching for discrete pancreatic cell subtypes from human islets of Langerhans: insulin-producing beta cells (CD9high/CD56+), glucagon-producing alpha cells (CD9- /CD56+) and trypsin-producing acinar cells (CD9- /CD56-). This strategy may assist future beta cell research and the development of diagnostic tools for diabetes. It can also be applied more generally for function-based purification of desired cell types from other limited and heterogeneous biological samples. PMID:25706282

  3. Novel therapy for type 1 diabetes: autologous hematopoietic stem cell transplantation.

    PubMed

    Li, Lirong; Gu, Weiqiong; Zhu, Dalong

    2012-12-01

    Type 1 diabetes is characterized pathologically by autoimmune insulitis-related islet β-cell destruction. Although intensive insulin therapy for patients with type 1 diabetes can correct hyperglycemia, this therapy does not prevent all diabetes-related complications. Recent studies have shown that autologous hematopoietic stem cell transplantation (HSCT) is a promising new approach for the treatment of type 1 diabetes by reconstitution of immunotolerance and preservation of islet β-cell function. Herein we discuss the therapeutic efficacy and potential mechanisms underlying the action of HSCT and other perspectives in the clinical management of type 1 diabetes.

  4. Cancer type-dependent genetic interactions between cancer driver alterations indicate plasticity of epistasis across cell types

    PubMed Central

    Park, Solip; Lehner, Ben

    2015-01-01

    Cancers, like many diseases, are normally caused by combinations of genetic alterations rather than by changes affecting single genes. It is well established that the genetic alterations that drive cancer often interact epistatically, having greater or weaker consequences in combination than expected from their individual effects. In a stringent statistical analysis of data from > 3,000 tumors, we find that the co-occurrence and mutual exclusivity relationships between cancer driver alterations change quite extensively in different types of cancer. This cannot be accounted for by variation in tumor heterogeneity or unrecognized cancer subtypes. Rather, it suggests that how genomic alterations interact cooperatively or partially redundantly to driver cancer changes in different types of cancers. This re-wiring of epistasis across cell types is likely to be a basic feature of genetic architecture, with important implications for understanding the evolution of multicellularity and human genetic diseases. In addition, if this plasticity of epistasis across cell types is also true for synthetic lethal interactions, a synthetic lethal strategy to kill cancer cells may frequently work in one type of cancer but prove ineffective in another. PMID:26227665

  5. Adenovirus type 12 gene 401 function and temperature sensitivity of cytochalasin B effects on transformed cells.

    PubMed

    Ledinko, N; Bhe, F T

    1980-01-01

    Rat (3Y1) cells transformed by wild-type adenovirus type 12 or the temperature-sensitive mutant ts401 with an active function required for transformation maintenance were exposed at the permissive(36 degrees) or nonpermissive (40 degrees) temperature to cytochalasin B (CB). At 40 degrees, the ts401-transformed cells, but not the wild-type transformants, exhibited, at least partially, the untransformed 3Y1 cell phenotype; most of the cells became bi- and trinucleated and DNA synthesis was inhibited. AT 36 degrees, both groups of cells became highly multinucleated, and there was no apparent inhibition of DNA synthesis by CB. These characteristics were exhibited also by the wild-type transformants at 40 degrees. These findings provide additional evidence that an active 401 gene function is required for maintenance of the adenovirus-transformed cell phenotype. PMID:7251331

  6. Output of Neurogliaform Cells to Various Neuron Types in the Human and Rat Cerebral Cortex

    PubMed Central

    Oláh, Szabolcs; Komlósi, Gergely; Szabadics, János; Varga, Csaba; Tóth, Éva; Barzó, Pál; Tamás, Gábor

    2007-01-01

    Neurogliaform cells in the rat elicit combined GABAA and GABAB receptor-mediated postsynaptic responses on cortical pyramidal cells and establish electrical synapses with various interneuron types. However, the involvement of GABAB receptors in postsynaptic effects of neurogliaform cells on other GABAergic interneurons is not clear. We measured the postsynaptic effects of neurogliaform cells in vitro applying simultaneous whole-cell recordings in human and rat cortex. Single action potentials of human neurogliaform cells evoked unitary IPSPs composed of GABAA and GABAB receptor-mediated components in various types of inteneuron and in pyramidal cells. Slow IPSPs were combined with homologous and heterologous electrical coupling between neurogliaform cells and several human interneuron types. In the rat, single action potentials in neurogliaform cells elicited GABAB receptor-mediated component in responses of neurogliaform, regular spiking, and fast spiking interneurons following the GABAA receptor-mediated component in postsynaptic responses. In conclusion, human and rat neurogliaform cells elicit slow IPSPs and reach GABAA and GABAB receptors on several interneuron types with a connection-specific involvement of GABAB receptors. The electrical synapses recorded between human neurogliaform cells and various interneuron types represent the first electrical synapses recorded in the human cortex. PMID:18946546

  7. A novel, cell-specific attenuation of a herpes simplex virus type 1 infection in vivo.

    PubMed

    Kienzle, T E; Chen, T M; Mrak, R E; Stroop, W G

    2001-04-01

    We have observed a cell-specific attenuation of herpes simplex virus type 1 strain 17syn+ in vivo that was dependent upon the cell type used to grow the virus. Direct corneal infection of rabbits with 17syn+ propagated in Vero cells caused 60% (6 of 10) to develop severe central nervous system (CNS) disease as evidenced by seizures and/or paralysis; all neurologically impaired rabbits died. In contrast, infection of rabbits with 17syn+ propagated in BHK-21 cells induced seizures and was fatal in 10% (1 of 10). The cell-specific attenuation of a 17syn+ occurred after one growth cycle in BHK-21 cells. To determine whether the decreased virulence of the BHK-21 cell-grown virus correlated with a less severe CNS inflammatory reaction, CNS tissues from rabbits infected with 17syn+ grown in Vero and BHK-21 cells were compared. Histopathological analyses revealed no differences in the location or severity of inflammatory lesions from rabbits infected with virus grown in either cell type. Virus-induced corneal disease was less dependent upon the cell type used to propagate the virus as there were no significant differences in the type or severity of observed corneal lesions. Possible explanations based on differences between Vero and BHK-21 cells are discussed.

  8. MINARETS WILDERNESS AND ADJACENT AREAS, CALIFORNIA.

    USGS Publications Warehouse

    Huber, N. King; Thurber, Horace K.

    1984-01-01

    A mineral survey of the Minarets Wilderness and adjacent areas in the central Sierra Nevada, California was conducted. The results of the survey indicate that the study area has a substantiated resource potential for small deposits of copper, silver, zinc, lead, and iron, and a probable mineral-resource potential for molybdenum. No energy-resource potential was identified in the study.

  9. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) DANGEROUS CARGOES CARRIAGE OF BULK SOLID MATERIALS... transporting a material that Table 148.10 of this part associates with a reference to this section, the following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  10. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) DANGEROUS CARGOES CARRIAGE OF BULK SOLID MATERIALS... transporting a material that Table 148.10 of this part associates with a reference to this section, the following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  11. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) DANGEROUS CARGOES CARRIAGE OF BULK SOLID MATERIALS... transporting a material that Table 148.10 of this part associates with a reference to this section, the following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  12. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) DANGEROUS CARGOES CARRIAGE OF BULK SOLID MATERIALS... transporting a material that Table 148.10 of this part associates with a reference to this section, the following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  13. Topographical Control of Ocular Cell Types for Tissue Engineering

    PubMed Central

    McHugh, Kevin J.; Saint-Geniez, Magali; Tao, Sarah L.

    2014-01-01

    Visual impairment affects over 285 million people worldwide and has a major impact on an individual’s quality of life. Tissue engineering has the potential to increase quality of life for many of these patients by preventing vision loss or restoring vision using cell-based therapies. However, these strategies will require an understanding of the microenvironmental factors that influence cell behavior. The eye is a well-organized organ whose structural complexity is essential for proper function. Interactions between ocular cells and their highly ordered extracellular matrix are necessary for maintaining key tissue properties including corneal transparency and retinal lamination. Therefore, it is not surprising that culturing these cells in vitro on traditional flat substrates result in irregular morphology. Instead, topographically patterned biomaterials better mimic native extracellular matrix and have been shown to elicit in vivo-like morphology and gene expression which is essential for tissue engineering. Herein we review multiple methods for producing well-controlled topography and discuss optimal biomaterial scaffold design for cells of the cornea, retina, and lens. PMID:23744715

  14. Topographical control of ocular cell types for tissue engineering.

    PubMed

    McHugh, Kevin J; Saint-Geniez, Magali; Tao, Sarah L

    2013-11-01

    Visual impairment affects over 285 million people worldwide and has a major impact on an individual's quality of life. Tissue engineering has the potential to increase the quality of life for many of these patients by preventing vision loss or restoring vision using cell-based therapies. However, these strategies will require an understanding of the microenvironmental factors that influence cell behavior. The eye is a well-organized organ whose structural complexity is essential for proper function. Interactions between ocular cells and their highly ordered extracellular matrix are necessary for maintaining key tissue properties including corneal transparency and retinal lamination. Therefore, it is not surprising that culturing these cells in vitro on traditional flat substrates result in irregular morphology. Instead, topographically patterned biomaterials better mimic native extracellular matrix and have been shown to elicit in vivo-like morphology and gene expression which is essential for tissue engineering. Herein we review multiple methods for producing well-controlled topography and discuss optimal biomaterial scaffold design for cells of the cornea, retina, and lens.

  15. Pulmonary Alveolar Type II Epithelial Cells and Adult Respiratory Distress Syndrome

    PubMed Central

    Mason, Robert J.

    1985-01-01

    During the past ten years, functions of alveolar type II cells have been well characterized with isolated cells in vitro. Some of the functions were well known from studies in vivo, but others such as transepithelial sodium transport were unsuspected. A better understanding of this important pulmonary cell type improves our knowledge of the pathophysiology of adult respiratory distress syndrome and may in time lead to new therapeutic strategies. ImagesFigure 1.Figure 2.Figure 3.Figure 4. PMID:3909639

  16. Cell cycle regulation and cell type-specific localization of the FtsZ division initiation protein in Caulobacter.

    PubMed Central

    Quardokus, E; Din, N; Brun, Y V

    1996-01-01

    Many genes involved in cell division and DNA replication and their protein products have been identified in bacteria; however, little is known about the cell cycle regulation of the intracellular concentration of these proteins. It has been shown that the level of the tubulin-like GTPase FtsZ is critical for the initiation of cell division in bacteria. We show that the concentration of FtsZ varies dramatically during the cell cycle of Caulobacter crescentus. Caulobacter produce two different cell types at each cell division: (i) a sessile stalked cell that can initiate DNA replication immediately after cell division and (ii) a motile swarmer cell in which DNA replication is blocked. After cell division, only the stalked cell contains FtsZ. FtsZ is synthesized slightly before the swarmer cells differentiate into stalked cells and the intracellular concentration of FtsZ is maximal at the beginning of cell division. Late in the cell cycle, after the completion of chromosome replication, the level of FtsZ decreases dramatically. This decrease is probably mostly due to the degradation of FtsZ in the swarmer compartment of the predivisional cell. Thus, the variation of FtsZ concentration parallels the pattern of DNA synthesis. Constitutive expression of FtsZ leads to defects in stalk biosynthesis suggesting a role for FtsZ in this developmental process in addition to its role in cell division. Images Fig. 2 Fig. 3 Fig. 4 PMID:8692812

  17. Current trends in type 1 diabetes mellitus--stem cells and beyond.

    PubMed

    Hingorjo, Mozaffer Rahim; Syed, Sadiqa; Qureshi, Masood A; Kumar, Ashok

    2007-12-01

    Search for a cure for type-1 diabetes mellitus has lead to many avenues of research, all having the same objective: to replace the lost beta cells and prevent their further destruction by the immune system. Transplantation of islets of Langerhans seems closer to achieving this goal with the recent introduction of new improved immunosuppressive protocols including monoclonal antibodies against the T-lymphocytes. But the need for acquiring beta cells in large numbers rather limits this approach. With the recent advancement in stem cell technology, it may be possible to gather enough stem cells for transplantation purposes. In this regard, embryonic stem cells have shown the greatest promise due to their capacity for unlimited proliferation and differentiation into any cell type. This review discusses the current direction of research regarding diabetes mellitus type-1, while explaining the progress being made in stem cell usage in finding a cure for the disease.

  18. The types of endocrine cells in the pancreas of Sunda porcupine (Hystrix javanica)

    PubMed Central

    Budipitojo, Teguh; Fibrianto, Yuda Heru; Mulyani, Guntari Titik

    2016-01-01

    Aim: To identify the types of endocrine cells in the pancreas of the Sunda porcupine (Hystrix javanica) and its immunolocalization. Materials and Methods: Five adult H. javanica were used without sexual distinction. The presences of endocrine cells (glucagon, insulin, somatostatin, and pancreatic polypeptide [PP]) in pancreatic tissues were detected using the avidin-biotin-peroxidase complex method. Results: The fusiform, round, and oval form endocrine cells were detected in the islets of Langerhans and exocrine parts. Most of the insulin cells were found in the central area, glucagon cells were identified in the central and peripheral areas, and somatostatin and PP cells were detected in the mantle area of the islets of Langerhans. Glucagon and somatostatin cells were also detected in smaller numbers of peripheral parts of the islet. In all of the islet parts, glucagon endocrine cells were most prevalent cell type and then, somatostatin, insulin, and PP. In the exocrine parts, PP, somatostatin, glucagon, and insulin endocrine cells were found in the inter-acinus part with moderate, moderate, a few and rare numbers, in that order. In the pancreatic duct, glucagon and somatostatin cells were found between epithelial cells in rare numbers. Conclusion: The pancreas of Sunda porcupine (H. javanica) contains four types of major pancreatic endocrine cells with approximately similar distribution patterns to the other rodents, except for abundant glucagon cells in the peripheral area of the islets of Langerhans. PMID:27397977

  19. Systematic and Cell Type-Specific Telomere Length Changes in Subsets of Lymphocytes

    PubMed Central

    Lin, Jue; Cheon, Joshua; Brown, Rashida; Coccia, Michael; Puterman, Eli; Aschbacher, Kirstin; Sinclair, Elizabeth; Epel, Elissa; Blackburn, Elizabeth H.

    2016-01-01

    Telomeres, the protective DNA-protein complexes at the ends of linear chromosomes, are important for genome stability. Leukocyte or peripheral blood mononuclear cell (PBMC) telomere length is a potential biomarker for human aging that integrates genetic, environmental, and lifestyle factors and is associated with mortality and risks for major diseases. However, only a limited number of studies have examined longitudinal changes of telomere length and few have reported data on sorted circulating immune cells. We examined the average telomere length (TL) in CD4+, CD8+CD28+, and CD8+CD28− T cells, B cells, and PBMCs, cross-sectionally and longitudinally, in a cohort of premenopausal women. We report that TL changes over 18 months were correlated among these three T cell types within the same participant. Additionally, PBMC TL change was also correlated with those of all three T cell types, and B cells. The rate of shortening for B cells was significantly greater than for the three T cell types. CD8+CD28− cells, despite having the shortest TL, showed significantly more rapid attrition when compared to CD8+CD28+ T cells. These results suggest systematically coordinated, yet cell type-specific responses to factors and pathways contribute to telomere length regulation. PMID:26977417

  20. List of gene variants developed for cancer cells from nine tissue types

    Cancer.gov

    NCI scientists have developed a comprehensive list of genetic variants for each of the types of cells that comprise what is known as the NCI-60 cell line collection. This new list adds depth to the most frequently studied human tumor cell lines in cancer

  1. Analysis of alterations adjacent to invasive vulvar carcinoma and their relationship with the associated carcinoma: a study of 67 cases.

    PubMed

    Vilmer, C; Cavelier-Balloy, B; Nogues, C; Trassard, M; Le Doussal, V

    1998-01-01

    A retrospective analysis of histological lesions adjacent to 67 invasive vulvar squamous cell carcinomas (SCC) was undertaken to analyse their nature, as well as their relationship to SCC. Patient age, clinical presentation and histological type of carcinoma, ISSVD classification of its adjacent lesions, disease-free and overall survival were reviewed. Severe undifferentiated vulvar intra-epithelial neoplasia (VIN3) was found in 19.4% of cases and vulvar lichen sclerosus (VLS) in 76.1% of cases. All VLS, except 2 cases, were associated with squamous cell hyperplasia (SCH), and a concomitant differentiated VIN was found in 76.6% of cases. Undifferentiated VIN3 was never associated with VLS. VLS was significantly associated with a keratinizing, well-differentiated SCC (98% of cases), while undifferentiated VIN3, was linked preferentially to 2 other types of SCC: in 77% of cases, a moderately-differentiated SCC with the same histological features as the so-called basaloid carcinoma and, in 23% of cases, a well-differentiated SCC with a variable extent of koilocytic atypia, similar to the so-called warty carcinoma. Carcinoma of the fourchette was more often associated with undifferentiated VIN3. Disease-free and overall survival were significantly better for carcinoma associated with undifferentiated VIN3 (p < 0.01 and p < 0.05, respectively). These findings suggest invasive vulvar SCC occurs on 2 distinct types of vulvar lesions: differentiated VIN and/or SCH associated with VLS and undifferentiated VIN3. Furthermore, the histological type of the carcinoma seems to differ according to adjacent lesions.

  2. Cell Type-specific Translational Profiling in the Xenopus laevis Retina

    PubMed Central

    Watson, F.L.; Mills, E. A.; Wang, X.; Guo, C.; Chen, D.F.; Marsh-Armstrong, N.

    2013-01-01

    Background Translating Ribosome Affinity Purification (TRAP), a method recently developed to generate cell type-specific translational profiles, relies on creating transgenic lines of animals in which a tagged ribosomal protein is placed under regulatory control of a cell type-specific promoter. An antibody is then used to affinity purify the tagged ribosomes so that cell type-specific mRNAs can be isolated from whole tissue lysates. Results Here, cell type-specific transgenic lines were generated to enable TRAP studies for retinal ganglion cells and rod photoreceptors in the Xenopus laevis retina. Using real time quantitative PCR for assessing expression levels of cell type-specific mRNAs, the TRAP method was shown to selectively isolate mRNAs expressed in the targeted cell and was efficient at purifying mRNAs expressed at both high and low levels. Statistical measures used to distinguish cell type-specific RNAs from low level background and non-specific RNAs showed TRAP to be highly effective in Xenopus. Conclusions TRAP can be used to purify mRNAs expressed in rod photoreceptors and retinal ganglion cells in Xenopus laevis. The generated transgenic lines will enable numerous studies into the development, disease and injury of the Xenopus laevis retina. PMID:23074098

  3. Hematologically and genetically distinct forms of sickle cell anemia in Africa. The Senegal type and the Benin type.

    PubMed

    Nagel, R L; Fabry, M E; Pagnier, J; Zohoun, I; Wajcman, H; Baudin, V; Labie, D

    1985-04-01

    Patients with sickle cell anemia vary in the hematologic and clinical features of their disease, in part because of variability in the presence of linked and unlinked genes that modify the expression of the disease. The hemoglobin S gene is strongly linked to three different haplotypes of polymorphic endonuclease-restriction sites of the beta-like gene cluster (genes in the vicinity of the beta-globin gene)--one prevalent in Atlantic West Africa, another in central West Africa, and yet another in Bantu-speaking Africa (equatorial, East, and southern Africa). We have studied the differences in the hematologic characteristics of patients with sickle cell anemia from the first two geographical areas. We find that the Senegalese (Atlantic West Africa) patients have higher levels of hemoglobin F, a preponderance of G gamma chains in hemoglobin F, a lower proportion of very dense red cells, and a lower percentage of irreversibly sickled cells than those from Benin (central West Africa). We interpret these data to mean that the gamma-chain composition and the hemoglobin F level are haplotype linked and that the decrease in the percentage of dense cells and irreversibly sickled cells is secondary to the elevation in the hemoglobin F level. Patients with sickle cell anemia in the New World probably correspond to various combinations of these types, in addition to the still hematologically undefined haplotype associated with sickle cell anemia in the Bantu-speaking areas of Africa. PMID:2579336

  4. Effect of adjacent insulating oxide layers on superconductivity of one unit cell thick YBa 2Cu 3O 7-δ layers in PrBa 2Cu 3O 7-δ/YBa 2Cu 3O 7-δ/insulating oxide trilayers

    NASA Astrophysics Data System (ADS)

    Bando, Yoshichika; Terashima, Takahito; Shimura, Ken-ichi; Daitoh, Yoshihiro; Yano, Yoshihiko

    1995-05-01

    One unit cell thick (1-UCT) YBa 2Cu 3O 7-δ (YBCO) adjacent to nonsuperconducting PrBa 2Cu 3O 7-δ (PrBCO) buffer layers have been grown epitaxially on SrTiO 3 (100) using molecular beam epitaxy (MBE). The layer thickness was accurately controlled by monitoring reflection high-energy electron diffraction (RHEED) specular intensity oscillations. Various oxide caps were grown epitaxially on the 1-UCT YBCO layer to form PrBCO/1-UCT YBCO/insulating oxide trilayers. It was found that superconducting 1-UCT YBCO layers were achieved when BaO with a good lattice match to YBCO was used as an insulating oxide in the trilayers. As the terminating atomic layer of 1-UCT YBCO layer is CuO 1-δ, the addition of an atomic BaO layer means the completion of the charge reservoir block of BaO-CuO 1-δ-BaO positioned above the CuO 2 bilayer (CuO 2YCuO 2). The PrBCO buffer layers provide another charge reservoir block positioned below the CuO 2 bilayer interposing Y. Therefore, the minimum unit for superconductivity in YBCO is the CuO 2 bilayer sandwiched between the charge reservoir blocks. In the present study, the systematic substitution of various AO layers for the one-sided atomic BaO layer of the charge reservoir block has been made by epitaxial growth of cap oxides such as rock-salt-type and perovskite-type oxides on the CuO 1-δ-terminated surface of 1-UCT YBCO layers. It is found that the small lattice mismatch between YBCO and the cap oxides, in addition to divalent A ions such as Ba 2+, Sr 2+, Ca 2+, Pb 2+ and Cd 2+ in atomic AO layers subsequent to the CuO 1-δ-terminated surface, is a requirement for superconductivity in 1-UCT YBCO layers.

  5. BRAF mutation is associated with a specific cell-type with features suggestive of senescence in ovarian serous borderline (atypical proliferative) tumors

    PubMed Central

    Zeppernick, Felix; Ardighieri, Laura; Hannibal, Charlotte G.; Vang, Russell; Junge, Jette; Kjaer, Susanne K.; Zhang, Rugang; Kurman, Robert J.; Shih, Ie-Ming

    2014-01-01

    Serous borderline tumor (SBT) also known as atypical proliferative serous tumor (APST) is the precursor of ovarian low-grade serous carcinoma (LGSC). In this study, we correlated the morphologic and immunohistochemical phenotypes of 71 APSTs and 18 LGSCs with the mutational status of KRAS and BRAF, the most common molecular genetic changes in these neoplasms. A subset of cells characterized by abundant eosinophilic cytoplasm (EC), discrete cell borders and bland nuclei was identified in all (100%) 25 BRAF mutated APSTs but in only 5 (10%) of 46 APSTs without BRAF mutations (p<0.0001). Among the 18 LGSCs, EC cells were found in only 2 and both contained BRAF mutations. The EC cells were present admixed with cuboidal and columnar cells lining the papillae and appeared to be budding from the surface, resulting in individual cells and clusters of detached cells “floating” above the papillae. Immunohistochemistry showed that the EC cells always expressed p16, a senescence-associated marker, and had a significantly lower Ki-67 labeling index than adjacent cuboidal and columnar cells (p=0.02). In vitro studies supported the interpretation that these cells were undergoing senescence as the same morphologic features could be reproduced in cultured epithelial cells by ectopic expression of BRAFV600E. Senescence was further established by markers such as SA-β-gal staining, expression of p16 and p21, and reduction in DNA synthesis. In conclusion, this study sheds light on the pathogenesis of this unique group of ovarian tumors by showing that BRAF mutation is associated with cellular senescence and the presence of a specific cell type characterized by abundant eosinophilic cytoplasm. This “oncogene-induced senescence” phenotype may represent a mechanism that prevents impedes progression of APSTs to LGSC. PMID:25188864

  6. A cavity type absorption cell for double resonance microwave spectroscopy.

    NASA Technical Reports Server (NTRS)

    Lee, M. C.; White, W. F.

    1972-01-01

    Description of an experimental dual resonant cavity absorption cell for observing microwave spectroscopic double-resonance effects. The device is composed of two Fabry-Perot interferometers excited by independent microwave sources and mounted at right angles in a suitable vacuum enclosure. The pumping transition is modulated by one source and the modulation induced on the rf absorption in the orthogonal cavity is detected.

  7. Human papillomavirus type 16 DNA in periungual squamous cell carcinomas

    SciTech Connect

    Moy, R.L.; Eliezri, Y.D.; Bennett, R.G. ); Nuovo, G.J.; Siverstein, S. Columbia Univ., New York, NY ); Zitelli, J.A. )

    1989-05-12

    Ten squamous cell carcinomas (in situ or invasive) of the fingernail region were analyzed for the presence of DNA sequences homologous to human papilloma-virus (HPV) by dot blot hybridization. In most patients, the lesions were verrucae of long-term duration that were refractory to conventional treatment methods. Eight of the lesions contained HPV DNA sequences, and in six of these the sequences were related to HPV 16 as deduced from low-stringency nucleic acid hybridization followed by low- and high-stringency washes. Furthermore, the restriction endonuclease digestion pattern of DNA isolated from four of these lesions was diagnostic of episomal HPV 16. The high-frequency association of HPV 16 with periungual squamous cell carcinoma is similar to that reported for HPV 16 with squamous cell carcinomas on mucous membranes at other sites, notably the genital tract. The findings suggest that HPV 16 may play an important role in the development of squamous cell carcinomas of the finger, most notably those lesions that are chronic and located in the periungual area.

  8. Induction of dendritic cell production of type I and type III interferons by wild-type and vaccine strains of measles virus: role of defective interfering RNAs.

    PubMed

    Shivakoti, Rupak; Siwek, Martina; Hauer, Debra; Schultz, Kimberly L W; Griffin, Diane E

    2013-07-01

    The innate immune response to viral infection frequently includes induction of type I interferons (IFN), but many viruses have evolved ways to block this response and increase virulence. In vitro studies of IFN production after infection of susceptible cells with measles virus (MeV) have often reported greater IFN synthesis after infection with vaccine than with wild-type strains of MeV. However, the possible presence in laboratory virus stocks of 5' copy-back defective interfering (DI) RNAs that induce IFN independent of the standard virus has frequently confounded interpretation of data from these studies. To further investigate MeV strain-dependent differences in IFN induction and the role of DI RNAs, monocyte-derived dendritic cells (moDCs) were infected with the wild-type Bilthoven strain and the vaccine Edmonston-Zagreb strain with and without DI RNAs. Production of type I IFN, type III IFN, and the interferon-stimulated genes (ISGs) Mx and ISG56 by infected cells was assessed with a flow cytometry-based IFN bioassay, quantitative reverse transcriptase PCR (RT-PCR), and immunoassays. Bilthoven infected moDCs less efficiently than Edmonston-Zagreb. Presence of DI RNAs in vaccine stocks resulted in greater maturation of moDCs, inhibition of virus replication, and induction of higher levels of IFN and ISGs. Production of type I IFN, type III IFN, and ISG mRNA and protein was determined by both the level of infection and the presence of DI RNAs. At the same levels of infection and in the absence of DI RNA, IFN induction was similar between wild-type and vaccine strains of MeV. PMID:23678166

  9. Boric acid solution concentration influencing p-type emitter formation in n-type crystalline Si solar cells

    NASA Astrophysics Data System (ADS)

    Singha, Bandana; Singh Solanki, Chetan

    2016-09-01

    Boric acid (BA) is a spin on dopant (BSoD) source which is used to form p+ emitters in n-type c-Si solar cells. High purity boric acid powder (99.99% pure) when mixed with deionized (DI) water can result in high quality p-type emitter with less amount of surface defects. In this work, we have used different concentrations of boric acid solution concentrations to fabricate p-type emitters with sheet resistance values < 90 Ω/□. The corresponding junction depths for the same are less than 500 nm as measured by SIMS analysis. Boron rich layer (BRL), which is considered as detrimental in emitter performance is found to be minimal for BA solution concentration less than 2% and hence useful for p-type emitter formation.

  10. Search for inhibitors against herpes simplex virus type-I in cell extracts derived from human lymphoblastoid cell lines.

    PubMed

    Lin, K H

    1977-06-01

    Cell extracts obtained from KB cells and 5 human lymphoblastoid cell lines including 2 from Burkitt's lymphoma (P3HR-1 and Raji), one each from nasopharyngeal carcinoma (no.223), acute lymphatic leukemia (MOLT-4) and a healthy person (NC-37) were tested for their inhibitory effects on the growth of herpes simplex virus type-1 (HSV-1) in green monkey kidney (GMK) cells by the plaque titration method. The relationship between the production of HSV-1 inhibitors and the degree of Epstein-Barr virus (EBV) genome repression in lymphoblastoid cells were also examined. Among the cell lines used P3HR-1 and no.223 cells produced a few EBV particles, Raji and NC-37 cells contained EBV genomes only, and MOLT-4 as well as KB cells were EBV genome-negative. The results revealed that P3HR-1 cell extract showed a tendency to inhibit HSV-1 growth in GMK cells but the other 4 lymphoblastoid cell lines and KB cells did not produce HSV-1 inhibitors, indicating that EBV genomes governing the formation of EBV structural antigens were not related to the production of HSV-1 growth inhibitors. The extracts from MOLT-4 cells, which are only a T lymphocyte cell line used in this study, stimulated HSV-1 growth in GMK cells significantly.

  11. Tousled-like kinase regulates cytokine-mediated communication between cooperating cell types during collective border cell migration

    PubMed Central

    Xiang, Wenjuan; Zhang, Dabing; Montell, Denise J.

    2016-01-01

    Collective cell migration is emerging as a major contributor to normal development and disease. Collective movement of border cells in the Drosophila ovary requires cooperation between two distinct cell types: four to six migratory cells surrounding two immotile cells called polar cells. Polar cells secrete a cytokine, Unpaired (Upd), which activates JAK/STAT signaling in neighboring cells, stimulating their motility. Without Upd, migration fails, causing sterility. Ectopic Upd expression is sufficient to stimulate motility in otherwise immobile cells. Thus regulation of Upd is key. Here we report a limited RNAi screen for nuclear proteins required for border cell migration, which revealed that the gene encoding Tousled-like kinase (Tlk) is required in polar cells for Upd expression without affecting polar cell fate. In the absence of Tlk, fewer border cells are recruited and motility is impaired, similar to inhibition of JAK/STAT signaling. We further show that Tlk in polar cells is required for JAK/STAT activation in border cells. Genetic interactions further confirmed Tlk as a new regulator of Upd/JAK/STAT signaling. These findings shed light on the molecular mechanisms regulating the cooperation of motile and nonmotile cells during collective invasion, a phenomenon that may also drive metastatic cancer. PMID:26510500

  12. Unraveling the contribution of pancreatic beta-cell suicide in autoimmune type 1 diabetes✩

    PubMed Central

    Jaberi-Douraki, Majid; Schnell, Santiago; Pietropaolo, Massimo; Khadra, Anmar

    2014-01-01

    In type 1 diabetes, an autoimmune disease mediated by autoreactive T-cells that attack insulin-secreting pancreatic beta-cells, it has been suggested that disease progression may additionally require protective mechanisms in the target tissue to impede such auto-destructive mechanisms. We hypothesize that the autoimmune attack against beta-cells causes endoplasmic reticulum stress by forcing the remaining beta-cells to synthesize and secrete defective insulin. To rescue beta-cell from the endoplasmic reticulum stress, beta-cells activate the unfolded protein response to restore protein homeostasis and normal insulin synthesis. Here we investigate the compensatory role of unfolded protein response by developing a multi-state model of type 1 diabetes that takes into account beta-cell destruction caused by pathogenic autoreactive T-cells and apoptosis triggered by endoplasmic reticulum stress. We discuss the mechanism of unfolded protein response activation and how it counters beta-cell extinction caused by an autoimmune attack and/or irreversible damage by endoplasmic reticulum stress. Our results reveal important insights about the balance between beta-cell destruction by autoimmune attack (beta-cell homicide) and beta-cell apoptosis by endoplasmic reticulum stress (beta-cell suicide). It also provides an explanation as to why the unfolded protein response may not be a successful therapeutic target to treat type 1 diabetes. PMID:24831415

  13. Lipocalin-type prostaglandin D synthase as a marker for the proliferative potential of melanocyte-lineage cells in the human skin.

    PubMed

    Shimanuki, Miwa; Takeda, Kazuhisa; Kawaguchi, Masakazu; Suzuki, Tamio; Shibahara, Shigeki

    2012-08-01

    Melanocytes in the human epidermis actively produce and secrete various substances, thereby contributing to the maintenance of the skin homeostasis. Lipocalin-type prostaglandin D synthase (L-PGDS) that catalyzes the formation of prostaglandin D(2) (PGD(2) ) may be one of such secreted molecules. Once secreted, L-PGDS functions as a transporter for lipophilic ligands, including all-trans retinoic acid (RA). L-PGDS, therefore, may possess pleiotropic functions in the skin through PGD(2) and RA. We aimed to identify the cell types that express L-PGDS in human skin and to explore the role of L-PGDS in the growth potential of melanocyte-lineage cells. Immunohistochemical analysis for L-PGDS expression was performed with the tissue sections that were prepared from five malignant melanomas, six nevus cell nevi and one Spitz nevus. Normal skin tissues adjacent to the excised melanoma tissues were also analyzed. L-PGDS is expressed in epidermal melanocytes but its expression is undetectable in keratinocytes. Moreover, L-PGDS is undetectable in most benign nevus cells, which may reflect the marginally accelerated proliferation of nevus cells. In contrast, L-PGDS is overexpressed in malignant melanomas, although the frequency of L-PGDS-positive cells was variable (15-50%), depending on the specimens. Lastly, RNA interference analysis against human L-PGDS was performed with short interfering RNA. Knockdown of L-PGDS expression with short interfering RNA in cultured cells suggests that L-PGDS may restrict cell proliferation through RA. In conclusion, L-PGDS expression may contribute to the restricted proliferation of epidermal melanocytes, but conversely its overexpression may reflect the dysregulated proliferation of melanoma cells.

  14. Analysis of Cells Targeted by Salmonella Type III Secretion In Vivo

    PubMed Central

    Geddes, Kaoru; Cruz, Frank; Heffron, Fred

    2007-01-01

    The type III secretion systems (TTSS) encoded in Salmonella pathogenicity island-1 and -2 (SPI-1 and -2) are virulence factors required for specific phases of Salmonella infection in animal hosts. However, the host cell types targeted by the TTSS have not been determined. To investigate this, we have constructed translational fusions between the ß-lactamase reporter and a broad array of TTSS effectors secreted via SPI-1, SPI-2, or both. Secretion of the fusion protein to a host cell was determined by cleavage of a specific fluorescent substrate. In cultured cells, secretion of all six effectors could be observed. However, two to four days following i.p. infection of mice, only effectors secreted by SPI-2 were detected in spleen cells. The cells targeted were identified via staining with nine different cell surface markers followed by FACS analysis as well as by conventional cytological methods. The targeted cells include B and T lymphocytes, neutrophils, monocytes, and dendritic cells, but not mature macrophages. To further investigate replication in these various cell types, Salmonella derivatives were constructed that express a red fluorescent protein. Bacteria could be seen in each of the cell types above; however, most viable bacteria were present in neutrophils. We find that Salmonella is capable of targeting most phagocytic and non-phagocytic cells in the spleen but has a surprisingly high preference for neutrophils. These findings suggest that Salmonella specifically target splenic neutrophils presumably to attenuate their microbicidal functions, thereby promoting intracellular survival and replication in the mouse. PMID:18159943

  15. Differential response of three cell types to dual stress of nitric oxide and radiation.

    PubMed

    Dhariwala, Fatema A; Narang, Himanshi; Krishna, Malini

    2012-06-01

    The perception of toxicity to nitric oxide (NO) and irradiation (IR) by three different cell types has been studied. The three cell types are the macrophage like RAW264.7 cells, EL4 lymphoma cells, and splenocytes, which represent the different components of a tumor. These three cell types respond differently to NO donors (SNP and SNAP) and radiation treatment. The macrophages were found to be most radio-resistant and insensitive to NO donors. The innate resistance of the macrophages was not due to its antioxidant defense system since there was no significant activation of the enzymes (superoxide dismutases, catalase, and glutathione peroxidase) in RAW264.7 cells after NO donor and irradiation. But the cell cycle arrest of the three cell types was different from each other. The EL4 cells were found to arrest in the G2/M phase while the macrophages were found arrested in the G1 phase of the cell cycle. Such specific killing of the tumor cell in response to NO donor while sparing the macrophages can be of immense importance to radiotherapy.

  16. Proton and Fe Ion-Induced Early and Late Chromosome Aberrations in Different Cell Types

    NASA Technical Reports Server (NTRS)

    Wu, Honglu; Lu, Tao; Yeshitla, Samrawit; Zhang, Ye; Kadhim, Munira

    2016-01-01

    An early stage of cancer development is believed to be genomic instability (GI) which accelerates the mutation rate in the descendants of the cells surviving radiation exposure. To investigate GI induced by charged particles, we exposed human lymphocytes, human fibroblast cells, and human mammary epithelial cells to high energy protons and Fe ions. In addition, we also investigated GI in bone marrow cells isolated from CBA/CaH (CBA) and C57BL/6 (C57) mice, by analyzing cell survival and chromosome aberrations in the cells after multiple cell divisions. Results analyzed so far from the experiments indicated different sensitivities to charged particles between CBA/CaH (CBA) and C57BL/6 (C57) mouse strains, suggesting that there are two main types of response to irradiation: 1) responses associated with survival of damaged cells and 2) responses associated with the induction of non-clonal chromosomal instability in the surviving progeny of stem cells. Previously, we reported that the RBE for initial chromosome damages was high in human lymphocytes exposed to Fe ions. Our results with different cell types demonstrated different RBE values between different cell types and between early and late chromosomal damages. This study also attempts to offer an explanation for the varying RBE values for different cancer types.

  17. GAD65-reactive T cells are activated in patients with autoimmune type 1a diabetes

    PubMed Central

    Viglietta, Vissia; Kent, Sally C.; Orban, Tihamer; Hafler, David A.

    2002-01-01

    Insulin-dependent type 1 diabetes is an autoimmune disease mediated by T lymphocytes recognizing pancreatic islet cell antigens. Glutamic acid decarboxylase 65 (GAD65) appears to be an important autoantigen in the disease. However, T cells from both patients with type 1 diabetes and healthy subjects vigorously proliferate in response to GAD65 stimulation ex vivo, leading us to postulate that the critical event in the onset of human diabetes is the activation of autoreactive T cells. Thus, we investigated whether GAD65-reactive T cells in patients with diabetes functioned as previously activated memory T cells, no longer requiring a second, costimulatory signal for clonal expansion. We found that in patients with new-onset type 1 diabetes, GAD65-reactive T cells were strikingly less dependent on CD28 and B7-1 costimulation to enter into cell cycle and proliferate than were equivalent cells derived from healthy controls. We hypothesize that these autoreactive T cells have been activated in vivo and have differentiated into memory cells, suggesting a pathogenic role in type 1 diabetes. In addition, we observed different effects with selective blockade of either B7-1 or B7-2 molecules; B7-1 appears to deliver a negative signal by engaging CTLA-4, while B7-2 engagement of CD28 upregulates T cell proliferation and cytokine secretion. PMID:11927616

  18. Principles of connectivity among morphologically defined cell types in adult neocortex.

    PubMed

    Jiang, Xiaolong; Shen, Shan; Cadwell, Cathryn R; Berens, Philipp; Sinz, Fabian; Ecker, Alexander S; Patel, Saumil; Tolias, Andreas S

    2015-11-27

    Since the work of Ramón y Cajal in the late 19th and early 20th centuries, neuroscientists have speculated that a complete understanding of neuronal cell types and their connections is key to explaining complex brain functions. However, a complete census of the constituent cell types and their wiring diagram in mature neocortex remains elusive. By combining octuple whole-cell recordings with an optimized avidin-biotin-peroxidase staining technique, we carried out a morphological and electrophysiological census of neuronal types in layers 1, 2/3, and 5 of mature neocortex and mapped the connectivity between more than 11,000 pairs of identified neurons. We categorized 15 types of interneurons, and each exhibited a characteristic pattern of connectivity with other interneuron types and pyramidal cells. The essential connectivity structure of the neocortical microcircuit could be captured by only a few connectivity motifs.

  19. Investigation of the diaphragm-type pressure cell

    NASA Technical Reports Server (NTRS)

    Theodorsen, Theodore

    1932-01-01

    This report relates to various improvements in the process of manufacture of the NACA standard pressure cell. Like most pressure recording devices employing thin diaphragms, they would in general show considerable change in calibration with temperature and also some change of calibration with time or aging effect. The required diaphragm thickness and the desirable rate of mechanical magnification have been determined on the basis of several hundred tests.

  20. Dendritic web-type solar cell mini-modules

    NASA Technical Reports Server (NTRS)

    Campbell, R. B.

    1985-01-01

    Twenty-five minimodules composed of dendritic web solar cells with nominal glass size of 12 by 40 cm were provided for study. The modules were identical with respect to design, materials, and manufacturing and assembly processes to full scale modules. The modules were also electrically functional. These minimodules were fabricated to provide test vehicle for environmental testing and to assess reliability of process and design procedures. The module design and performance are outlined.

  1. Development and testing of shingle-type solar cell molecules

    NASA Technical Reports Server (NTRS)

    Shepard, N. F.

    1978-01-01

    The details of a shingle module design which produces in excess of 97 watts/sq m of module area at 1 kW/sq m insolation and at 60 C are reported. This selected design employs a tempered glass coverplate to provide the primary solar cell structural support. The fabrication and testing of a preproduction module of this design has demonstrated that this selected approach will meet the environmental testing requirements imposed by the contract.

  2. Type I and Type II Interferon Coordinately Regulate Suppressive Dendritic Cell Fate and Function during Viral Persistence.

    PubMed

    Cunningham, Cameron R; Champhekar, Ameya; Tullius, Michael V; Dillon, Barbara Jane; Zhen, Anjie; de la Fuente, Justin Rafael; Herskovitz, Jonathan; Elsaesser, Heidi; Snell, Laura M; Wilson, Elizabeth B; de la Torre, Juan Carlos; Kitchen, Scott G; Horwitz, Marcus A; Bensinger, Steven J; Smale, Stephen T; Brooks, David G

    2016-01-01

    Persistent viral infections are simultaneously associated with chronic inflammation and highly potent immunosuppressive programs mediated by IL-10 and PDL1 that attenuate antiviral T cell responses. Inhibiting these suppressive signals enhances T cell function to control persistent infection; yet, the underlying signals and mechanisms that program immunosuppressive cell fates and functions are not well understood. Herein, we use lymphocytic choriomeningitis virus infection (LCMV) to demonstrate that the induction and functional programming of immunosuppressive dendritic cells (DCs) during viral persistence are separable mechanisms programmed by factors primarily considered pro-inflammatory. IFNγ first induces the de novo development of naive monocytes into DCs with immunosuppressive potential. Type I interferon (IFN-I) then directly targets these newly generated DCs to program their potent T cell immunosuppressive functions while simultaneously inhibiting conventional DCs with T cell stimulating capacity. These mechanisms of monocyte conversion are constant throughout persistent infection, establishing a system to continuously interpret and shape the immunologic environment. MyD88 signaling was required for the differentiation of suppressive DCs, whereas inhibition of stimulatory DCs was dependent on MAVS signaling, demonstrating a bifurcation in the pathogen recognition pathways that promote distinct elements of IFN-I mediated immunosuppression. Further, a similar suppressive DC origin and differentiation was also observed in Mycobacterium tuberculosis infection, HIV infection and cancer. Ultimately, targeting the underlying mechanisms that induce immunosuppression could simultaneously prevent multiple suppressive signals to further restore T cell function and control persistent infections.

  3. Type I and Type II Interferon Coordinately Regulate Suppressive Dendritic Cell Fate and Function during Viral Persistence.

    PubMed

    Cunningham, Cameron R; Champhekar, Ameya; Tullius, Michael V; Dillon, Barbara Jane; Zhen, Anjie; de la Fuente, Justin Rafael; Herskovitz, Jonathan; Elsaesser, Heidi; Snell, Laura M; Wilson, Elizabeth B; de la Torre, Juan Carlos; Kitchen, Scott G; Horwitz, Marcus A; Bensinger, Steven J; Smale, Stephen T; Brooks, David G

    2016-01-01

    Persistent viral infections are simultaneously associated with chronic inflammation and highly potent immunosuppressive programs mediated by IL-10 and PDL1 that attenuate antiviral T cell responses. Inhibiting these suppressive signals enhances T cell function to control persistent infection; yet, the underlying signals and mechanisms that program immunosuppressive cell fates and functions are not well understood. Herein, we use lymphocytic choriomeningitis virus infection (LCMV) to demonstrate that the induction and functional programming of immunosuppressive dendritic cells (DCs) during viral persistence are separable mechanisms programmed by factors primarily considered pro-inflammatory. IFNγ first induces the de novo development of naive monocytes into DCs with immunosuppressive potential. Type I interferon (IFN-I) then directly targets these newly generated DCs to program their potent T cell immunosuppressive functions while simultaneously inhibiting conventional DCs with T cell stimulating capacity. These mechanisms of monocyte conversion are constant throughout persistent infection, establishing a system to continuously interpret and shape the immunologic environment. MyD88 signaling was required for the differentiation of suppressive DCs, whereas inhibition of stimulatory DCs was dependent on MAVS signaling, demonstrating a bifurcation in the pathogen recognition pathways that promote distinct elements of IFN-I mediated immunosuppression. Further, a similar suppressive DC origin and differentiation was also observed in Mycobacterium tuberculosis infection, HIV infection and cancer. Ultimately, targeting the underlying mechanisms that induce immunosuppression could simultaneously prevent multiple suppressive signals to further restore T cell function and control persistent infections. PMID:26808628

  4. Type I and Type II Interferon Coordinately Regulate Suppressive Dendritic Cell Fate and Function during Viral Persistence

    PubMed Central

    Cunningham, Cameron R.; Champhekar, Ameya; Tullius, Michael V.; Dillon, Barbara Jane; Zhen, Anjie; de la Fuente, Justin Rafael; Herskovitz, Jonathan; Elsaesser, Heidi; Snell, Laura M.; Wilson, Elizabeth B.; de la Torre, Juan Carlos; Kitchen, Scott G.; Horwitz, Marcus A.; Bensinger, Steven J.; Smale, Stephen T.; Brooks, David G.

    2016-01-01

    Persistent viral infections are simultaneously associated with chronic inflammation and highly potent immunosuppressive programs mediated by IL-10 and PDL1 that attenuate antiviral T cell responses. Inhibiting these suppressive signals enhances T cell function to control persistent infection; yet, the underlying signals and mechanisms that program immunosuppressive cell fates and functions are not well understood. Herein, we use lymphocytic choriomeningitis virus infection (LCMV) to demonstrate that the induction and functional programming of immunosuppressive dendritic cells (DCs) during viral persistence are separable mechanisms programmed by factors primarily considered pro-inflammatory. IFNγ first induces the de novo development of naive monocytes into DCs with immunosuppressive potential. Type I interferon (IFN-I) then directly targets these newly generated DCs to program their potent T cell immunosuppressive functions while simultaneously inhibiting conventional DCs with T cell stimulating capacity. These mechanisms of monocyte conversion are constant throughout persistent infection, establishing a system to continuously interpret and shape the immunologic environment. MyD88 signaling was required for the differentiation of suppressive DCs, whereas inhibition of stimulatory DCs was dependent on MAVS signaling, demonstrating a bifurcation in the pathogen recognition pathways that promote distinct elements of IFN-I mediated immunosuppression. Further, a similar suppressive DC origin and differentiation was also observed in Mycobacterium tuberculosis infection, HIV infection and cancer. Ultimately, targeting the underlying mechanisms that induce immunosuppression could simultaneously prevent multiple suppressive signals to further restore T cell function and control persistent infections. PMID:26808628

  5. Regulation of expression driven by human immunodeficiency virus type 1 and human T-cell leukemia virus type I long terminal repeats in pluripotential human embryonic cells

    SciTech Connect

    Maio, J.; Brown, F.L. )

    1988-04-01

    Human pluripotential embryonic teratocarcinoma cells differentially expressed gene activity controlled by the human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type I (HTLV-I) long terminal repeats (LTRs) when differentiation was induced by the morphogen all-trans retinoic acid. The alterations occurred after commitment and before the appearance of the multiple cell types characteristic of these pluripotential cells. After commitment, gene activity controlled by the HIV-1 LTR markedly increased, whereas that controlled by the HTLV-I LTR decreased. Steady-state mRNA levels and nuclear run-on transcription indicated that the increased HIV-1-directed activity during differentiation occurred posttranscriptionally, whereas the decreased HTLV-I activity was at the transcriptional level. Phorbol esters did not cause commitment but strongly enhanced expression by both viral LTRs at the transcriptional level. Differentiating cells gradually lost the ability to respond to phorbol ester stimulation. Experiments with a deletion mutant of the HIV-1 LTR suggested that this was due to imposition of negative regulation during differentiation that was not reversed by phorbol ester induction. Cycloheximide, with or without phorbol ester, slightly stimulated HIV-1-directed activity at the transcriptional level and massively increased the amounts of steady-state mRNA by posttranscriptional superinduction. It appeared, however, that new nuclear protein synthesis was required for maximal transcriptional stimulation by phorbol esters. Thus, changing cellular regulatory mechanisms influenced human retrovirus expression during human embryonic cell differentiation.

  6. Epithelial cell types of the primary ureter of Helix aspersa: Ultrastructural and cytochemical characteristics.

    PubMed

    Sánchez-Aguayo, I; Ballesteros, F; Hidalgo, J; López-Campos, J L

    1987-01-01

    The present study describes the morphological characteristics which determine the structural polarity of the principal and ciliated cells in the primary ureter epithelium of Helix aspersa. These characteristics are analysed on the basis of the function performed by both cell types. The presence of paniculate glycogen and the location of glycoconjugates associated with cell membranes of the epithelial cells is revealed by the method of Thiéry.

  7. Decipher the Molecular Response of Plant Single Cell Types to Environmental Stresses

    DOE PAGES

    Nourbakhsh-Rey, Mehrnoush; Libault, Marc

    2016-01-01

    The analysis of the molecular response of entire plants or organs to environmental stresses suffers from the cellular complexity of the samples used. Specifically, this cellular complexity masks cell-specific responses to environmental stresses and logically leads to the dilution of the molecular changes occurring in each cell type composing the tissue/organ/plant in response to the stress. Therefore, to generate a more accurate picture of these responses, scientists are focusing on plant single cell type approaches. Several cell types are now considered as models such as the pollen, the trichomes, the cotton fiber, various root cell types including the root hairmore » cell, and the guard cell of stomata. Among them, several have been used to characterize plant response to abiotic and biotic stresses. Lastly, in this review, we are describing the various -omic studies performed on these different plant single cell type models to better understand plant cell response to biotic and abiotic stresses.« less

  8. Rat coronaviruses infect rat alveolar type I epithelial cells and induce expression of CXC chemokines

    PubMed Central

    Miura, Tanya A.; Wang, Jieru; Holmes, Kathryn V.; Mason, Robert J.

    2007-01-01

    We analyzed the ability of two rat coronavirus (RCoV) strains, sialodacryoadenitis virus (SDAV) and Parker’s RCoV (RCoV-P), to infect rat alveolar type I cells and induce chemokine expression. Primary rat alveolar type II cells were transdifferentiated into the type I cell phenotype. Type I cells were productively infected with SDAV and RCoV-P, and both live virus and UV-inactivated virus induced mRNA and protein expression of three CXC chemokines: CINC-2, CINC-3, and LIX, which are neutrophil chemoattractants. Dual immunolabeling of type I cells for viral antigen and CXC chemokines showed that chemokines were expressed primarily by uninfected cells. Virus-induced chemokine expression was reduced by the IL-1 receptor antagonist, suggesting that IL-1 produced by infected cells induces uninfected cells to express chemokines. Primary cultures of alveolar epithelial cells are an important model for the early events in viral infection that lead to pulmonary inflammation. PMID:17804032

  9. Haemoglobin synthesis in K562 erythroleukaemia cells is affected by intimate contact with monolayers of various human cell types.

    PubMed

    Zuhrie, S R; Pearson, J D; Wickramasinghe, S N

    1988-01-01

    The haemoglobin content of K562 erythroleukaemia cells was affected by co-culture over monolayers of various human cell types. Haemoglobin synthesis was increased after co-culture with umbilical-cord-derived endothelial cells and most monolayers of bone-marrow-derived macrophages, and inhibited after co-culture with two fibroblast lines, blood-monocyte-derived macrophages, a neuroglial cell line (U-251 MG) and most monolayers of bone-marrow-derived stromal cells. These effects were modified when a thin layer of agar was placed over the monolayers. Cell-free culture media conditioned by all but two of the seven types of monolayer studied inhibited haemoglobin synthesis by K562 cells; those conditioned by blood-monocyte-derived macrophages and two of 11 monolayers of bone-marrow-derived macrophages stimulated haemoglobin synthesis. Thus, the haemoglobin content of K562 cells appeared to be influenced both by intimate contact between K562 cells and the cells of the monolayers and by humoral factors released by the monolayers. The data support the concept that erythroid differentiation is partly dependent on intimate contact between erythroid progenitor cells and microenvironmental cells.

  10. Mouse Adenovirus Type 1 Infection of Natural Killer Cell-Deficient Mice

    PubMed Central

    Welton, Amanda R.; Gralinski, Lisa E.; Spindler, Katherine R.

    2008-01-01

    Natural killer (NK) cells contribute to the initial nonspecific response to viral infection, and viruses exhibit a range of sensitivities to NK cells in vivo. We investigated the role of NK cells in infection of mice by mouse adenovirus type 1 (MAV-1) using antibody-mediated depletion and knockout mice. MAV-1 causes encephalomyelitis and replicates to highest levels in brains. NK cell-depleted mice infected with MAV-1 showed brain viral loads 8-20 days p.i. that were similar to wild-type control non-depleted mice. Mice genetically deficient for NK cells behaved similarly to wild-type control mice with respect to brain viral loads and survival. We conclude that NK cells are not required to control virus replication in the brains of MAV-1-infected mice. PMID:18155121

  11. Renal-type clear cell carcinoma of the prostate: a diagnostic challenge.

    PubMed

    Patne, Shashikant C U; Johri, Nidhi; Katiyar, Richa; Trivedi, Sameer; Dwivedi, Uday Shankar

    2015-01-01

    A 72-year-old male presented with urinary symptoms. His serum prostate specific antigen level was 65.2 ng/ml. His radical prostatectomy specimen showed clear cell lesion reminiscent of the clear cell renal cell carcinoma along with acinar type of prostatic adenocarcinoma, Gleason score 4 + 4. The lesional clear cells were positive for pancytokeratin, epithelial membrane antigen, CD10, vimentin, and AMACR while negative for 34βE12, CK7, prostate specific antigen, and PAX8. The final diagnosis was renal-type clear cell carcinoma of the prostate. A follow-up of 20 months did not show metastasis. We herein report fifth case of renal-type clear cell carcinoma of the prostate. PMID:26498435

  12. Fuel cell arrangement

    DOEpatents

    Isenberg, A.O.

    1987-05-12

    A fuel cell arrangement is provided wherein cylindrical cells of the solid oxide electrolyte type are arranged in planar arrays where the cells within a plane are parallel. Planes of cells are stacked with cells of adjacent planes perpendicular to one another. Air is provided to the interior of the cells through feed tubes which pass through a preheat chamber. Fuel is provided to the fuel cells through a channel in the center of the cell stack; the fuel then passes the exterior of the cells and combines with the oxygen-depleted air in the preheat chamber. 3 figs.

  13. Fuel cell arrangement

    DOEpatents

    Isenberg, Arnold O.

    1987-05-12

    A fuel cell arrangement is provided wherein cylindrical cells of the solid oxide electrolyte type are arranged in planar arrays where the cells within a plane are parallel. Planes of cells are stacked with cells of adjacent planes perpendicular to one another. Air is provided to the interior of the cells through feed tubes which pass through a preheat chamber. Fuel is provided to the fuel cells through a channel in the center of the cell stack; the fuel then passes the exterior of the cells and combines with the oxygen-depleted air in the preheat chamber.

  14. Infection of human endothelial cells by human T-lymphotropic virus type I.

    PubMed Central

    Ho, D D; Rota, T R; Hirsch, M S

    1984-01-01

    We studied the effects of human T-lymphotropic virus type I (HTLV-I) on human endothelial cells in vitro. During cocultivation with an HTLV-I producer cell line (C91/PL), endothelial cells formed characteristic multinucleated syncytial giant cells. Inoculation with concentrated cell-free supernatant fluid from C91/PL cultures produced similar cytopathic effects, which were neutralized by pretreatment with HTLV-I specific human serum. HTLV-I antigens were detected in the cytoplasm of the multinucleated cells by indirect immunofluorescence. When endothelial cells showed maximal cytopathic changes, reverse transcriptase activity was demonstrated in the supernatant fluid and HTLV-I was isolated by cocultivation with peripheral blood mononuclear cells. This study demonstrates that HTLV-I tropism is not limited to lymphoid cells but extends to human endothelial cells as well. Images PMID:6095308

  15. Adjacent channel interference degradation with minimum shift keyed modulation

    NASA Technical Reports Server (NTRS)

    Ziemer, R. E.

    1981-01-01

    Computer simulation results for degradation in signal-to-noise ratio for various values of bit error probability are given for minimum shift-keyed-type signaling in the presence of adjacent channel interference. A serial modulator structure which utilizes spectral shaping is characterized in terms of envelope deviation and bandwidth efficiency. This serial generation technique is convenient for implementation at high data rates and results in signal spectra with lower sidelobe levels than conventional minimum shift-keyed modulation at the expense of moderate envelope deviation. Because of the lower sidelobe levels, the resulting spectra allow denser channel packing than does ideal MSK.

  16. Interaction of Cracks Between Two Adjacent Indents in Glass

    NASA Technical Reports Server (NTRS)

    Choi, S. R.; Salem, J. A.

    1993-01-01

    Experimental observations of the interaction behavior of cracks between two adjacent indents were made using an indentation technique in soda-lime glass. It was specifically demonstrated how one indent crack initiates and propagates in the vicinity of another indent crack. Several types of crack interactions were examined by changing the orientation and distance of one indent relative to the other. It was found that the residual stress field produced by elastic/plastic indentation has a significant influence on controlling the mode of crack interaction. The interaction of an indent crack with a free surface was also investigated for glass and ceramic specimens.

  17. Adjacent Segment Pathology after Lumbar Spinal Fusion.

    PubMed

    Lee, Jae Chul; Choi, Sung-Woo

    2015-10-01

    One of the major clinical issues encountered after lumbar spinal fusion is the development of adjacent segment pathology (ASP) caused by increased mechanical stress at adjacent segments, and resulting in various radiographic changes and clinical symptoms. This condition may require surgical intervention. The incidence of ASP varies with both the definition and methodology adopted in individual studies; various risk factors for this condition have been identified, although a significant controversy still exists regarding their significance. Motion-preserving devices have been developed, and some studies have shown their efficacy of preventing ASP. Surgeons should be aware of the risk factors of ASP when planning a surgery, and accordingly counsel their patients preoperatively. PMID:26435804

  18. Adjacent Segment Pathology after Lumbar Spinal Fusion

    PubMed Central

    Lee, Jae Chul

    2015-01-01

    One of the major clinical issues encountered after lumbar spinal fusion is the development of adjacent segment pathology (ASP) caused by increased mechanical stress at adjacent segments, and resulting in various radiographic changes and clinical symptoms. This condition may require surgical intervention. The incidence of ASP varies with both the definition and methodology adopted in individual studies; various risk factors for this condition have been identified, although a significant controversy still exists regarding their significance. Motion-preserving devices have been developed, and some studies have shown their efficacy of preventing ASP. Surgeons should be aware of the risk factors of ASP when planning a surgery, and accordingly counsel their patients preoperatively. PMID:26435804

  19. Photoresponse diversity among the five types of intrinsically photosensitive retinal ganglion cells

    PubMed Central

    Zhao, Xiwu; Stafford, Ben K; Godin, Ashley L; King, W Michael; Wong, Kwoon Y

    2014-01-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) mediate non-image-forming visual responses, including pupillary constriction, circadian photoentrainment and suppression of pineal melatonin secretion. Five morphological types of ipRGCs, M1–M5, have been identified in mice. In order to understand their functions better, we studied the photoresponses of all five cell types, by whole-cell recording from fluorescently labelled ipRGCs visualized using multiphoton microscopy. All ipRGC types generated melanopsin-based (‘intrinsic’) as well as synaptically driven (‘extrinsic’) light responses. The intrinsic photoresponses of M1 cells were lower threshold, higher amplitude and faster than those of M2–M5. The peak amplitudes of extrinsic light responses differed among the ipRGC types; however, the responses of all cell types had comparable thresholds, kinetics and waveforms, and all cells received rod input. While all five types exhibited inhibitory amacrine-cell and excitatory bipolar-cell inputs from the ‘on’ channel, M1 and M3 received additional ‘off’-channel inhibition, possibly through their ‘off’-sublamina dendrites. The M2–M5 ipRGCs had centre–surround-organized receptive fields, implicating a capacity to detect spatial contrast. In contrast, the receptive fields of M1 cells lacked surround antagonism, which might be caused by the surround of the inhibitory input nullifying the surround of the excitatory input. All ipRGCs responded robustly to a wide range of motion speeds, and M1–M4 cells appeared tuned to different speeds, suggesting that they might analyse the speed of motion. Retrograde labelling revealed that M1–M4 cells project to the superior colliculus, suggesting that the contrast and motion information signalled by these cells could be used by this sensorimotor area to detect novel objects and motion in the visual field. PMID:24396062

  20. Ontology based molecular signatures for immune cell types via gene expression analysis

    PubMed Central

    2013-01-01

    Background New technologies are focusing on characterizing cell types to better understand their heterogeneity. With large volumes of cellular data being generated, innovative methods are needed to structure the resulting data analyses. Here, we describe an ‘Ontologically BAsed Molecular Signature’ (OBAMS) method that identifies novel cellular biomarkers and infers biological functions as characteristics of particular cell types. This method finds molecular signatures for immune cell types based on mapping biological samples to the Cell Ontology (CL) and navigating the space of all possible pairwise comparisons between cell types to find genes whose expression is core to a particular cell type’s identity. Results We illustrate this ontological approach by evaluating expression data available from the Immunological Genome project (IGP) to identify unique biomarkers of mature B cell subtypes. We find that using OBAMS, candidate biomarkers can be identified at every strata of cellular identity from broad classifications to very granular. Furthermore, we show that Gene Ontology can be used to cluster cell types by shared biological processes in order to find candidate genes responsible for somatic hypermutation in germinal center B cells. Moreover, through in silico experiments based on this approach, we have identified genes sets that represent genes overexpressed in germinal center B cells and identify genes uniquely expressed in these B cells compared to other B cell types. Conclusions This work demonstrates the utility of incorporating structured ontological knowledge into biological data analysis – providing a new method for defining novel biomarkers and providing an opportunity for new biological insights. PMID:24004649

  1. Photoresponse diversity among the five types of intrinsically photosensitive retinal ganglion cells.

    PubMed

    Zhao, Xiwu; Stafford, Ben K; Godin, Ashley L; King, W Michael; Wong, Kwoon Y

    2014-04-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) mediate non-image-forming visual responses, including pupillary constriction, circadian photoentrainment and suppression of pineal melatonin secretion. Five morphological types of ipRGCs, M1-M5, have been identified in mice. In order to understand their functions better, we studied the photoresponses of all five cell types, by whole-cell recording from fluorescently labelled ipRGCs visualized using multiphoton microscopy. All ipRGC types generated melanopsin-based ('intrinsic') as well as synaptically driven ('extrinsic') light responses. The intrinsic photoresponses of M1 cells were lower threshold, higher amplitude and faster than those of M2-M5. The peak amplitudes of extrinsic light responses differed among the ipRGC types; however, the responses of all cell types had comparable thresholds, kinetics and waveforms, and all cells received rod input. While all five types exhibited inhibitory amacrine-cell and excitatory bipolar-cell inputs from the 'on' channel, M1 and M3 received additional 'off'-channel inhibition, possibly through their 'off'-sublamina dendrites. The M2-M5 ipRGCs had centre-surround-organized receptive fields, implicating a capacity to detect spatial contrast. In contrast, the receptive fields of M1 cells lacked surround antagonism, which might be caused by the surround of the inhibitory input nullifying the surround of the excitatory input. All ipRGCs responded robustly to a wide range of motion speeds, and M1-M4 cells appeared tuned to different speeds, suggesting that they might analyse the speed of motion. Retrograde labelling revealed that M1-M4 cells project to the superior colliculus, suggesting that the contrast and motion information signalled by these cells could be used by this sensorimotor area to detect novel objects and motion in the visual field. PMID:24396062

  2. Some effects of vitamin A deficiency on the isolated rat lung alveolar type II cell.

    PubMed

    Zachman, R D; Chen, X; Verma, A K; Grummer, M A

    1992-01-01

    Alveolar Type II cells were isolated from control and vitamin A deficient rats and allowed to form a monolayer in plastic dishes for 16-18 hours. The vitamin A content (retinol plus retinyl palmitate) of deficient cells was 50-75% less than in control cells on a per mg protein basis. Isolated Type II cells took up [3H]-retinol, synthesized [3H]-retinyl palmitate, and after 4 hours, 24% of the radioactivity in the Type II cells was [3H]-retinoic acid. Deficiency did not appear to alter retinoic acid synthesis. Phosphatidylcholine (PC) and disaturated phosphatidylcholine (DSPC) synthesis, were slightly less in deficient cells compared to control (95 and 85% respectively). In addition, 10(-6) M and 10(-5) M retinoic acid in the reaction media stimulated both PC and DSPC synthesis by 120-140% in control cells. The stimulating effect of retinoic acid was present in deficient cells as well, but less pronounced (120% with 10(-5) M). Vitamin A deficient Type II cells also had less basal levels of both tissue transglutaminase and epidermal transglutaminase activity than control cells. PMID:1355470

  3. Nectin spot: a novel type of nectin-mediated cell adhesion apparatus.

    PubMed

    Mizutani, Kiyohito; Takai, Yoshimi

    2016-09-15

    Nectins are Ca(2+)-independent immunoglobulin (Ig) superfamily cell adhesion molecules constituting a family with four members, all of which have three Ig-like loops at their extracellular regions. Nectins play roles in the formation of a variety of cell-cell adhesion apparatuses. There are at least three types of nectin-mediated cell adhesions: afadin- and cadherin-dependent, afadin-dependent and cadherin-independent, and afadin- and cadherin-independent. In addition, nectins trans-interact with nectin-like molecules (Necls) with three Ig-like loops and other Ig-like molecules with one to three Ig-like loops. Furthermore, nectins and Necls cis-interact with membrane receptors and integrins, some of which are associated with the nectin-mediated cell adhesions, and play roles in the regulation of many cellular functions, such as cell polarization, movement, proliferation, differentiation, and survival, co-operatively with these cell surface proteins. The nectin-mediated cell adhesions are implicated in a variety of diseases, including genetic disorders, neural disorders, and cancers. Of the three types of nectin-mediated cell adhesions, the afadin- and cadherin-dependent apparatus has been most extensively investigated, but the examples of the third type of apparatus independent of afadin and cadherin are recently increasing and its morphological and functional properties have been well characterized. We review here recent advances in research on this type of nectin-mediated cell adhesion apparatus, which is named nectin spot. PMID:27621480

  4. Adenovirus vectors targeting distinct cell types in the retina.

    PubMed

    Sweigard, J Harry; Cashman, Siobhan M; Kumar-Singh, Rajendra

    2010-04-01

    Purpose. Gene therapy for a number of retinal diseases necessitates efficient transduction of photoreceptor cells. Whereas adenovirus (Ad) serotype 5 (Ad5) does not transduce photoreceptors efficiently, previous studies have demonstrated improved photoreceptor transduction by Ad5 pseudotyped with Ad35 (Ad5/F35) or Ad37 (Ad5/F37) fiber or by the deletion of the RGD domain in the Ad5 penton base (Ad5DeltaRGD). However, each of these constructs contained a different transgene cassette, preventing the evaluation of the relative performance of these vectors, an important consideration before the use of these vectors in the clinic. The aim of this study was to evaluate these vectors in the retina and to attempt photoreceptor-specific transgene expression. Methods. Three Ad5-based vectors containing the same expression cassette were generated and injected into the subretinal space of adult mice. Eyes were analyzed for green fluorescence protein expression in flat-mounts, cross-sections, quantitative RT-PCR, and a modified stereological technique. A 257-bp fragment derived from the mouse opsin promoter was analyzed in the context of photoreceptor-specific transgene expression. Results. Each virus tested efficiently transduced the retinal pigment epithelium. The authors found no evidence that Ad5/F35 or Ad5/F37 transduced photoreceptors. Instead, they found that Ad5/F37 transduced Müller cells. Robust photoreceptor transduction by Ad5DeltaRGD was detected. Photoreceptor-specific transgene expression from the 257-bp mouse opsin promoter in the context of Ad5DeltaRGD vectors was found. Conclusions. Adenovirus vectors may be designed with tropism to distinct cell populations. Robust photoreceptor-specific transgene expression can be achieved in the context of Ad5DeltaRGD vectors.

  5. Cell type specific gene expression analysis of prostate needle biopsies resolves tumor tissue heterogeneity.

    PubMed

    Krönig, Malte; Walter, Max; Drendel, Vanessa; Werner, Martin; Jilg, Cordula A; Richter, Andreas S; Backofen, Rolf; McGarry, David; Follo, Marie; Schultze-Seemann, Wolfgang; Schüle, Roland

    2015-01-20

    A lack of cell surface markers for the specific identification, isolation and subsequent analysis of living prostate tumor cells hampers progress in the field. Specific characterization of tumor cells and their microenvironment in a multi-parameter molecular assay could significantly improve prognostic accuracy for the heterogeneous prostate tumor tissue. Novel functionalized gold-nano particles allow fluorescence-based detection of absolute mRNA expression levels in living cells by fluorescent activated flow cytometry (FACS). We use of this technique to separate prostate tumor and benign cells in human prostate needle biopsies based on the expression levels of the tumor marker alpha-methylacyl-CoA racemase (AMACR). We combined RNA and protein detection of living cells by FACS to gate for epithelial cell adhesion molecule (EPCAM) positive tumor and benign cells, EPCAM/CD45 double negative mesenchymal cells and CD45 positive infiltrating lymphocytes. EPCAM positive epithelial cells were further sub-gated into AMACR high and low expressing cells. Two hundred cells from each population and several biopsies from the same patient were analyzed using a multiplexed gene expression profile to generate a cell type resolved profile of the specimen. This technique provides the basis for the clinical evaluation of cell type resolved gene expression profiles as pre-therapeutic prognostic markers for prostate cancer.

  6. Cell type specific gene expression analysis of prostate needle biopsies resolves tumor tissue heterogeneity

    PubMed Central

    Krönig, Malte; Walter, Max; Drendel, Vanessa; Werner, Martin; Jilg, Cordula A.; Richter, Andreas S.; Backofen, Rolf; McGarry, David; Follo, Marie; Schultze-Seemann, Wolfgang; Schüle, Roland

    2015-01-01

    A lack of cell surface markers for the specific identification, isolation and subsequent analysis of living prostate tumor cells hampers progress in the field. Specific characterization of tumor cells and their microenvironment in a multi-parameter molecular assay could significantly improve prognostic accuracy for the heterogeneous prostate tumor tissue. Novel functionalized gold-nano particles allow fluorescence-based detection of absolute mRNA expression levels in living cells by fluorescent activated flow cytometry (FACS). We use of this technique to separate prostate tumor and benign cells in human prostate needle biopsies based on the expression levels of the tumor marker alpha-methylacyl-CoA racemase (AMACR). We combined RNA and protein detection of living cells by FACS to gate for epithelial cell adhesion molecule (EPCAM) positive tumor and benign cells, EPCAM/CD45 double negative mesenchymal cells and CD45 positive infiltrating lymphocytes. EPCAM positive epithelial cells were further sub-gated into AMACR high and low expressing cells. Two hundred cells from each population and several biopsies from the same patient were analyzed using a multiplexed gene expression profile to generate a cell type resolved profile of the specimen. This technique provides the basis for the clinical evaluation of cell type resolved gene expression profiles as pre-therapeutic prognostic markers for prostate cancer. PMID:25514598

  7. Establishment of a vascular endothelial cell-reactive type II NKT cell clone from a rat model of autoimmune vasculitis.

    PubMed

    Iinuma, Chihiro; Waki, Masashi; Kawakami, Ai; Yamaguchi, Madoka; Tomaru, Utano; Sasaki, Naomi; Masuda, Sakiko; Matsui, Yuki; Iwasaki, Sari; Baba, Tomohisa; Kasahara, Masanori; Yoshiki, Takashi; Paletta, Daniel; Herrmann, Thomas; Ishizu, Akihiro

    2015-02-01

    We previously generated a rat model that spontaneously developed small vessel vasculitis (SVV). In this study, a T cell clone reactive with rat vascular endothelial cells (REC) was established and named VASC-1. Intravenous injection of VASC-1 induced SVV in normal recipients. VASC-1 was a TCRαβ/CD3-positive CD4/CD8 double-negative T cell clone with expression of NKG2D. The cytokine mRNA profile under unstimulated condition was positive for IL-4 and IFN-γ but negative for IL-2 and IL-10. After interaction with REC, the mRNA expression of IL-2, IL-5 and IL-6 was induced in VASC-1, which was inhibited by blocking of CD1d on the REC surface. Although the protein levels of these cytokines seemed to be lower than the detection limit in the culture medium, IFN-γ was detectable. The production of IFN-γ from the VASC-1 stimulated with LPS-pre-treated REC was inhibited by the CD1d blockade on the REC. These findings indicated VASC-1 as an NKT cell clone. The NKT cell pool includes two major subsets, namely types I and II. Type I NKT cells are characterized by expression of semi-invariant TCRs and the potential to bind to marine sponge-derived α-galactosylceramide (α-GalCer) loaded on CD1d; whereas, type II NKT cells do not manifest these characteristics. VASC-1 exhibited a usage of TCR other than the type I invariant TCR α chain and did not bind to α-GalCer-loaded CD1d; therefore, it was determined as a type II NKT cell clone. The collective evidence suggested that REC-reactive type II NKT cells could be involved in the pathogenesis of SVV in rats.

  8. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells

    PubMed Central

    Salopiata, Florian; Depner, Sofia; Wäsch, Marvin; Böhm, Martin E.; Mücke, Oliver; Plass, Christoph; Lehmann, Wolf D.; Kreutz, Clemens; Timmer, Jens; Klingmüller, Ursula

    2016-01-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  9. Wild-type human p53 transactivates the human proliferating cell nuclear antigen promoter

    SciTech Connect

    Shivakumar, C.V.; Brown, D.R.; Deb, S.; Deb, S.P.

    1995-12-01

    The p53 tumor suppressor protein negatively regulates cell growth and somatic mutations in the p53 gene lead to uncontrolled cell growth and oncogenesis. This report describes research which demonstrates, using a number of different cell lines, that at low levels, wild-type p53 transactivates the human proliferating cell nuclear antigen (PCNA) promoter. When expressed at similar levels, tumor-derived p53 mutants did not transactivate the PCNA promoter. It also reports the identification of a wild-type human p53-binding site on the human PCNA promote. 84 refs., 5 figs, 3 tabs.

  10. Replication of human immunodeficiency virus type 1 in primary dendritic cell cultures.

    PubMed Central

    Langhoff, E; Terwilliger, E F; Bos, H J; Kalland, K H; Poznansky, M C; Bacon, O M; Haseltine, W A

    1991-01-01

    The ability of the human immunodeficiency virus type 1 (HIV-1) to replicate in primary blood dendritic cells was investigated. Dendritic cells compose less than 1% of the circulating leukocytes and are nondividing cells. Highly purified preparations of dendritic cells were obtained using recent advances in cell fractionation. The results of these experiments show that dendritic cells, in contrast to monocytes and T cells, support the active replication of all strains of HIV-1 tested, including T-cell tropic and monocyte/macrophage tropic isolates. The dendritic cell cultures supported much more virus production than did cultures of primary unseparated T cells, CD4+ T cells, and adherent as well as nonadherent monocytes. Replication of HIV-1 in dendritic cells produces no noticeable cytopathic effect nor does it decrease total cell number. The ability of the nonreplicating dendritic cells to support high levels of replication of HIV-1 suggests that this antigen-presenting cell population, which is also capable of supporting clonal T-cell growth, may play a central role in HIV pathogenesis, serving as a source of continued infection of CD4+ T cells and as a reservoir of virus infection. Images PMID:1910172

  11. In vitro infection of natural killer cells with different human immunodeficiency virus type 1 isolates.

    PubMed Central

    Chehimi, J; Bandyopadhyay, S; Prakash, K; Perussia, B; Hassan, N F; Kawashima, H; Campbell, D; Kornbluth, J; Starr, S E

    1991-01-01

    Natural killer (NK) cells are a discrete subset of leukocytes, distinct from T and B lymphocytes. NK cells mediate spontaneous non-MHC-restricted killing of a wide variety of target cells without prior sensitization and appear to be involved in initial protection against certain viral infections. Depressed NK cell-mediated cytotoxicity, one of the many immunological defects observed in AIDS patients, may contribute to secondary virus infections. Here we report that clonal and purified polyclonal populations of NK cells, which expressed neither surface CD4 nor CD4 mRNA, were susceptible to infection with various isolates of human immunodeficiency virus type 1 (HIV-1). Viral replication was demonstrated by detection of p24 antigen intracellularly and in culture supernatants, by the presence of HIV DNA within infected cells, and by the ability of supernatants derived from HIV-infected NK cells to infect peripheral blood mononuclear cells or CD4+ cell lines. Infection of NK cells was not blocked by anti-CD4 or anti-Fc gamma RIII monoclonal antibodies. NK cells from HIV-infected and uninfected cultures were similar in their ability to lyse three different target cells. Considerable numbers of cells died in HIV-infected NK cell cultures. These results suggest that loss of NK cells in AIDS patients is a direct effect of HIV infection but that reduced NK cell function involves another mechanism. The possibility that NK cells serve as a potential reservoir for HIV-1 must be considered. Images PMID:1672164

  12. Cell Type-Specific Modulation of Respiratory Chain Supercomplex Organization

    PubMed Central

    Sun, Dayan; Li, Bin; Qiu, Ruyi; Fang, Hezhi; Lyu, Jianxin

    2016-01-01

    Respiratory chain complexes are organized into large supercomplexes among which supercomplex In + IIIn + IVn is the only one that can directly transfer electrons from NADH to oxygen. Recently, it was reported that the formation of supercomplex In + IIIn + IVn in mice largely depends on their genetic background. However, in this study, we showed that the composition of supercomplex In + IIIn + IVn is well conserved in various mouse and human cell lines. Strikingly, we found that a minimal supercomplex In + IIIn, termed “lowest supercomplex” (LSC) in this study because of its migration at the lowest position close to complex V dimers in blue native polyacrylamide gel electrophoresis, was associated with complex IV to form a supercomplex In + IIIn + IVn in some, but not all of the human and mouse cells. In addition, we observed that the 3697G>A mutation in mitochondrial-encoded NADH dehydrogenase 1 (ND1) in one patient with Leigh’s disease specifically affected the assembly of supercomplex In + IIIn + IVn containing LSC, leading to decreased cellular respiration and ATP generation. In conclusion, we showed the existence of LSC In + IIIn + IVn and impairment of this supercomplex causes disease. PMID:27338358

  13. Cell Type-Specific Modulation of Respiratory Chain Supercomplex Organization.

    PubMed

    Sun, Dayan; Li, Bin; Qiu, Ruyi; Fang, Hezhi; Lyu, Jianxin

    2016-01-01

    Respiratory chain complexes are organized into large supercomplexes among which supercomplex In + IIIn + IVn is the only one that can directly transfer electrons from NADH to oxygen. Recently, it was reported that the formation of supercomplex In + IIIn + IVn in mice largely depends on their genetic background. However, in this study, we showed that the composition of supercomplex In + IIIn + IVn is well conserved in various mouse and human cell lines. Strikingly, we found that a minimal supercomplex In + IIIn, termed "lowest supercomplex" (LSC) in this study because of its migration at the lowest position close to complex V dimers in blue native polyacrylamide gel electrophoresis, was associated with complex IV to form a supercomplex In + IIIn + IVn in some, but not all of the human and mouse cells. In addition, we observed that the 3697G>A mutation in mitochondrial-encoded NADH dehydrogenase 1 (ND1) in one patient with Leigh's disease specifically affected the assembly of supercomplex In + IIIn + IVn containing LSC, leading to decreased cellular respiration and ATP generation. In conclusion, we showed the existence of LSC In + IIIn + IVn and impairment of this supercomplex causes disease. PMID:27338358

  14. Type I interferons produced by dendritic cells promote their phenotypic and functional activation.

    PubMed

    Montoya, Maria; Schiavoni, Giovanna; Mattei, Fabrizio; Gresser, Ion; Belardelli, Filippo; Borrow, Persephone; Tough, David F

    2002-05-01

    Resting dendritic cells (DCs) are resident in most tissues and can be activated by environmental stimuli to mature into potent antigen-presenting cells. One important stimulus for DC activation is infection; DCs can be triggered through receptors that recognize microbial components directly or by contact with infection-induced cytokines. We show here that murine DCs undergo phenotypic maturation upon exposure to type I interferons (type I IFNs) in vivo or in vitro. Moreover, DCs either derived from bone marrow cells in vitro or isolated from the spleens of normal animals express IFN-alpha and IFN-beta, suggesting that type I IFNs can act in an autocrine manner to activate DCs. Consistent with this idea, the ability to respond to type I IFN was required for the generation of fully activated DCs from bone marrow precursors, as DCs derived from the bone marrow of mice lacking a functional receptor for type I IFN had reduced expression of costimulatory and adhesion molecules and a diminished ability to stimulate naive T-cell proliferation compared with DCs derived from control bone marrow. Furthermore, the addition of neutralizing anti-IFN-alpha/beta antibody to purified splenic DCs in vitro partially blocked the "spontaneous" activation of these cells, inhibiting the up-regulation of costimulatory molecules, secretion of IFN-gamma, and T-cell stimulatory activity. These results show that DCs both secrete and respond to type I IFN, identifying type I interferons as autocrine DC activators. PMID:11964292

  15. Electrical filtering in gerbil isolated type I semicircular canal hair cells

    NASA Technical Reports Server (NTRS)

    Rennie, K. J.; Ricci, A. J.; Correia, M. J.

    1996-01-01

    1. Membrane potential responses of dissociated gerbil type I semicircular canal hair cells to current injections in whole cell current-clamp have been measured. The input resistance of type I cells was 21.4 +/- 14.3 (SD) M omega, (n = 25). Around the zero-current potential (Vz = -66.6 +/- 9.3 mV, n = 25), pulsed current injections (from approximately -200 to 750 pA) produced only small-amplitude, pulse-like changes in membrane potential. 2. Injecting constant current to hyperpolarize the membrane to around -100 mV resulted in a approximately 10-fold increase in membrane resistance. Current pulses superimposed on this constant hyperpolarization produced larger and more complex membrane potential changes. Depolarizing currents > or = 200 pA caused a rapid transient peak voltage before a plateau. 3. Membrane voltage was able to faithfully follow sine-wave current injections around Vz over the range 1-1,000 Hz with < 25% attenuation at 1 kHz. A previously described K conductance, IKI, which is active at Vz, produces the low input resistance and frequency response. This was confirmed by pharmacologically blocking IKI. This conductance, present in type I cells but not type II hair cells, would appear to confer on type I cells a lower gain, but a much broader bandwidth at Vz, than seen in type II cells.

  16. Experimental radiation pneumonitis. Corticosteroids increase the replicative activity of alveolar type 2 cells

    SciTech Connect

    Gross, N.J.; Narine, K.R.

    1988-09-01

    Corticosteroid administration during radiation pneumonitis in mice markedly improves the physiologic abnormalities and decreases mortality, an effect that has been attributed to the stimulation of surfactant synthesis and secretion by type 2 alveolar epithelial cells. In the present experiments we explored the effects of corticosteroids on the replicative activity of type 2 cells of lethally irradiated lungs at the height of the radiation reaction. The labeling index of type 2 cells of irradiated mice was increased threefold above that of sham-irradiated controls. Corticosteroids given continuously from 10 weeks after thoracic irradiation further increased the type 2 cell labeling index another threefold above that of irradiated untreated mice. The enhanced reproductive activity of type 2 cells following thoracic irradiation is seen as a protective response that is augmented by corticosteroids, whose effect may be both to improve the physiology of the alveolar surface and to maintain the population of alveolar epithelial cells. The bearing of this result on the controversial role of the type 2 cell as a target in radiation pneumonitis is discussed.

  17. Cells that emerge from embryonic explants produce fibers of type IV collagen.

    PubMed

    Chen, J M; Little, C D

    1985-10-01

    Double immunofluorescence staining experiments designed to examine the synthesis and deposition of collagen types I and IV in cultured explants of embryonic mouse lung revealed the presence of connective tissue-like fibers that were immunoreactive with anti-type IV collagen antibodies. This observation is contrary to the widely accepted belief that type IV collagen is found only in sheet-like arrangements beneath epithelia or as a sheath-like layer enveloping bundles of nerve or muscle cells. The extracellular matrix produced by cells that migrate from embryonic mouse lung rudiments in vitro was examined by double indirect immunofluorescence microscopy. Affinity-purified monospecific polyclonal antibodies were used to examine cells after growth on glass or native collagen substrata. The data show that embryonic mesenchymal cells can produce organized fibers of type IV collagen that are not contained within a basement membrane, and that embryonic epithelial cells deposit fibers and strands of type IV collagen beneath their basal surface when grown on glass; however, when grown on a rat tail collagen substratum the epithelial cells produce a fine meshwork. To our knowledge this work represents the first report that type IV collagen can be organized by cells into a fibrous extracellular matrix that is not a basement membrane.

  18. Cell of origin of distinct cultured rat liver epithelial cells, as typed by cytokeratin and surface component selective expression.

    PubMed

    Marceau, N; Germain, L; Goyette, R; Noël, M; Gourdeau, H

    1986-08-01

    The cell of origin of the nonparenchymal epithelioid cells that emerge in liver cell cultures is unknown. Cultures of rat hepatocytes and several types of nonparenchymal cells obtained by selective tissue dispersion procedures were typed with monoclonal antibodies to rat liver cytokeratin and vimentin, polyvalent antibodies to cow hoof cytokeratins and porcine lens vimentin, and monoclonal antibodies to surface membrane components of ductular oval cells and hepatocytes. Immunoblot analysis revealed that, in cultured rat liver nonparenchymal epithelial cells, the anti-rat hepatocyte cytokeratin antibody recognized a cytokeratin of relative mass (Mr) 55,000 and the anti-cow hoof cytokeratin antibody reacted with a cytokeratin of Mr 52,000, while the anti-vimentin antibodies detected vimentin in both cultured rat fibroblasts and nonparenchymal epithelial cells. Analyses on the specificity of anti-cytokeratin and anti-vimentin antibodies toward the various cellular structures of liver by double immunofluorescence staining of frozen tissue sections revealed unique reactivity patterns. For example, hepatocytes were only stained with anti-Mr 55,000 cytokeratin antibody, while the sinusoidal cells reacted only with the anti-vimentin antibodies. In contrast, epithelial cells of the bile ductular structures and mesothelial cells of the Glisson capsula reacted with all the anti-cytokeratin and anti-vimentin antibodies. It should be stressed, however, that the reaction of the anti-vimentin antibodies on bile ductular cells was weak. The same analysis on tissue sections using the anti-ductular oval cell antibody revealed that it reacted with bile duct structures but not with the Glisson capsula. The anti-hepatocyte antibody reacted only with the parenchymal cells. The differential reactivity of the anti-cytokeratin and anti-vimentin antibodies with the various liver cell compartments was confirmed in primary cultures of hepatocytes, sinusoidal cells, and bile ductular cells

  19. Gentamicin is primarily localized in vestibular type I hair cells after intratympanic administration.

    PubMed

    Lyford-Pike, Sofia; Vogelheim, Casey; Chu, Eugene; Della Santina, Charles C; Carey, John P

    2007-12-01

    Intratympanic (IT) gentamicin injections are effective in the control of episodic vertigo due to Ménière's disease. Histological studies in animals have found that the loss of type I vestibular hair cells far exceeds that of type II cells after IT gentamicin treatment. The objective of this study was to determine whether this selective toxicity for type I hair cells might be due to selective concentration of the drug by these cells. Gentamicin was localized within the vestibular epithelium by both direct and indirect methods. Gentamicin conjugated to Texas Red(R) was used as a direct tracer, and anti-gentamicin antibody provided an indirect means of localization. Conjugated or unconjugated gentamicin was injected into the left tympanic space of chinchillas. The animals were killed and fixed 1 or 3 weeks post-treatment. Confocal fluorescence microscopy was used to determine the localization of gentamicin in semicircular canal cristae. Results from the animals killed within 1 week of administration showed that numerous type I hair cells still remained throughout the epithelium. The mean intensity in grayscale units (0-255) of anti-gentamicin labeling for type I hair cells was 28.14 (95% CI 24.60-31.69), for type II hair cells was 17.09 (14.99-19.20), and for support cells was 5.35 (5.34-5.46; p < 0.001, ANOVA). Anti-gentamicin antibody labeling appeared in the majority of type I hair cells throughout their cytoplasm, but with greater intensity at the apex (p < 0.001). Intensity of fluorescence with Texas-Red conjugated gentamicin was 25.38 (22.83-27.94) in type I hair cells, 15.60 (14.73-16.48) in type II cells, and 12.62 (12.06-13.17) in support cells (p < 0.001, ANOVA). These results suggest that type I hair cells are more susceptible to gentamicin because they more avidly take up or retain the drug in the early period after administration. PMID:17899270

  20. Characteristics of a human cell line transformed by DNA from human adenovirus type 5.

    PubMed

    Graham, F L; Smiley, J; Russell, W C; Nairn, R

    1977-07-01

    Human embryonic kidney cells have been transformed by exposing cells to sheared fragments of adenovirus type 5 DNA. The transformed cells (designated 293 cells) exhibited many of the characteristics of transformation including the elaboration of a virus-specific tumour antigen. Analysis of the polypeptides synthesized in the 293 cells by labelling with 35S-methionine and SDS PAGE showed a variable pattern of synthesis, different in a number of respects from that seen in otheruman cells. On labelling the surface of cells by lactoperoxidase catalysed radio-iodination, the absence of a labelled polypeptide analogous to the 250 K (LETS) glycoprotein was noted. Hybridization of labelled cellular RNA with restriction fragments of adenovirus type 5 DNA indicated transcription of a portion of the adenovirus genome at the conventional left hand end. PMID:886304

  1. Fireworks in the primate retina: in vitro photodynamics reveals diverse LGN-projecting ganglion cell types.

    PubMed

    Dacey, Dennis M; Peterson, Beth B; Robinson, Farrel R; Gamlin, Paul D

    2003-01-01

    Diverse cell types and parallel pathways are characteristic of the vertebrate nervous system, yet it remains a challenge to define the basic components of most neural structures. We describe a process termed retrograde photodynamics that allowed us to rapidly make the link between morphology, physiology, and connectivity for ganglion cells in the macaque retina that project to the lateral geniculate nucleus (LGN). Rhodamine dextran injected into the LGN was transported retrogradely and sequestered within the cytoplasm of ganglion cell bodies. Exposure of the retina to light in vitro liberated the tracer and allowed it to diffuse throughout the dendrites, revealing the cell's complete morphology. Eight previously unknown LGN-projecting cell types were identified. Cells could also be targeted in vitro for intracellular recording and physiological analysis. The photodynamic process was also observed in pyramidal cells in a rat neocortical slice.

  2. Methylated DNA-binding protein is present in various mammalian cell types

    SciTech Connect

    Supakar, P.C.; Weist, D.; Zhang, D.; Inamdar, N.; Zhang, Xianyang; Khan, R.; Ehrlich, M. ); Ehrlich, K.C. )

    1988-08-25

    A DNA-binding protein from human placenta, methylated DNA-binding protein (MDBP), binds to certain DNA sequences only when they contain 5-methylcytosine (m{sup 5}C) residues at specific positions. The authors found a very similar DNA-binding activity in nuclear extracts of rat tissues, calf thymus, human embryonal carcinoma cells, HeLa cells, and mouse LTK cells. Like human placental MDBP, the analogous DNA-binding proteins from the above mammalian cell lines formed a number of different low-electrophoretic-mobility complexes with a 14-bp MDBP-specific oligonucleotide duplex. All of these complexes exhibited the same DNA methylation specificity and DNA sequence specificity. Although MDBP activity was found in various mammalian cell types, it was not detected in extracts of cultured mosquito cells and so may be associated only with cells with vertebrate-type DNA methylation.

  3. A Generic and Cell-Type-Specific Wound Response Precedes Regeneration in Planarians.

    PubMed

    Wurtzel, Omri; Cote, Lauren E; Poirier, Amber; Satija, Rahul; Regev, Aviv; Reddien, Peter W

    2015-12-01

    Regeneration starts with injury. Yet how injuries affect gene expression in different cell types and how distinct injuries differ in gene expression remain unclear. We defined the transcriptomes of major cell types of planarians--flatworms that regenerate from nearly any injury--and identified 1,214 tissue-specific markers across 13 cell types. RNA sequencing on 619 single cells revealed that wound-induced genes were expressed either in nearly all cell types or specifically in one of three cell types (stem cells, muscle, or epidermis). Time course experiments following different injuries indicated that a generic wound response is activated with any injury regardless of the regenerative outcome. Only one gene, notum, was differentially expressed early between anterior- and posterior-facing wounds. Injury-specific transcriptional responses emerged 30 hr after injury, involving context-dependent patterning and stem-cell-specialization genes. The regenerative requirement of every injury is different; however, our work demonstrates that all injuries start with a common transcriptional response.

  4. Vitamin E alters alveolar type II cell phospholipid synthesis in oxygen and air

    SciTech Connect

    Kennedy, K.A.; Snyder, J.M.; Stenzel, W.; Saito, K.; Warshaw, J.B. )

    1990-11-01

    Newborn rats were injected with vitamin E or placebo daily until 6 days after birth. The effect of vitamin E pretreatment on in vitro surfactant phospholipid synthesis was examined in isolated type II cells exposed to oxygen or air form 24 h in vitro. Type II cells were also isolated from untreated 6-day-old rats and cultured for 24 h in oxygen or air with control medium or vitamin E supplemented medium. These cells were used to examine the effect of vitamin E exposure in vitro on type II cell phospholipid synthesis and ultrastructure. Phosphatidylcholine (PC) synthesis was reduced in cells cultured in oxygen as compared with air. This decrease was not prevented by in vivo pretreatment or in vitro supplementation with vitamin E. Vitamin E pretreatment increased the ratio of disaturated PC to total PC and increased phosphatidylglycerol synthesis. The volume density of lamellar bodies in type II cells was increased in cells maintained in oxygen. Vitamin E did not affect the volume density of lamellar bodies. We conclude that in vitro hyperoxia inhibits alveolar type II cell phosphatidylcholine synthesis without decreasing lamellar body volume density and that supplemental vitamin E does not prevent hyperoxia-induced decrease in phosphatidylcholine synthesis.

  5. Neuronal merlin influences ERBB2 receptor expression on Schwann cells through neuregulin 1 type III signalling

    PubMed Central

    Schulz, Alexander; Kyselyova, Anna; Baader, Stephan L.; Jung, Marie Juliane; Zoch, Ansgar; Mautner, Victor-Felix

    2014-01-01

    Axonal surface proteins encompass a group of heterogeneous molecules, which exert a variety of different functions in the highly interdependent relationship between axons and Schwann cells. We recently revealed that the tumour suppressor protein merlin, mutated in the hereditary tumour syndrome neurofibromatosis type 2, impacts significantly on axon structure maintenance in the peripheral nervous system. We now report on a role of neuronal merlin in the regulation of the axonal surface protein neuregulin 1 important for modulating Schwann cell differentiation and myelination. Specifically, neuregulin 1 type III expression is reduced in sciatic nerve tissue of neuron-specific knockout animals as well as in biopsies from seven patients with neurofibromatosis type 2. In vitro experiments performed on both the P19 neuronal cell line and primary dorsal root ganglion cells demonstrate the influence of merlin on neuregulin 1 type III expression. Moreover, expression of ERBB2, a Schwann cell receptor for neuregulin 1 ligands is increased in nerve tissue of both neuron-specific merlin knockout animals and patients with neurofibromatosis type 2, demonstrating for the first time that axonal merlin indirectly regulates Schwann cell behaviour. Collectively, we have identified that neuronally expressed merlin can influence Schwann cell activity in a cell-extrinsic manner. PMID:24309211

  6. Effect of type 2 cell mitosis on the surfactant system of injured mouse lungs

    SciTech Connect

    Smith, L.J.

    1983-09-01

    This study was designed to evaluate the effect of type 2 cell proliferation, and specifically mitosis, on the surfactant system after lung injury. Lung injury was produced in mice with butylated hydroxytoluene (BHT). The lamellar body (LB) volume density and the LB area of tritiated thymidine (/sup 3/H-T) labeled and mitotic type 2 cells were determined by combining light microscopic autoradiography with electron microscopic morphometry. Over a 48-hour period, the LB volume density of proliferating (/sup 3/H-T-labeled) type 2 cells decreased from 20.7% to 7.6% and the LB area per cell decreased from 9.1 to 2.4 ..mu..m/sup 2/. These changes were closely related to type 2 cell mitosis, since the LB volume density decreased from 19.2% to 2.9% and the LB area per cell decreased from 9.1 to 1.7 ..mu.. m/sup 2/ between prophase and telophase, but they were independent of the time elapsed since injury. These results indicate that mitosis influenced the LB content of type 2 cells after lung injury and suggest a previously unrecognized link between cell division and the surfactant system of the lung. 38 references, 5 figures, 2 tables.

  7. [Molecular pathogenesis of peripheral T cell lymphoma (2): extranodal NK/T cell lymphoma, nasal type, adult T cell leukemia/lymphoma and enteropathy associated T cell lymphoma].

    PubMed

    Couronné, Lucile; Bastard, Christian; Gaulard, Philippe; Hermine, Olivier; Bernard, Olivier

    2015-11-01

    Peripheral T-cell lymphomas (PTCL) belong to the group of non-Hodgkin lymphoma and particularly that of mature T /NK cells lymphoproliferative neoplasms. The 2008 WHO classification describes different PTCL entities with varying prevalence. With the exception of histologic subtype "ALK positive anaplastic large cell lymphoma", PTCL are characterized by a poor prognosis. The mechanisms underlying the pathogenesis of these lymphomas are not yet fully understood, but development of genomic high-throughput analysis techniques now allows to extensively identify the molecular abnormalities present in tumor cells. This review aims to summarize the current knowledge and recent advances about the molecular events occurring at the origin or during the natural history of main entities of PTCL. The first part published in the October issue was focused on the three more frequent entities, i.e. angioimmunoblastic T-cell lymphoma, peripheral T-cell lymphoma, not otherwise specified, and anaplastic large cell lymphoma. The second part presented herein will describe other subtypes less frequent and of poor prognosis : extranodal NK/T-cell lymphoma, nasal type, adult T-cell leukemia/lymphoma, and enteropathy-associated T-cell lymphoma. PMID:26576610

  8. Autoantigen-specific B-cell depletion overcomes failed immune tolerance in type 1 diabetes.

    PubMed

    Henry, Rachel A; Kendall, Peggy L; Thomas, James W

    2012-08-01

    Eliminating autoantigen-specific B cells is an attractive alternative to global B-cell depletion for autoimmune disease treatment. To identify the potential for targeting a key autoimmune B-cell specificity in type 1 diabetes, insulin-binding B cells were tracked within a polyclonal repertoire using heavy chain B-cell receptor (BCR) transgenic (VH125Tg) mice. Insulin-specific B cells are rare in the periphery of nonautoimmune VH125Tg/C57BL/6 mice and WT/NOD autoimmune mice, whereas they clearly populate 1% of mature B-cell subsets in VH125Tg/NOD mice. Autoantigen upregulates CD86 in anti-insulin B cells, suggesting they are competent to interact with T cells. Endogenous insulin occupies anti-insulin BCR beginning with antigen commitment in bone marrow parenchyma, as identified by a second anti-insulin monoclonal antibody. Administration of this monoclonal antibody selectively eliminates insulin-reactive B cells in vivo and prevents disease in WT/NOD mice. Unexpectedly, developing B cells are less amenable to depletion, despite increased BCR sensitivity. These findings exemplify how a critical type 1 diabetes B-cell specificity escapes immune tolerance checkpoints. Disease liability is corrected by eliminating this B-cell specificity, providing proof of concept for a novel therapeutic approach for autoimmune disease. PMID:22698916

  9. Cold plasma selectivity in the interaction with various types of the cells

    NASA Astrophysics Data System (ADS)

    Volotskova, Olga; Stepp, Mary Ann; Keidar, Michael

    2011-10-01

    Present research in the area of cold atmospheric plasma (CAP) demonstrates great potential in various areas including medicine and biology. Depending on their configuration they can be used for wound healing, sterilization, targeted cell/tissue removal, and cancer treatments. Here we explore potential mechanisms by which CAP alters cell migration and influences cell adhesion. The migration studies are focused on the CAP interaction with fibroblasts and corneal epithelial cells. Data show that various types of cells have different thresholds (treatment times) required to achieve maximum inhibition of cell migration which is around ~30-40%. Studies to assess the impact of CAP treatment on the activation state of integrins and focal adhesion size by immunofluorescence showed more active b1 integrin on the cell surface and large focal adhesions after CAP treatment. Based on these data, a thermodynamic model is presented to explain how CAP leads to integrin activation and focal adhesion assembly. Also responses of the various types of the cells to the cold plasma treatment on the example of the epithelial keratinocytes, papilloma and carcinoma cells are studied. Cell cycle, migration and cell vitality analysis were performed. The goal of this study is to understand the mechanism by which the CAP jet alters cell migration, influences adhesion and cell survival.

  10. C-type lectins do not act as functional receptors for filovirus entry into cells

    SciTech Connect

    Matsuno, Keita; Nakayama, Eri; Noyori, Osamu; Marzi, Andrea; Ebihara, Hideki; Irimura, Tatsuro; Feldmann, Heinz; Takada, Ayato

    2010-12-03

    Research highlights: {yields} Filovirus glycoprotein (GP) having a deficient receptor binding region were generated. {yields} Mutant GPs mediated virus entry less efficiently than wild-type GP. {yields} Mutant GPs bound to C-type lectins but not mediated entire steps of cellular entry. {yields} C-type lectins do not independently mediate filovirus entry into cells. {yields} Other molecule(s) are required for C-type lectin-mediated entry of filoviruses. -- Abstract: Cellular C-type lectins have been reported to facilitate filovirus infection by binding to glycans on filovirus glycoprotein (GP). However, it is not clearly known whether interaction between C-type lectins and GP mediates all the steps of virus entry (i.e., attachment, internalization, and membrane fusion). In this study, we generated vesicular stomatitis viruses pseudotyped with mutant GPs that have impaired structures of the putative receptor binding regions and thus reduced ability to infect the monkey kidney cells that are routinely used for virus propagation. We found that infectivities of viruses with the mutant GPs dropped in C-type lectin-expressing cells, parallel with those in the monkey kidney cells, whereas binding activities of these GPs to the C-type lectins were not correlated with the reduced infectivities. These results suggest that C-type lectin-mediated entry of filoviruses requires other cellular molecule(s) that may be involved in virion internalization or membrane fusion.

  11. MUCOSAL IMMUNOLOGY. The microbiota regulates type 2 immunity through RORγt⁺ T cells.

    PubMed

    Ohnmacht, Caspar; Park, Joo-Hong; Cording, Sascha; Wing, James B; Atarashi, Koji; Obata, Yuuki; Gaboriau-Routhiau, Valérie; Marques, Rute; Dulauroy, Sophie; Fedoseeva, Maria; Busslinger, Meinrad; Cerf-Bensussan, Nadine; Boneca, Ivo G; Voehringer, David; Hase, Koji; Honda, Kenya; Sakaguchi, Shimon; Eberl, Gérard

    2015-08-28

    Changes to the symbiotic microbiota early in life, or the absence of it, can lead to exacerbated type 2 immunity and allergic inflammations. Although it is unclear how the microbiota regulates type 2 immunity, it is a strong inducer of proinflammatory T helper 17 (T(H)17) cells and regulatory T cells (T(regs)) in the intestine. Here, we report that microbiota-induced T(regs) express the nuclear hormone receptor RORγt and differentiate along a pathway that also leads to T(H)17 cells. In the absence of RORγt(+) T(regs), T(H)2-driven defense against helminths is more efficient, whereas T(H)2-associated pathology is exacerbated. Thus, the microbiota regulates type 2 responses through the induction of type 3 RORγt(+) T(regs) and T(H)17 cells and acts as a key factor in balancing immune responses at mucosal surfaces. PMID:26160380

  12. A Mammalian enhancer trap resource for discovering and manipulating neuronal cell types

    PubMed Central

    Shima, Yasuyuki; Sugino, Ken; Hempel, Chris Martin; Shima, Masami; Taneja, Praveen; Bullis, James B; Mehta, Sonam; Lois, Carlos; Nelson, Sacha B

    2016-01-01

    There is a continuing need for driver strains to enable cell-type-specific manipulation in the nervous system. Each cell type expresses a unique set of genes, and recapitulating expression of marker genes by BAC transgenesis or knock-in has generated useful transgenic mouse lines. However, since genes are often expressed in many cell types, many of these lines have relatively broad expression patterns. We report an alternative transgenic approach capturing distal enhancers for more focused expression. We identified an enhancer trap probe often producing restricted reporter expression and developed efficient enhancer trap screening with the PiggyBac transposon. We established more than 200 lines and found many lines that label small subsets of neurons in brain substructures, including known and novel cell types. Images and other information about each line are available online (enhancertrap.bio.brandeis.edu). DOI: http://dx.doi.org/10.7554/eLife.13503.001 PMID:26999799

  13. Type I interferons regulate eomesodermin expression and the development of unconventional memory CD8(+) T cells.

    PubMed

    Martinet, Valérie; Tonon, Sandrine; Torres, David; Azouz, Abdulkader; Nguyen, Muriel; Kohler, Arnaud; Flamand, Véronique; Mao, Chai-An; Klein, William H; Leo, Oberdan; Goriely, Stanislas

    2015-05-08

    CD8(+) T-cell memory phenotype and function are acquired after antigen-driven activation. Memory-like cells may also arise in absence of antigenic exposure in the thymus or in the periphery. Eomesodermin (Eomes) is a key transcription factor for the development of these unconventional memory cells. Herein, we show that type I interferon signalling in CD8(+) T cells directly activates Eomes gene expression. Consistent with this observation, the phenotype, function and age-dependent expansion of 'virtual memory' CD8(+) T cells are strongly affected in absence of type I interferon signalling. In addition, type I interferons induce a sustained expansion of 'virtual memory' CD8(+) T cells in an Eomes-dependent fashion. We further show that the development of 'innate thymic' CD8(+) T cells is dependent on the same pathway. In conclusion, we demonstrate that type I interferon signalling in CD8(+) T cells drives Eomes expression and thereby regulates the function and homeostasis of memory-like CD8(+) T cells.

  14. Stem cell-based therapies and immunomodulatory approaches in newly diagnosed type 1 diabetes.

    PubMed

    Couri, Carlos Eduardo Barra; Voltarelli, Júlio César

    2011-03-01

    Type 1 diabetes mellitus is an autoimmune disease against pancreatic β cells. The autoimmune response begins months or years before the clinical presentation. At the time of hyperglycemic symptoms a small amount of β cell mass still remains. The main therapeutic option to type 1 diabetes mellitus is daily insulin injections which is shown to promote tighter glucose control and to reduce much of diabetic chronic complications. Subgroup analysis of the Diabetes Control and Complication Trial (DCCT) showed another important aspect related to long term complications of diabetes, ie, patients with initially larger residual β cell mass suffered less microvascular complications and less hypoglycemic events than those patients with small amounts of β cells at diagnosis. In face of this, β cell preservation has become another important target in the management of type 1 diabetes and its related complications. In this review, we summarize various immunomodulatory regimens ever used in humans, including stem cell-based strategies, aiming at blocking autoimmunity against pancreatic β cells and at promoting β cell preservation and/or possible β cell regeneration in recent-onset type 1 diabetes.

  15. Complex heterogeneous tissue constructs containing multiple cell types prepared by inkjet printing technology.

    PubMed

    Xu, Tao; Zhao, Weixin; Zhu, Jian-Ming; Albanna, Mohammad Z; Yoo, James J; Atala, Anthony

    2013-01-01

    This study was designed to develop a versatile method for fabricating complex and heterogeneous three-dimensional (3D) tissue constructs using simultaneous ink-jetting of multiple cell types. Human amniotic fluid-derived stem cells (hAFSCs), canine smooth muscle cells (dSMCs), and bovine aortic endothelial cells (bECs), were separately mixed with ionic cross-linker calcium chloride (CaCl(2)), loaded into separate ink cartridges and printed using a modified thermal inkjet printer. The three cell types were delivered layer-by-layer to pre-determined locations in a sodium alginate-collagen composite located in a chamber under the printer. The reaction between CaCl(2) and sodium alginate resulted in a rapid formation of a solid composite gel and the printed cells were anchored in designated areas within the gel. The printing process was repeated for several cycles leading to a complex 3D multi-cell hybrid construct. The biological functions of the 3D printed constructs were evaluated in vitro and in vivo. Each of the printed cell types maintained their viability and normal proliferation rates, phenotypic expression, and physiological functions within the heterogeneous constructs. The bioprinted constructs were able to survive and mature into functional tissues with adequate vascularization in vivo. These findings demonstrate the feasibility of fabricating complex heterogeneous tissue constructs containing multiple cell types using inkjet printing technology.

  16. Cell-type-specific enrichment of risk-associated regulatory elements at ovarian cancer susceptibility loci

    PubMed Central

    Coetzee, Simon G.; Shen, Howard C.; Hazelett, Dennis J.; Lawrenson, Kate; Kuchenbaecker, Karoline; Tyrer, Jonathan; Rhie, Suhn K.; Levanon, Keren; Karst, Alison; Drapkin, Ronny; Ramus, Susan J.; Couch, Fergus J.; Offit, Kenneth; Chenevix-Trench, Georgia; Monteiro, Alvaro N.A.; Antoniou, Antonis; Freedman, Matthew; Coetzee, Gerhard A.; Pharoah, Paul D.P.; Noushmehr, Houtan; Gayther, Simon A.

    2015-01-01

    Understanding the regulatory landscape of the human genome is a central question in complex trait genetics. Most single-nucleotide polymorphisms (SNPs) associated with cancer risk lie in non-protein-coding regions, implicating regulatory DNA elements as functional targets of susceptibility variants. Here, we describe genome-wide annotation of regions of open chromatin and histone modification in fallopian tube and ovarian surface epithelial cells (FTSECs, OSECs), the debated cellular origins of high-grade serous ovarian cancers (HGSOCs) and in endometriosis epithelial cells (EECs), the likely precursor of clear cell ovarian carcinomas (CCOCs). The regulatory architecture of these cell types was compared with normal human mammary epithelial cells and LNCaP prostate cancer cells. We observed similar positional patterns of global enhancer signatures across the three different ovarian cancer precursor cell types, and evidence of tissue-specific regulatory signatures compared to non-gynecological cell types. We found significant enrichment for risk-associated SNPs intersecting regulatory biofeatures at 17 known HGSOC susceptibility loci in FTSECs (P = 3.8 × 10−30), OSECs (P = 2.4 × 10−23) and HMECs (P = 6.7 × 10−15) but not for EECs (P = 0.45) or LNCaP cells (P = 0.88). Hierarchical clustering of risk SNPs conditioned on the six different cell types indicates FTSECs and OSECs are highly related (96% of samples using multi-scale bootstrapping) suggesting both cell types may be precursors of HGSOC. These data represent the first description of regulatory catalogues of normal precursor cells for different ovarian cancer subtypes, and provide unique insights into the tissue specific regulatory variation with respect to the likely functional targets of germline genetic susceptibility variants for ovarian cancer. PMID:25804953

  17. [Membrane type 1 matrix metalloproteinase (MT1-MMP) and the regulators of its activity as invasive factors in squamous cell cervical carcinomas].

    PubMed

    Timoshenko, O S; Gureeva, T A; Kugaevskaia, E V; Solov'eva, N I

    2014-01-01

    Membrane type 1 matrix metalloproteinase (MT1MMP) is one of matrix metalloproteinases (MMP), which play а key role in tumor invasion and metastasis. The aim of this study was to elucidate the peculiarities of expression of MT1MMP and endogenous regulators of its activity: the activator - furin and the inhibitor - TIMP-2, as invasive factors of squamous cell cervical carcinomas (SCC). The study was carried out using 11 specimens of SCC and 11 specimens of morphologically normal tissue adjacent to the tumor. It was shown that the increase of MT1-MMP and furin expression and low of TIMP-2 expression makes the main contribution to the destructive (invasive) potential of SCC. Moreover, substantial expression of MT1-MMP was registered in the specimens of morphologically normal adjoining to tumor tissue. This expression was found to make an additional contribution to the destructive potential of the cervical tumor.

  18. Studies on quantitative analysis and automatic recognition of cell types of lung cancer.

    PubMed

    Chen, Yi-Chen; Hu, Kuang-Hu; Li, Fang-Zhen; Li, Shu-Yu; Su, Wan-Fang; Huang, Zhi-Ying; Hu, Ying-Xiong

    2006-01-01

    Recognition of lung cancer cells is very important to the clinical diagnosis of lung cancer. In this paper we present a novel method to extract the structure characteristics of lung cancer cells and automatically recognize their types. Firstly soft mathematical morphology methods are used to enhance the grayscale image, to improve the definition of images, and to eliminate most of disturbance, noise and information of subordinate images, so the contour of target lung cancer cell and biological shape characteristic parameters can be extracted accurately. Then the minimum distance classifier is introduced to realize the automatic recognition of different types of lung cancer cells. A software system named "CANCER.LUNG" is established to demonstrate the efficiency of this method. The clinical experiments show that this method can accurately and objectively recognize the type of lung cancer cells, which can significantly improve the pathology research on the pathological changes of lung cancer and clinical assistant diagnoses.

  19. Cell-type specific regulation of gene expression by simian virus 40 T antigens

    SciTech Connect

    Cantalupo, Paul G.; Saenz-Robles, Maria Teresa; Rathi, Abhilasha V.; Beerman, Rebecca W.; Patterson, William H.; Whitehead, Robert H.; Pipas, James M.

    2009-03-30

    SV40 transforms cells through the action of two oncoproteins, large T antigen and small t antigen. Small t antigen targets phosphatase PP2A, while large T antigen stimulates cell proliferation and survival by action on multiple proteins, including the tumor suppressors Rb and p53. Large T antigen also binds components of the transcription initiation complex and several transcription factors. We examined global gene expression in SV40-transformed mouse embryo fibroblasts, and in enterocytes obtained from transgenic mice. SV40 transformation alters the expression of approximately 800 cellular genes in both systems. Much of this regulation is observed in both MEFs and enterocytes and is consistent with T antigen action on the Rb-E2F pathway. However, the regulation of many genes is cell-type specific, suggesting that unique signaling pathways are activated in different cell types upon transformation, and that the consequences of SV40 transformation depends on the type of cell targeted.

  20. GGPPS-mediated Rab27A geranylgeranylation regulates β cell dysfunction during type 2 diabetes development by affecting insulin granule docked pool formation.

    PubMed

    Jiang, Shan; Shen, Di; Jia, Wen-Jun; Han, Xiao; Shen, Ning; Tao, Weiwei; Gao, Xiang; Xue, Bin; Li, Chao-Jun

    2016-01-01

    Loss of first-phase insulin secretion associated with β cell dysfunction is an independent predictor of type 2 diabetes mellitus (T2DM) onset. Here we found that a critical enzyme involved in protein prenylation, geranylgeranyl pyrophosphate synthase (GGPPS), is required to maintain first-phase insulin secretion. GGPPS shows a biphasic expression pattern in islets of db/db mice during the progression of T2DM: GGPPS is increased during the insulin compensatory period, followed by a decrease during β cell dysfunction. Ggpps deletion in β cells results in typical T2DM β cell dysfunction, with blunted glucose-stimulated insulin secretion and consequent insulin secretion insufficiency. However, the number and size of islets and insulin biosynthesis are unaltered. Transmission electron microscopy shows a reduced number of insulin granules adjacent to the cellular membrane, suggesting a defect in docked granule pool formation, while the reserve pool is unaffected. Ggpps ablation depletes GGPP and impairs Rab27A geranylgeranylation, which is responsible for the docked pool deficiency in Ggpps-null mice. Moreover, GGPPS re-expression or GGPP administration restore glucose-stimulated insulin secretion in Ggpps-null islets. These results suggest that GGPPS-controlled protein geranylgeranylation, which regulates formation of the insulin granule docked pool, is critical for β cell function and insulin release during the development of T2DM.

  1. SEQUENTIAL CELLULAR CHANGES PRODUCED BY TYPES 5 AND 7 ADENOVIRUSES IN HELA CELLS AND IN HUMAN AMNIOTIC CELLS

    PubMed Central

    Boyer, Georgiana S.; Denny, Floyd W.; Ginsberg, Harold S.

    1959-01-01

    The sequential cytological changes which develop in tissue culture cells infected with adenovirus types 5 and 7 are described and compared with those produced by adenovirus types 1, 2, 3, and 4. The evidence that is presented indicates that types 1, 2, and 5 belong to one major subdivision of the adenovirus group and types 3, 4, and 7 to another. That the host cell nucleus is the principal site of adenovirus synthesis has been confirmed by fluorescent antibody studies. They have demonstrated the occurrence of type-specific adenovirus antigen in the characteristic intranuclear inclusions and other virus-induced structures reported to contain virus-like particles or shown by electronmicroscopy. PMID:13803575

  2. Molecular beacon-enabled purification of living cells by targeting cell type-specific mRNAs.

    PubMed

    Wile, Brian M; Ban, Kiwon; Yoon, Young-Sup; Bao, Gang

    2014-10-01

    Molecular beacons (MBs) are dual-labeled oligonucleotides that fluoresce only in the presence of complementary mRNA. The use of MBs to target specific mRNAs allows sorting of specific cells from a mixed cell population. In contrast to existing approaches that are limited by available surface markers or selectable metabolic characteristics, the MB-based method enables the isolation of a wide variety of cells. For example, the ability to purify specific cell types derived from pluripotent stem cells (PSCs) is important for basic research and therapeutics. In addition to providing a general protocol for MB design, validation and nucleofection into cells, we describe how to isolate a specific cell population from differentiating PSCs. By using this protocol, we have successfully isolated cardiomyocytes differentiated from mouse or human PSCs (hPSCs) with ∼ 97% purity, as confirmed by electrophysiology and immunocytochemistry. After designing MBs, their ordering and validation requires 2 weeks, and the isolation process requires 3 h.

  3. Hypoxia-Induced Modulation of Apoptosis and BCL-2 Family Proteins in Different Cancer Cell Types

    PubMed Central

    Sermeus, Audrey; Genin, Marie; Maincent, Amélie; Fransolet, Maude; Notte, Annick; Leclere, Lionel; Riquier, Hélène; Arnould, Thierry; Michiels, Carine

    2012-01-01

    Hypoxia plays an important role in the resistance of tumour cells to chemotherapy. However, the exact mechanisms underlying this process are not well understood. Moreover, according to the cell lines, hypoxia differently influences cell death. The study of the effects of hypoxia on the apoptosis induced by 5 chemotherapeutic drugs in 7 cancer cell types showed that hypoxia generally inhibited the drug-induced apoptosis. In most cases, the effect of hypoxia was the same for all the drugs in one cell type. The expression profile of 93 genes involved in apoptosis as well as the protein level of BCL-2 family proteins were then investigated. In HepG2 cells that are strongly protected against cell death by hypoxia, hypoxia decreased the abundance of nearly all the pro-apoptotic BCL-2 family proteins while none of them are decreased in A549 cells that are not protected against cell death by hypoxia. In HepG2 cells, hypoxia decreased NOXA and BAD abundance and modified the electrophoretic mobility of BIMEL. BIM and NOXA are important mediators of etoposide-induced cell death in HepG2 cells and the hypoxia-induced modification of these proteins abundance or post-translational modifications partly account for chemoresistance. Finally, the modulation of the abundance and/or of the post-translational modifications of most proteins of the BCL-2 family by hypoxia involves p53-dependent and –independent pathways and is cell type-dependent. A better understanding of these cell-to-cell variations is crucial in order to overcome hypoxia-induced resistance and to ameliorate cancer therapy. PMID:23139748

  4. Role of cell type and animal species in tumor metastasis

    NASA Astrophysics Data System (ADS)

    Solban, Nicolas; Georgakoudi, Irene; Rice, William L.; Lin, Charles; Hasan, Tayyaba

    2004-06-01

    Photodynamic therapy (PDT) is now a reasonably well-known therapeutic option and is approved as a first line treatment of age-related macular degeneration (AMD), a non-oncologic condition. For most cancer applications PDT is approved mainly as a palliative or adjunctive treatment often when all other options have failed. As the modality evolves toward becoming a first-line or curative option, long-term effects of processes involved will need to be studied. Cellular and tissue responses to PDT are more complex than responses to the more conventional therapies, perhaps because PDT is inherently a binary (or ternary) therapy. In addition to the nature and localization of the photosensitizer (PS), the timing of illumination after administration, the mode of administration and the PS and light doses, the efficacy and selectivity of responses are also determined by the physiology and geometry of tumors, the inherent survivability of tumor cells (in circulation and other anatomic sites) and cellular and molecular responses to PDT. The overall outcome of photodynamic treatment in the long term is determined by a combination, in varying degrees, of all of the above factors. In order to enhance and broaden the application of PDT to complex anatomical sites, an understanding of these factors would be useful. In the laboratory, the outcome is also dependent on the specific animal models being studied. This manuscript discusses preliminary studies along these lines using a variety of tools and implications, if any, of the results obtained.

  5. Regenerative Effects of Mesenchymal Stem Cells: Contribution of Muse Cells, a Novel Pluripotent Stem Cell Type that Resides in Mesenchymal Cells.

    PubMed

    Wakao, Shohei; Kuroda, Yasumasa; Ogura, Fumitaka; Shigemoto, Taeko; Dezawa, Mari

    2012-11-08

    Mesenchymal stem cells (MSCs) are easily accessible and safe for regenerative medicine. MSCs exert trophic, immunomodulatory, anti-apoptotic, and tissue regeneration effects in a variety of tissues and organs, but their entity remains an enigma. Because MSCs are generally harvested from mesenchymal tissues, such as bone marrow, adipose tissue, or umbilical cord as adherent cells, MSCs comprise crude cell populations and are heterogeneous. The specific cells responsible for each effect have not been clarified. The most interesting property of MSCs is that, despite being adult stem cells that belong to the mesenchymal tissue lineage, they are able to differentiate into a broad spectrum of cells beyond the boundary of mesodermal lineage cells into ectodermal or endodermal lineages, and repair tissues. The broad spectrum of differentiation ability and tissue-repairing effects of MSCs might be mediated in part by the presence of a novel pluripotent stem cell type recently found in adult human mesenchymal tissues, termed multilineage-differentiating stress enduring (Muse) cells. Here we review recently updated studies of the regenerative effects of MSCs and discuss their potential in regenerative medicine.

  6. Redirecting cell-type specific cytokine responses with engineered interleukin-4 superkines

    PubMed Central

    Junttila, Ilkka S.; Creusot, Remi J.; Moraga, Ignacio; Bates, Darren L.; Wong, Michael T.; Alonso, Michael N.; Suhoski, Megan M.; Lupardus, Patrick; Meier-Schellersheim, Martin; Engleman, Edgar G.; Utz, Paul J.; Fathman, C. Garrison; Paul, William E.; Garcia, K. Christopher

    2012-01-01

    Cytokines dimerize their receptors, with binding of the “second chain” triggering signaling. In the interleukin (IL)-4/13 system, different cell types express varying levels of alternative second receptor chains (γc or IL-13Rα1), forming functionally distinct Type-I or Type-II complexes. We manipulated the affinity and specificity of second chain recruitment by human IL-4. A Type-I receptor-selective IL-4 ‘superkine’ with 3700-fold higher affinity for γc was 3-10 fold more potent than wild-type IL-4. Conversely, a variant with high affinity for IL-13Rα1 more potently activated cells expressing the Type-II receptor, and induced differentiation of dendritic cells from monocytes, implicating the Type-II receptor in this process. Superkines exhibited signaling advantages on cells with lower second chain levels. Comparative transcriptional analysis reveals that the superkines induce largely redundant gene expression profiles. Variable second chain levels can be exploited to redirect cytokines towards distinct cell subsets and elicit novel actions, potentially improving the selectivity of cytokine therapy. PMID:23103943

  7. Nivolumab, an Anti-Programmed Cell Death-1 Antibody, Induces Fulminant Type 1 Diabetes.

    PubMed

    Miyoshi, Yuka; Ogawa, Osamu; Oyama, Yu

    2016-01-01

    Programmed cell death-1 (PD-1), an immunoreceptor, is located on T cells and pro-B cells and interacts with its ligands to inhibit T cell activation and proliferation, thereby promoting immunological self-tolerance. Nivolumab, an anti-PD1 antibody, blocks PD-1 and can restore anticancer immune responses by abrogating PD-1 pathway-mediated T-cell inhibition. Autoimmune adverse events are expected with PD-1 therapy. Fulminant type 1 diabetes is the subtype of type 1 diabetes. The clinical feature is the extremely rapid progression of hyperglycemia and ketoacidosis. Here we describe a 66-year-old woman with advanced melanoma who was treated with nivolumab. After 4 months and six doses of the medicine, the patient was admitted to the hospital with complaints of nausea and vomiting. The laboratory data showed ketonuria, hyperglycemia (531 mg/dl), high anion gap metabolic acidosis, HbA1c (7.3%), and absence of insulin-secreting capacity. These data are compatible with the criteria of fulminant type 1 diabetes. The patient was diagnosed with diabetic ketoacidosis because of fulminant type 1 diabetes. The findings of this case indicated that nivolumab can cause fulminant type 1 diabetes. Diabetic ketoacidosis due to fulminant type 1 diabetes is potentially fatal condition. Thus, diabetic ketoacidosis due to fulminant type 1 diabetes should be considered in the differential diagnosis when patients treated with nivolumab complain of gastrointestinal symptoms. PMID:27297738

  8. Discovering cell types in flow cytometry data with random matrix theory

    NASA Astrophysics Data System (ADS)

    Shen, Yang; Nussenblatt, Robert; Losert, Wolfgang

    Flow cytometry is a widely used experimental technique in immunology research. During the experiments, peripheral blood mononuclear cells (PBMC) from a single patient, labeled with multiple fluorescent stains that bind to different proteins, are illuminated by a laser. The intensity of each stain on a single cell is recorded and reflects the amount of protein expressed by that cell. The data analysis focuses on identifying specific cell types related to a disease. Different cell types can be identified by the type and amount of protein they express. To date, this has most often been done manually by labelling a protein as expressed or not while ignoring the amount of expression. Using a cross correlation matrix of stain intensities, which contains both information on the proteins expressed and their amount, has been largely ignored by researchers as it suffers from measurement noise. Here we present an algorithm to identify cell types in flow cytometry data which uses random matrix theory (RMT) to reduce noise in a cross correlation matrix. We demonstrate our method using a published flow cytometry data set. Compared with previous analysis techniques, we were able to rediscover relevant cell types in an automatic way. Department of Physics, University of Maryland, College Park, MD 20742.

  9. Interleukin-9 and T helper type 9 cells in rheumatic diseases.

    PubMed

    Ciccia, F; Guggino, G; Ferrante, A; Cipriani, P; Giacomelli, R; Triolo, G

    2016-08-01

    Interleukin (IL)-9 is a 28-30 kDa monomeric glycosylated polypeptide belonging to the IL-7/IL-9 family of proteins that bind to a composite receptor consisting of the private receptor IL-9R and the IL-2 receptor, gamma (IL-2RG), a common gamma subunit shared by the receptors of many different cytokines. The IL-9R is expressed widely and IL-9 impacts a number of effector cells, such as effector T cells, B cells, innate lymphoid cells, mast cells, polymorphonuclear cells, epithelial cells and smooth muscle cells, playing an important role in regulating inflammatory immunity. The critical role of IL-9 in promoting cellular and humoral immune responses makes it an important focus of potential therapeutic interventions. Recently, a defined subset of T helper type cells, Th9 cells, has been identified by the potent production of IL-9. The involvement of the Th9 cell subset has been described in many types of inflammatory diseases, namely atopic diseases, helminth infections, experimental autoimmune encephalomyelitis and ulcerative colitis. In this review, we summarize the IL-9 biological activities, highlighting roles for IL-9 and Th9 cells in rheumatoid and psoriatic arthritis, systemic vasculitis, systemic lupus erythematosus and systemic sclerosis. PMID:27159882

  10. A versatile Multisite Gateway-compatible promoter and transgenic line collection for cell type-specific functional genomics in Arabidopsis.

    PubMed

    Marquès-Bueno, Maria Mar; Morao, Ana K; Cayrel, Anne; Platre, Matthieu P; Barberon, Marie; Caillieux, Erwann; Colot, Vincent; Jaillais, Yvon; Roudier, François; Vert, Grégory

    2016-01-01

    Multicellular organisms are composed of many cell types that acquire their specific fate through a precisely controlled pattern of gene expression in time and space dictated in part by cell type-specific promoter activity. Understanding the contribution of highly specialized cell types in the development of a whole organism requires the ability to isolate or analyze different cell types separately. We have characterized and validated a large collection of root cell type-specific promoters and have generated cell type-specific marker lines. These benchmarked promoters can be readily used to evaluate cell type-specific complementation of mutant phenotypes, or to knockdown gene expression using targeted expression of artificial miRNA. We also generated vectors and characterized transgenic lines for cell type-specific induction of gene expression and cell type-specific isolation of nuclei for RNA and chromatin profiling. Vectors and seeds from transgenic Arabidopsis plants will be freely available, and will promote rapid progress in cell type-specific functional genomics. We demonstrate the power of this promoter set for analysis of complex biological processes by investigating the contribution of root cell types in the IRT1-dependent root iron uptake. Our findings revealed the complex spatial expression pattern of IRT1 in both root epidermis and phloem companion cells and the requirement for IRT1 to be expressed in both cell types for proper iron homeostasis.

  11. A versatile Multisite Gateway-compatible promoter and transgenic line collection for cell type-specific functional genomics in Arabidopsis

    PubMed Central

    Platre, Matthieu Pierre; Barberon, Marie; Caillieux, Erwann; Colot, Vincent

    2016-01-01

    Summary Multicellular organisms are composed of many cell types that acquire their specific fate through a precisely controlled pattern of gene expression in time and space dictated in part by cell type-specific promoter activity. Understanding the contribution of highly specialized cell types in the development of a whole organism requires the ability to isolate or analyze different cell types separately. We have characterized and validated a large collection of root cell type-specific promoters and have generated cell type-specific marker lines. These benchmarked promoters can be readily used to evaluate cell type-specific complementation of mutant phenotypes, or to knockdown gene expression using targeted expression of artificial miRNA. We also generated vectors and characterized transgenic lines for cell type-specific induction of gene expression and cell type-specific isolation of nuclei for RNA and chromatin profiling. Vectors and seeds from transgenic Arabidopsis plants will be freely available, and will promote rapid progress in cell type-specific functional genomics. We demonstrate the power of this promoter set for analysis of complex biological processes by investigating the contribution of root cell types in the IRT1-dependent root iron uptake. Our findings revealed the complex spatial expression pattern of IRT1 in both root epidermis and phloem companion cells and the requirement for IRT1 to be expressed in both cell types for proper iron homeostasis. PMID:26662936

  12. N-Glycoprotein Surfaceomes of Four Developmentally Distinct Mouse Cell Types

    PubMed Central

    Kropp, Erin M.; Bhattacharya, Subarna; Waas, Matthew; Chuppa, Sandra L.; Hadjantonakis, Anna-Katerina; Boheler, Kenneth R.; Gundry, Rebekah L.

    2014-01-01

    Purpose Detailed knowledge of cell surface proteins present during early embryonic development remains limited for most cell lineages. Due to the relevance of cell surface proteins in their functional roles controlling cell signaling and their utility as accessible, non-genetic markers for cell identification and sorting, the goal of this study was to provide new information regarding the cell surface proteins present during early mouse embryonic development. Experimental Design Using the Cell Surface Capture Technology, the cell surface N-glycoproteomes of three cell lines and one in vitro differentiated cell type representing distinct cell fates and stages in mouse embryogenesis were assessed. Results Altogether, more than 600 cell surface N-glycoproteins were identified represented by >5500 N-glycopeptides. Conclusions and Clinical Relevance The development of new, informative cell surface markers for the reliable identification and isolation of functionally defined subsets of cells from early developmental stages will advance the use of stem cell technologies for mechanistic developmental studies, including disease modeling and drug discovery. PMID:24920426

  13. Human oocytes reprogram adult somatic nuclei of a type 1 diabetic to diploid pluripotent stem cells.

    PubMed

    Yamada, Mitsutoshi; Johannesson, Bjarki; Sagi, Ido; Burnett, Lisa Cole; Kort, Daniel H; Prosser, Robert W; Paull, Daniel; Nestor, Michael W; Freeby, Matthew; Greenberg, Ellen; Goland, Robin S; Leibel, Rudolph L; Solomon, Susan L; Benvenisty, Nissim; Sauer, Mark V; Egli, Dieter

    2014-06-26

    The transfer of somatic cell nuclei into oocytes can give rise to pluripotent stem cells that are consistently equivalent to embryonic stem cells, holding promise for autologous cell replacement therapy. Although methods to induce pluripotent stem cells from somatic cells by transcription factors are widely used in basic research, numerous differences between induced pluripotent stem cells and embryonic stem cells have been reported, potentially affecting their clinical use. Because of the therapeutic potential of diploid embryonic stem-cell lines derived from adult cells of diseased human subjects, we have systematically investigated the parameters affecting efficiency of blastocyst development and stem-cell derivation. Here we show that improvements to the oocyte activation protocol, including the use of both kinase and translation inhibitors, and cell culture in the presence of histone deacetylase inhibitors, promote development to the blastocyst stage. Developmental efficiency varied between oocyte donors, and was inversely related to the number of days of hormonal stimulation required for oocyte maturation, whereas the daily dose of gonadotropin or the total number of metaphase II oocytes retrieved did not affect developmental outcome. Because the use of concentrated Sendai virus for cell fusion induced an increase in intracellular calcium concentration, causing premature oocyte activation, we used diluted Sendai virus in calcium-free medium. Using this modified nuclear transfer protocol, we derived diploid pluripotent stem-cell lines from somatic cells of a newborn and, for the first time, an adult, a female with type 1 diabetes.

  14. Breaking the In Vitro Alveolar Type II Cell Proliferation Barrier while Retaining Ion Transport Properties

    PubMed Central

    Dang, Hong; Cheluvaraju, Chaitra; Jones, Lisa C.; Liu, Xuefeng; O’Neal, Wanda K.; Randell, Scott H.; Schlegel, Richard; Boucher, Richard C.

    2014-01-01

    Alveolar type (AT)I and ATII cells are central to maintaining normal alveolar fluid homeostasis. When disrupted, they contribute to the pathogenesis of acute lung injury (ALI) and acute respiratory distress syndrome. Research on ATII cells has been limited by the inability to propagate primary cells in vitro to study their specific functional properties. Moreover, primary ATII cells in vitro quickly transdifferentiate into nonproliferative “ATI-like” cells under traditional culture conditions. Recent studies have demonstrated that normal and tumor cells grown in culture with a combination of fibroblast (feeder cells) and a pharmacological Rho kinase inhibitor (Y-27632) exhibit indefinite cell proliferation that resembled a “conditionally reprogrammed cell” phenotype. Using this coculture system, we found that primary human ATII cells (1) proliferated at an exponential rate, (2) established epithelial colonies expressing ATII-specific and “ATI-like” mRNA and proteins after serial passage, (3) up-regulated genes important in cell proliferation and migration, and (4) on removal of feeder cells and Rho kinase inhibitor under air–liquid interface conditions, exhibited bioelectric and volume transport characteristics similar to freshly cultured ATII cells. Collectively, our results demonstrate that this novel coculture technique breaks the in vitro ATII cell proliferation barrier while retaining cell-specific functional properties. This work will allow for a significant increase in studies designed to elucidate ATII cell function with the goal of accelerating the development of novel therapies for alveolar diseases. PMID:24191670

  15. The Cellular and Molecular Mechanisms of Immuno-Suppression by Human Type 1 Regulatory T Cells

    PubMed Central

    Gregori, Silvia; Goudy, Kevin S.; Roncarolo, Maria Grazia

    2011-01-01

    The immuno-regulatory mechanisms of IL-10-producing type 1 regulatory T (Tr1) cells have been widely studied over the years. However, several recent discoveries have shed new light on the cellular and molecular mechanisms that human Tr1 cells use to control immune responses and induce tolerance. In this review we outline the well known and newly discovered regulatory properties of human Tr1 cells and provide an in-depth comparison of the known suppressor mechanisms of Tr1 cells with FOXP3+ Treg. We also highlight the role that Tr1 cells play in promoting and maintaining tolerance in autoimmunity, allergy, and transplantation. PMID:22566914

  16. Two types of ON direction-selective ganglion cells in rabbit retina.

    PubMed

    Kanjhan, Refik; Sivyer, Benjamin

    2010-10-11

    Direction-selective ganglion cells (DSGCs) respond with robust spiking to image motion in a particular direction. Previously, two main types of DSGCs have been described in rabbit retina: the ON-OFF DSGCs respond to both increases and decreases in illumination, whereas the ON DSGCs respond only to increases in illumination. In this study, we show that there are two distinct types of ON DSGCs, which can be separated by differences in their receptive-field properties, dendritic morphology and tracer-coupling pattern. While both types show robust direction-selectivity, one type responds to increases in illumination with sustained firing, whereas the other responds with relatively transient firing. The two types of ON DSGCs also have distinct dendritic morphologies: the sustained cells give rise to shorter and more numerous terminal dendrites, which are distributed throughout the dendritic field forming a space-filling lattice. In addition, the transient ON DSGCs, but not the sustained ON DSGCs, show tracer-coupling to a mosaic of amacrine cells when filled with Neurobiotin. Both types of ON DSGCs have been encountered in previous studies but were not recognized as distinct types. We propose that the two types also differ in their central projections, with only the sustained cells projecting to the medial terminal nucleus (MTN) of the accessory optic system (AOS).

  17. Comparing Corn Types for Differences in Cell Wall Characteristics and p-Coumaroylation of Lignin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study was undertaken to compare cell wall characteristics, including levels of pCA and lignin in diverse corn (Zea mays L.) germplasm. Five different types of corn (Zea mays L.) germplasm (four commercial and Teosinte) were grown in the greenhouse in individual pots. For each corn type, replica...

  18. Cardiorenal Syndrome Type 5: In Vitro Cytotoxicity Effects on Renal Tubular Cells and Inflammatory Profile

    PubMed Central

    Brocca, Alessandra; Virzì, Grazia Maria; Pasqualin, Chiara; Pastori, Silvia; Marcante, Stefano; de Cal, Massimo; Ronco, Claudio

    2015-01-01

    Background. Cardiorenal Syndrome Type 5 (CRS Type 5) reflects concomitant cardiac and renal dysfunctions in the setting of a wide spectrum of systemic disorders. Our aim was to study in vitro effects of CRS Type 5 plasma on renal tubular cells (RTCs), in terms of cellular death and the characterization of inflammatory plasma profile in these patients. Material and Methods. We enrolled 11 CRS Type 5 patients from ICU and 16 healthy controls. Plasma from patients and controls was incubated with renal tubular cells (RTCs) and cell death was evaluated. Plasma cytokines were detected. Results. RTCs incubated with CRS Type 5 plasma showed significantly higher apoptosis and necrosis with respect to controls. Plasma cytokine profile of CRS Type 5 patients was significantly different from controls: we observed the production of pro- and anti-inflammatory mediators in these patients. Caspase-3, caspase-8, and caspase-9 were activated in cells treated with CRS Type 5 plasma compared to controls. Conclusions. Our results underline the cytotoxic effect of CRS Type 5 mediators on RTC viability, probably due to the activation of both intrinsic and extrinsic pathways of apoptosis and to the deregulation of cytokine release. The consequence may be the damage of distant organs which lead to the worsening of condition of patients. PMID:26266085

  19. Infection of human synovial cells by human T cell lymphotropic virus type I. Proliferation and granulocyte/macrophage colony-stimulating factor production by synovial cells.

    PubMed Central

    Sakai, M; Eguchi, K; Terada, K; Nakashima, M; Yamashita, I; Ida, H; Kawabe, Y; Aoyagi, T; Takino, H; Nakamura, T

    1993-01-01

    The present study was performed to clarify the relationship between human T cell lymphotropic virus type I (HTLV-I) infection and chronic inflammatory arthropathy. To determine the ability of HTLV-I to infect synovial cells and the effect on synovial cell proliferation, synovial cells were cocultured with the HTLV-I-producing T cell lines (MT-2 or HCT-1). After coculture with HTLV-I-infected T cells, the synovial cells expressed HTLV-I-specific core antigens, and HTLV-I proviral DNA was detected from the synovial cells by polymerase chain reaction. These cocultured synovial cells with HTLV-I-infected T cells proliferated more actively than the synovial cells cocultured with uninfected T cells. This stimulatory effect of HTLV-I-infected T cells on synovial cell proliferation seems necessary to contact each other. After being cocultured with MT-2 cells, synovial cells proliferated more actively than control cells even after several passages. Furthermore, HTLV-I-infected synovial cells produced significant amounts of granulocyte/macrophage colony-stimulating factor. These results suggest that HTLV-I can infect synovial cells, resulting their active proliferation and may be involved in the pathogenesis of proliferative synovitis similar to that found in rheumatoid arthritis. Images PMID:8408648

  20. Th17 Cells in Type 1 Diabetes: Role in the Pathogenesis and Regulation by Gut Microbiome

    PubMed Central

    Li, Yangyang; Liu, Yu; Chu, Cong-Qiu

    2015-01-01

    Type 1 diabetes (T1D) is an autoimmune disease which is characterized by progressive destruction of insulin producing pancreatic islet β cells. The risk of developing T1D is determined by both genetic and environmental factors. A growing body of evidence supports an important role of T helper type 17 (Th17) cells along with impaired T regulatory (Treg) cells in the development of T1D in animal models and humans. Alteration of gut microbiota has been implicated to be responsible for the imbalance between Th17 and Treg cells. However, there is controversy concerning a pathogenic versus protective role of Th17 cells in murine models of diabetes in the context of influence of gut microbiota. In this review we will summarize current knowledge about Th17 cells and gut microbiota involved in T1D and propose Th17 targeted therapy in children with islet autoimmunity to prevent progression to overt diabetes. PMID:26843788

  1. The replication and titration of iridescent virus type 22 in Spodoptera frugiperda cells.

    PubMed

    Brown, D A; Lescott, T; Harrap, K A; Kelly, D C

    1978-01-01

    A plaque assay for iridescent virus type 22 (from Simulium sp.) using Spodoptera frugiperda cells has been devised, and the kinetics of growth of the virus in this cell line have been determined. The virus particle/p.f.u. ratio was 75 +/- 8, and the p.f.u./TCID50 ratio was 0.56 +/- 0.11.

  2. Poliovirus type 1 infection of murine PRNP-knockout neuronal cells.

    PubMed

    Baj, Andreina; Bettaccini, Alessia; Nishimura, Takuya; Onodera, Takashi; Toniolo, Antonio

    2005-07-01

    Transfection of the prion protein gene (Prnp) into prion-deficient mouse cells was shown to reduce the replication of coxsackievirus B3, an enterovirus. Because mice can be susceptible to poliovirus infection by parenteral routes, the authors tested the susceptibility to poliovirus-1 (PV-1) of a panel of murine neuronal cell lines differing in their ability to express Prnp. The investigated cell lines (prionless HpL3.4 cells, HpL3.4 cells transfected with a Prnp vector, HpL3.4 cells transfected with a void vector, wild-type Hw3.5 Prnp(+/+) cells) expressed the murine homologue (Tage4) of human poliovirus receptor (CD155/hPVR). PV-1 infection of Prnp(-/-) HpL3.4 cells resulted in the production of high viral titers, though viral antigens could be detected in only 0.5% to 2% of cells. Wild-type Prnp(+/+) cells and prionless cells transfected with the Prnp gene were not permissive to PV-1. Results of viral titration and immunofluorescence were confirmed by conventional polymerase chain reaction (PCR) and quantitative real-time PCR. Exposure to PV-1 had no influence on the gene expression profile of Prnp(+/+) cells. In contrast, PV-1 infection was associated with upregulation of several genes in permissive Prnp(-/-) cell cultures: type I interferon (IFN) genes, IFN-related developmental regulator 1 (IFNRD1), tumor necrosis factor superfamily member 13b (TNFSF13b), interleukin (IL) - 7, granulocyte/macrophage colony-stimulating factors (CSFs), hepatocyte growth factor (HGF), vascular endothelial growth factor-A, transforming growth factors beta1 and beta3 (TGFb1, TGFb3), as well as a variety of bone morphogenetic proteins endowed with neuroprotective activity. Distinction of permissive from nonpermissive neuronal cells on the basis of Prnp expression suggests that prion-deficient mice could represent an extraordinarily sensitive animal model for poliovirus infection.

  3. New World Hantaviruses Activate IFNλ Production in Type I IFN-Deficient Vero E6 Cells

    PubMed Central

    Prescott, Joseph; Hall, Pamela; Acuna-Retamar, Mariana; Ye, Chunyan; Wathelet, Marc G.; Ebihara, Hideki; Feldmann, Heinz; Hjelle, Brian

    2010-01-01

    Background Hantaviruses indigenous to the New World are the etiologic agents of hantavirus cardiopulmonary syndrome (HCPS). These viruses induce a strong interferon-stimulated gene (ISG) response in human endothelial cells. African green monkey-derived Vero E6 cells are used to propagate hantaviruses as well as many other viruses. The utility of the Vero E6 cell line for virus production is thought to owe to their lack of genes encoding type I interferons (IFN), rendering them unable to mount an efficient innate immune response to virus infection. Interferon λ, a more recently characterized type III IFN, is transcriptionally controlled much like the type I IFNs, and activates the innate immune system in a similar manner. Methodology/Principal Findings We show that Vero E6 cells respond to hantavirus infection by secreting abundant IFNλ. Three New World hantaviruses were similarly able to induce IFNλ expression in this cell line. The IFNλ contained within virus preparations generated with Vero E6 cells independently activates ISGs when used to infect several non-endothelial cell lines, whereas innate immune responses by endothelial cells are specifically due to viral infection. We show further that Sin Nombre virus replicates to high titer in human hepatoma cells (Huh7) without inducing ISGs. Conclusions/Significance Herein we report that Vero E6 cells respond to viral infection with a highly active antiviral response, including secretion of abundant IFNλ. This cytokine is biologically active, and when contained within viral preparations and presented to human epithelioid cell lines, results in the robust activation of innate immune responses. We also show that both Huh7 and A549 cell lines do not respond to hantavirus infection, confirming that the cytoplasmic RNA helicase pathways possessed by these cells are not involved in hantavirus recognition. We demonstrate that Vero E6 actively respond to virus infection and inhibiting IFNλ production in these cells

  4. Detection of Clostridium novyi type B alpha toxin by cell culture systems.

    PubMed

    Borrmann, E; Schulze, F

    1999-07-01

    Ten permanent cell lines were examined for their reaction to the Clostridium novyi alpha toxin. The action of the toxin was determined after 3 days by microscopic examination and the MTT assay. The alpha toxin exhibited the strongest effect on ESH-L cells rather than other cell lines. Vero and SFT-R cells reacted in a comparable way, but less sensitively. We were able to show that the cytopathic effect on the three types of cells was neutralised by the international standard for gas gangrene antitoxin (C. novyi) but in no case by heterologous antisera. Our results have shown that the three cell lines were specific indicators for the detection of the cytopathic effect of alpha toxin. The cytopathic effect can be measured reproducibly by the cell culture assay used. These results are suitable as the starting point for the development of the neutralisation test using cell cultures.

  5. Can thymic epithelial cells be infected by human T-lymphotropic virus type 1?

    PubMed

    Moreira-Ramos, Klaysa; Castro, Flávia Madeira Monteiro de; Linhares-Lacerda, Leandra; Savino, Wilson

    2011-09-01

    The human T-lymphotropic virus type-1 (HTLV-1) is the cause of adult T cell leukaemias/lymphoma. Because thymic epithelial cells (TEC) express recently defined receptors for the virus, it seemed conceivable that these cells might be a target for HTLV-1 infection. We developed an in vitro co-culture system comprising HTLV-1+-infected T cells and human TECs. Infected T cells did adhere to TECs and, after 24 h, the viral proteins gp46 and p19 were observed in TECs. After incubating TECs with culture supernatants from HTLV-1+-infected T cells, we detected gp46 on TEC membranes and the HTLV-1 tax gene integrated in the TEC genome. In conclusion, the human thymic epithelium can be infected in vitro by HTLV-1, not only via cell-cell contact, but also via exposure to virus-containing medium. PMID:22012233

  6. Causes of upregulation of glycolysis in lymphocytes upon stimulation. A comparison with other cell types.

    PubMed

    Stark, Heiko; Fichtner, Maximilian; König, Rainer; Lorkowski, Stefan; Schuster, Stefan

    2015-11-01

    In this review, we revisit the metabolic shift from respiration to glycolysis in lymphocytes upon activation, which is known as the Warburg effect in tumour cells. We compare the situation in lymphocytes with those in several other cell types, such as muscle cells, Kupffer cells, microglia cells, astrocytes, stem cells, tumour cells and various unicellular organisms (e.g. yeasts). We critically discuss and compare several explanations put forward in the literature for the observation that proliferating cells adopt this apparently less efficient pathway: hypoxia, poisoning of competitors by end products, higher ATP production rate, higher precursor supply, regulatory effects, and avoiding harmful effects (e.g. by reactive oxygen species). We conclude that in the case of lymphocytes, increased ATP production rate and precursor supply are the main advantages of upregulating glycolysis.

  7. Retinyl ester synthesis by isolated adult rabbit lung type II cells.

    PubMed

    Zachman, R D; Tsao, F H

    1988-01-01

    Type II alveolar cells were isolated from adult rabbit lungs and then cultured on monolayers for 16 hours. These cells were then covered with buffered medium containing [3H]-retinol. After 30-120 minutes incubation, the cells were extracted with Hexane: Ethanol and the hexane extract analyzed by HPLC. A linear synthesis of [3H]-retinyl palmitate with time of incubation was demonstrated. PMID:3170088

  8. Anaesthetics may change the shape of isolated type I hair cells.

    PubMed

    Scarfone, E; Ulfendahl, M; Figueroa, L; Flock, A

    1991-08-01

    Type I hair cells isolated from animals anaesthetised with barbiturates or ether were found to be shorter and to lack a prominent 'neck' region when compared to cells isolated from non-anaesthetised animals. Ketamine did not have this effect. The changes observed could have important implications for the physiology of inner ear receptors. These findings infer that care should be taken in the choice of anaesthetics used in studies on cells from the inner ear.

  9. Ultrastructural characteristics of type A epithelioid cells during BCG-granulomatosis and treatment with lysosomotropic isoniazid.

    PubMed

    Shkurupii, V A; Kozyaev, M A; Nadeev, A P

    2006-04-01

    We studied BCG-granulomas, their cellular composition, and ultrastructure of type A epithelioid cells in the liver of male BALB/c mice with spontaneous granulomatous inflammation. The animals received free isoniazid or isoniazid conjugated with lysosomotropic intracellularly prolonged matrix (dialdehyde dextran, molecular weight 65-75 kDa). Lysosomotropic isoniazid was accumulated in the vacuolar apparatus of epithelioid cells and produced a stimulatory effect on plastic processes in these cells.

  10. Therapeutic effect of mesenchymal multipotent stromal cells on memory in animals with Alzheimer-type neurodegeneration.

    PubMed

    Bobkova, N V; Poltavtseva, R A; Samokhin, A N; Sukhikh, G T

    2013-11-01

    Transplantation of human mesenchymal multipotent stromal cells improved spatial memory in bulbectomized mice with Alzheimer-type neurodegeneration. The positive effect was observed in 1 month after intracerebral transplantation and in 3 months after systemic injection of mesenchymal multipotent stromal cells. No cases of malignant transformation were noted. These findings indicate prospects of using mesenchymal multipotent stromal cells for the therapy of Alzheimer disease and the possibility of their systemic administration for attaining the therapeutic effect.

  11. A Single-Cell-Type Real-Time PCR Method Based on a Modified Patch-Pipette Cell Harvesting System.

    PubMed

    Song, Yuanlong; Zhang, Miaomiao; Tao, Xiaoqing; Xu, Zifen; Zhang, Liangpin; Zheng, Yunjie; Zhu, Minjie; Gao, Linlin

    2016-09-01

    Real-time PCR is a powerful tool for quantifying nucleic acid expression. Real-time PCR is conventionally performed at the tissue level to guarantee an abundance of nucleic acid for detection. The precision and reliability of this method, however, is limited by usually being composed of a mixture of different cell types. Single-cell PCR, in contrast, eliminates the purity problem of the cell source. However, use of this method is usually impeded by difficulties in cell harvesting and stringent requirements for processing of very small quantities of nucleic acids. In this study, we combined the advantages of the high purity of selected cells in single-cell PCR with the greater nucleic acid quantities and thus greater ease of tissue-level PCR. The key aspect of our method is to use a modified patch-clamp pipette to harvest several selected cells of the same type. This method is therefore especially useful for cells that can be morphologically or histologically identified such as primary sensory neurons, striated muscle fibers and cells labeled with fluorescent makers. PMID:27271017

  12. Vascular endothelial cells have impaired capacity to present immunodominant, antigenic peptides: a mechanism of cell type-specific immune escape.

    PubMed

    Kummer, Marco; Lev, Avital; Reiter, Yoram; Biedermann, Barbara C

    2005-02-15

    Vascular endothelial cells (EC) are an exposed target tissue in the course of CTL-mediated alloimmune diseases such as graft-vs-host disease (GVHD) or solid organ transplant rejection. The outcome of an interaction between CTL and target cells is determined by the amount of Ag presented and the costimulatory signals delivered by the target cells. We compared human EC with leukocytes and epithelial cells as targets for peptide-specific, MHC class I-restricted CTL clones. EC were poor targets for immunodominant CTL. Both endogenously processed antigenic proteins and exogenously added antigenic peptides are presented at 50- to 5000-fold lower levels on EC compared with any other target cell analyzed. This quantitative difference fully explained the poor CTL-mediated killing of EC. There was no evidence that lack of costimulation would contribute significantly to this cell type-specific difference in CTL activation. An HLA-A2-specific CTL clone that killed a broad selection of HLA A2-positive target cells equally well, killed EC less efficiently. Our data suggest that EC present a different Ag repertoire compared with other cell types. By this mechanism, these cells may escape an attack by effector CTL, which have been educated by professional APCs and are specific for immunodominant antigenic peptides. PMID:15699122

  13. Collagen Type I Improves the Differentiation of Human Embryonic Stem Cells towards Definitive Endoderm

    PubMed Central

    Rasmussen, Camilla Holzmann; Petersen, Dorthe Roenn; Moeller, Jonas Bech

    2015-01-01

    Human embryonic stem cells have the ability to generate all cell types in the body and can potentially provide an unlimited source of cells for cell replacement therapy to treat degenerative diseases such as diabetes. Current differentiation protocols of human embryonic stem cells towards insulin producing beta cells focus on soluble molecules whereas the impact of cell-matrix interactions has been mainly unattended. In this study almost 500 different extracellular matrix protein combinations were screened to systemically identify extracellular matrix proteins that influence differentiation of human embryonic stem cells to the definitive endoderm lineage. The percentage of definitive endoderm cells after differentiation on collagen I and fibronectin was >85% and 65%, respectively. The cells on collagen I substrates displayed different morphology and gene expression during differentiation as assessed by time lapse studies compared to cells on the other tested substrates. Global gene expression analysis showed that cells differentiated on collagen I were largely similar to cells on fibronectin after completed differentiation. Collectively, the data suggest that collagen I induces a more rapid and consistent differentiation of stem cells to definitive endoderm. The results shed light on the importance of extracellular matrix proteins for differentiation and also points to a cost effective and easy method to improve differentiation. PMID:26713616

  14. Hyperactivity of ON-type retinal ganglion cells in streptozotocin-induced diabetic mice.

    PubMed

    Yu, Jun; Wang, Lu; Weng, Shi-Jun; Yang, Xiong-Li; Zhang, Dao-Qi; Zhong, Yong-Mei

    2013-01-01

    Impairment of visual function has been detected in the early stage of diabetes but the underlying neural mechanisms involved are largely unknown. Morphological and functional alterations of retinal ganglion cells, the final output neurons of the vertebrate retina, are thought to be the major cause of visual defects in diabetes but direct evidence to support this notion is limited. In this study we investigated functional changes of retinal ganglion cells in a type 1-like diabetic mouse model. Our results demonstrated that the spontaneous spiking activity of ON-type retinal ganglion cells was increased in streptozotocin-diabetic mice after 3 to 4 months of diabetes. At this stage of diabetes, no apoptotic signals or cell loss were detected in the ganglion cell layer of the retina, suggesting that the functional alterations in ganglion cells occur prior to massive ganglion cell apoptosis. Furthermore, we found that the increased activity of ON-type ganglion cells was mainly a result of reduced inhibitory signaling to the cells in diabetes. This novel mechanism provides insight into how visual function is impaired in diabetic retinopathy. PMID:24069457

  15. Cell-Type-Specific mRNA Purification by Translating Ribosome Affinity Purification (TRAP)

    PubMed Central

    Heiman, Myriam; Kulicke, Ruth; Fenster, Robert J.; Greengard, Paul; Heintz, Nathaniel

    2014-01-01

    Cellular diversity and architectural complexity create barriers to understanding the function of the mammalian central nervous system (CNS) at a molecular level. To address this problem, we recently developed a methodology that provides the ability to profile the entire translated mRNA complement of any genetically defined cell population. This methodology, which we termed translating ribosome affinity purification, or TRAP, combines cell-type-specific transgene expression with affinity purification of translating ribosomes. TRAP can be used to study the cell-type-specific mRNA profiles of any genetically defined cell type, and has been successfully used to date in organisms ranging from D. melanogaster to mice and human cultured cells. Unlike other methodologies that rely upon micro-dissection, cell panning, or cell sorting, the TRAP methodology bypasses the need for tissue fixation or single-cell suspensions (and potential artifacts these treatments introduce), and reports on mRNAs in the entire cell body. This protocol provides a step-by-step guide to implementing the TRAP methodology, which takes two days to complete once all materials are in hand. PMID:24810037

  16. Mesenchymal Stem Cells Sense Three Dimensional Type I Collagen through Discoidin Domain Receptor 1.

    PubMed

    Lund, A W; Stegemann, J P; Plopper, G E

    2009-01-01

    The extracellular matrix provides structural and organizational cues for tissue development and defines and maintains cellular phenotype during cell fate determination. Multipotent mesenchymal stem cells use this matrix to tightly regulate the balance between their differentiation potential and self-renewal in the native niche. When understood, the mechanisms that govern cell-matrix crosstalk during differentiation will allow for efficient engineering of natural and synthetic matrices to specifically direct and maintain stem cell phenotype. This work identifies the discoidin domain receptor 1 (DDR1), a collagen activated receptor tyrosine kinase, as a potential link through which stem cells sense and respond to the 3D organization of their extracellular matrix microenvironment. DDR1 is dependent upon both the structure and proteolytic state of its collagen ligand and is specifically expressed and localized in three dimensional type I collagen culture. Inhibition of DDR1 expression results in decreased osteogenic potential, increased cell spreading, stress fiber formation and ERK1/2 phosphorylation. Additionally, loss of DDR1 activity alters the cell-mediated organization of the naïve type I collagen matrix. Taken together, these results demonstrate a role for DDR1 in the stem cell response to and interaction with three dimensional type I collagen. Dynamic changes in cell shape in 3D culture and the tuning of the local ECM microstructure, directs crosstalk between DDR1 and two dimensional mechanisms of osteogenesis that can alter their traditional roles.

  17. Cell-type-specific neuroanatomy of cliques of autism-related genes in the mouse brain

    PubMed Central

    Grange, Pascal; Menashe, Idan; Hawrylycz, Michael

    2015-01-01

    Two cliques of genes identified computationally for their high co-expression in the mouse brain according to the Allen Brain Atlas, and for their enrichment in genes related to autism spectrum disorder (ASD), have recently been shown to be highly co-expressed in the cerebellar cortex, compared to what could be expected by chance. Moreover, the expression of these cliques of genes is not homogeneous across the cerebellar cortex, and it has been noted that their expression pattern seems to highlight the granular layer. However, this observation was only made by eye, and recent advances in computational neuroanatomy allow to rank cell types in the mouse brain (characterized by their transcriptome profiles) according to the similarity between their spatial density profiles and the spatial expression profiles of the cliques. We establish by Monte Carlo simulation that with probability at least 99%, the expression profiles of the two cliques are more similar to the density profile of granule cells than 99% of the expression of cliques containing the same number of genes (Purkinje cells also score above 99% in one of the cliques). Thresholding the expression profiles shows that the signal is more intense in the granular layer. Finally, we work out pairs of cell types whose combined expression profiles are more similar to the expression profiles of the cliques than any single cell type. These pairs predominantly consist of one cortical pyramidal cell and one cerebellar cell (which can be either a granule cell or a Purkinje cell). PMID:26074809

  18. RNA interference-mediated silencing of speckle-type POZ protein promotes apoptosis of renal cell cancer cells

    PubMed Central

    Liu, Xiaoxia; Sun, Guiling; Sun, Xiuju

    2016-01-01

    This study aimed to investigate the effects of silencing the speckle-type POZ protein (SPOP) gene on renal cell cancer (RCC) cells and to explore its possible mechanism. The A498 and ACHN RCC cells were transfected with small interference RNA (siRNA)-SPOP by lipofection methods. The silencing efficiency was monitored by quantitative real-time polymerase chain reaction and Western blot. The effects of SPOP silencing on cell apoptosis, cell viability, colony formation ability, cell migration ability, and chemosensitivity to Sorafenib were assessed by flow cytometry, an MTT assay, a colony formation assay, a trans-well migration assay, and a CCK-8 assay, respectively. Its effects on the expression of several cytokines were determined by a protein microarray. Relevant signaling pathways were also analyzed. Compared with the control group, the cell apoptosis rate was significantly higher; the cell viability, the colony formation, and migration ability were significantly decreased in the siRNA-SPOP group. The protein microarray screening showed that the expression of vascular endothelial growth factor receptor, matrix metallopeptidase-9, vascular cell adhesion molecule-1, and stromal cell-derived factor-1 in the siRNA group was significantly decreased and that the expression of granulocyte–macrophage colony-stimulating factor and E-cadherin was significantly increased (P<0.05). The relevant signaling pathways were the integrin-mediated cell surface interactions pathway and extracellular matrix organization signal pathway. SPOP gene silencing induced cell apoptosis, decreased cell viability, colony formation, and migration ability, and elevated the drug sensitivity in the RCC cells. A possible mechanism is that silencing SPOP induces the differential expression of E-cadherin, vascular endothelial growth factor receptor, matrix metallopeptidase-9, and vascular cell adhesion molecule, which are related to the integrin-mediated cell surface interactions and extracellular

  19. RNA interference-mediated silencing of speckle-type POZ protein promotes apoptosis of renal cell cancer cells.

    PubMed

    Liu, Xiaoxia; Sun, Guiling; Sun, Xiuju

    2016-01-01

    This study aimed to investigate the effects of silencing the speckle-type POZ protein (SPOP) gene on renal cell cancer (RCC) cells and to explore its possible mechanism. The A498 and ACHN RCC cells were transfected with small interference RNA (siRNA)-SPOP by lipofection methods. The silencing efficiency was monitored by quantitative real-time polymerase chain reaction and Western blot. The effects of SPOP silencing on cell apoptosis, cell viability, colony formation ability, cell migration ability, and chemosensitivity to Sorafenib were assessed by flow cytometry, an MTT assay, a colony formation assay, a trans-well migration assay, and a CCK-8 assay, respectively. Its effects on the expression of several cytokines were determined by a protein microarray. Relevant signaling pathways were also analyzed. Compared with the control group, the cell apoptosis rate was significantly higher; the cell viability, the colony formation, and migration ability were significantly decreased in the siRNA-SPOP group. The protein microarray screening showed that the expression of vascular endothelial growth factor receptor, matrix metallopeptidase-9, vascular cell adhesion molecule-1, and stromal cell-derived factor-1 in the siRNA group was significantly decreased and that the expression of granulocyte-macrophage colony-stimulating factor and E-cadherin was significantly increased (P<0.05). The relevant signaling pathways were the integrin-mediated cell surface interactions pathway and extracellular matrix organization signal pathway. SPOP gene silencing induced cell apoptosis, decreased cell viability, colony formation, and migration ability, and elevated the drug sensitivity in the RCC cells. A possible mechanism is that silencing SPOP induces the differential expression of E-cadherin, vascular endothelial growth factor receptor, matrix metallopeptidase-9, and vascular cell adhesion molecule, which are related to the integrin-mediated cell surface interactions and extracellular

  20. Is Hydrogen Peroxide a Suitable Apoptosis Inducer for All Cell Types?

    PubMed Central

    Xiang, Jinmei; Wan, Chunyun; Guo, Rui

    2016-01-01

    Hydrogen peroxide is currently the most widely used apoptosis inducer due to its broad cytotoxic efficacy against nearly all cell types. However, equivalent cytotoxicity is achieved over a wide range of doses, although the reasons for this differential sensitivity are not always clear. In this study, three kinds of cells, the 293T cell line, primary fibroblasts, and terminally differentiated myocardial cells, were treated with a wide range of H2O2 doses. Times to apoptosis initiation and end were measured cytochemically and the changes in expression of caspase-9, P53, NF-κB, and RIP were determined by RT-PCR. The 293T cell line was the most sensitive to H2O2, undergoing necroptosis and/or apoptosis at all concentrations from 0.1 to 1.6 mM. At > 0.4 mM, H2O2 also caused necroptosis in primary cells. At < 0.4 mM, however, primary cells exhibited classic signs of apoptosis, although they tended to survive for 36 hours in < 0.2 mM H2O2. Thus, H2O2 is a broadly effective apoptosis inducer, but the dose range differs by cell type. For cell lines, a low dose is required and the exposure time must be reduced compared to primary cells to avoid cell death primarily by necroptosis or necrosis. PMID:27595106

  1. Is Hydrogen Peroxide a Suitable Apoptosis Inducer for All Cell Types?

    PubMed Central

    Xiang, Jinmei; Wan, Chunyun; Guo, Rui

    2016-01-01

    Hydrogen peroxide is currently the most widely used apoptosis inducer due to its broad cytotoxic efficacy against nearly all cell types. However, equivalent cytotoxicity is achieved over a wide range of doses, although the reasons for this differential sensitivity are not always clear. In this study, three kinds of cells, the 293T cell line, primary fibroblasts, and terminally differentiated myocardial cells, were treated with a wide range of H2O2 doses. Times to apoptosis initiation and end were measured cytochemically and the changes in expression of caspase-9, P53, NF-κB, and RIP were determined by RT-PCR. The 293T cell line was the most sensitive to H2O2, undergoing necroptosis and/or apoptosis at all concentrations from 0.1 to 1.6 mM. At > 0.4 mM, H2O2 also caused necroptosis in primary cells. At < 0.4 mM, however, primary cells exhibited classic signs of apoptosis, although they tended to survive for 36 hours in < 0.2 mM H2O2. Thus, H2O2 is a broadly effective apoptosis inducer, but the dose range differs by cell type. For cell lines, a low dose is required and the exposure time must be reduced compared to primary cells to avoid cell death primarily by necroptosis or necrosis.

  2. Regulating the beta cell mass as a strategy for type-2 diabetes treatment.

    PubMed

    Song, Imane; Muller, Christo; Louw, Johan; Bouwens, Luc

    2015-01-01

    The incidence of type 2 diabetes (T2D) increases dramatically worldwide and has created an enormous health care burden. Obesity, dyslipidemia and insulin resistance are major risk factors for the development of T2D, but the major factor leading to the disease is failure of the insulin-producing beta cell mass to compensate for increasing insulin demands of the body. Progression of the disease further diminishes the beta cell mass as a result of lipotoxicity and glucotoxicity for which beta cells are particularly sensitive. Hence, treatment aiming to prevent beta cell loss or increase the number of beta cells could inhibit diabetes progression or lead to restoration of normal metabolism. Whereas current and new antidiabetic drugs are mainly targeting insulin secretion and action or glucose uptake, newer interventions must be found that prevent beta cell loss or increase beta cell number. The targets for this are beta cell proliferation, neogenesis and survival. This review examines major evidence from animal experiments suggesting that it is feasible to regulate the beta cell mass by bioactive compounds like growth factors, cytokines, hormones, phytochemicals and small molecules. Often the mode of action remains unclear due to inadequate methods to assess the effects of the compounds on the beta cell dynamics. Furthermore, a major challenge is to identify compounds with sufficient specificity in order to avoid unwanted effects on other cell types. Provided such safety issues can be solved, this may provide a curative approach for diabetes treatment.

  3. Postnatal disappearance of type A intercalated cells in carbonic anhydrase II-deficient mice.

    PubMed

    Brion, L P; Suarez, C; Saenger, P

    2001-06-01

    Despite chronic acidosis, collecting ducts in adult carbonic anhydrase II-deficient (CAD mice) are depleted of intercalated cells, including those of type A, which are acid-secreting cells. We hypothesized that this depletion could occur during postnatal development. Principal cells were identified by immunofluorescence using an antibody to rat aquaporin-2 (AQP-2), and type A intercalated cells using an antibody specific for anion exchanger (AE1). In CAD mice the proportion of AQP2-positive cells, normal at 11 days, increased progressively in the cortical (CCD) and outer medullary collecting duct (OMCD), to reach almost 100% in the OMCD in adults. The percentage of AE1-positive cells in the OMCD of CAD mice decreased by half by 6 weeks of age and further by adulthood. In controls, however, the proportion of AQP2-positive cells and that of AE1-positive cells in the OMCD remained stable after 10 days of age. AE1-positive cells accounted for the majority of intercalated cells in the OMCD. The mechanisms leading to selective postnatal cell depletion in the collecting duct in CAD mice remain to be determined.

  4. Molecular modelling and experimental studies of mutation and cell-adhesion sites in the fibronectin type III and whey acidic protein domains of human anosmin-1.

    PubMed Central

    Robertson, A; MacColl, G S; Nash, J A; Boehm, M K; Perkins, S J; Bouloux, P M

    2001-01-01

    Anosmin-1, the gene product of the KAL gene, is implicated in the pathogenesis of X-linked Kallmann's syndrome. Anosmin-1 protein expression is restricted to the basement membrane and interstitial matrix of tissues affected in this syndrome during development. The anosmin-1 sequence indicates an N-terminal cysteine-rich domain, a whey acidic protein (WAP) domain, four fibronectin type III (FnIII) domains and a C-terminal histidine-rich region, and shows similarity with cell-adhesion molecules, such as neural cell-adhesion molecule, TAG-1 and L1. We investigated the structural and functional significance of three loss-of-function missense mutations of anosmin-1 using comparative modelling of the four FnIII and the WAP domains based on known NMR and crystal structures. Three missense mutation-encoded amino acid substitutions, N267K, E514K and F517L, were mapped to structurally defined positions on the GFCC' beta-sheet face of the first and third FnIII domains. Electrostatic maps demonstrated large basic surfaces containing clusters of conserved predicted heparan sulphate-binding residues adjacent to these mutation sites. To examine these modelling results anosmin-1 was expressed in insect cells. The incorporation of the three mutations into recombinant anosmin-1 had no effect on its secretion. The removal of two dibasic motifs that may constitute potential physiological cleavage sites for anosmin-1 had no effect on cleavage. Peptides based on the anosmin-1 sequences R254--K285 and P504--K527 were then synthesized in order to assess the effect of the three mutations on cellular adhesion, using cell lines that represented potential functional targets of anosmin-1. Peptides (10 microg/ml) incorporating the N267K and E514K substitutions promoted enhanced adhesion to 13.S.1.24 rat olfactory epithelial cells and canine MDCK1 kidney epithelial cells (P<0.01) compared with the wild-type peptides. This result was attributed to the introduction of a lysine residue adjacent to

  5. Analysis of type II diabetes mellitus adipose-derived stem cells for tissue engineering applications

    PubMed Central

    Minteer, Danielle Marie; Young, Matthew T; Lin, Yen-Chih; Over, Patrick J; Rubin, J Peter; Gerlach, Jorg C

    2015-01-01

    To address the functionality of diabetic adipose-derived stem cells in tissue engineering applications, adipose-derived stem cells isolated from patients with and without type II diabetes mellitus were cultured in bioreactor culture systems. The adipose-derived stem cells were differentiated into adipocytes and maintained as functional adipocytes. The bioreactor system utilizes a hollow fiber–based technology for three-dimensional perfusion of tissues in vitro, creating a model in which long-term culture of adipocytes is feasible, and providing a potential tool useful for drug discovery. Daily metabolic activity of the adipose-derived stem cells was analyzed within the medium recirculating throughout the bioreactor system. At experiment termination, tissues were extracted from bioreactors for immunohistological analyses in addition to gene and protein expression. Type II diabetic adipose-derived stem cells did not exhibit significantly different glucose consumption compared to adipose-derived stem cells from patients without type II diabetes (p > 0.05, N = 3). Expression of mature adipocyte genes was not significantly different between diabetic/non-diabetic groups (p > 0.05, N = 3). Protein expression of adipose tissue grown within all bioreactors was verified by Western blotting.The results from this small-scale study reveal adipose-derived stem cells from patients with type II diabetes when removed from diabetic environments behave metabolically similar to the same cells of non-diabetic patients when cultured in a three-dimensional perfusion bioreactor, suggesting that glucose transport across the adipocyte cell membrane, the hindrance of which being characteristic of type II diabetes, is dependent on environment. The presented observation describes a tissue-engineered tool for long-term cell culture and, following future adjustments to the culture environment and increased sample sizes, potentially for anti-diabetic drug testing. PMID:26090087

  6. [Nasal type natural killer/T cell lymphoma: case series and literature review].

    PubMed

    Düzlü, Mehmet; Ant, Ayça; Tutar, Hakan; Karamert, Recep; Şahin, Melih; Sayar, Erolcan; Cesur, Nesibe

    2016-01-01

    Nasal type natural killer/T-cell lymphoma is a rare type of extranodal non-Hodgkin lymphoma which originates from nasal cavity and paranasal sinuses. Exact diagnosis of nasal natural killer/T-cell lymphoma, which is a rapidly progressive clinical condition, may be established by immunohistochemical analysis on biopsy material after clinical suspicion. In this article, we report four cases of nasal natural killer/T-cell lymphoma who were followed-up in our clinic and discuss the diagnosis and treatment of the disease in light of the literature data. PMID:27405082

  7. Cell type-dependent Erk-Akt pathway crosstalk regulates the proliferation of fetal neural progenitor cells.

    PubMed

    Rhim, Ji Heon; Luo, Xiangjian; Gao, Dongbing; Xu, Xiaoyun; Zhou, Tieling; Li, Fuhai; Wang, Ping; Wong, Stephen T C; Xia, Xiaofeng

    2016-01-01

    Neural progenitor (NP) cells are the multipotent cells that produce neurons and glia in the central nervous system. Compounds regulating their proliferation are key to both understanding brain development and unlocking their potential in regenerative repair. We discuss a chemical screen that unexpectedly identified inhibitors of Erk signaling potently promoting the self-renewing divisions of fetal NP cells. This occurred through crosstalk between Erk and Akt signaling cascades. The crosstalk mechanism is cell type-specific, and is not detected in adult NP cells as well as brain tumor cells. The mechanism was also shown to be independent from the GSK-3 signaling pathway, which has been reported to be a major regulator of NP cell homeostasis and inhibitors to which were also identified in the screen. In vitro Erk inhibition led to the prolonged rapid expansion of fetal NP cells while retaining their multipotency. In vivo inhibitor administration significantly inhibited the neuronal differentiation, and resulted in increased proliferative progenitor cells in the ventricular/subventricular zone (VZ/SVZ) of the embryonic cortex. Our results uncovered a novel regulating pathway for NP cell proliferation in the developing brain. The discovery provides a pharmacological basis for in vitro expansion and in vivo manipulation of NP cells. PMID:27211495

  8. Importance of brain‑type fatty acid binding protein for cell-biological processes in human renal carcinoma cells.

    PubMed

    Tölle, Angelika; Krause, Hans; Miller, Kurt; Jung, Klaus; Stephan, Carsten

    2011-05-01

    The molecular mechanisms underlying renal cell carcinoma (RCC) development and progression are still not completely understood. The importance of fatty acid binding proteins (FABP) for the progression of carcinomas has been shown for several tumors. However, the importance of brain-type FABP (B‑FABP) in cell-biological processes in renal carcinoma cells is unknown. Therefore, it was the aim of this study to evaluate the role of B‑FABP in processes such as proliferation, migration and invasion. By using the approach of down- and up-regulation of B‑FABP in human kidney carcinoma cells Caki‑2 and Caki‑1, the potential participation of B‑FABP in proliferation, migration and invasion was demonstrated. B‑FABP was down-regulated at both mRNA and protein levels following treatment of Caki‑2 cells with B‑FABP siRNA. Down-regulation of B‑FABP decreased cell proliferation and migration but did not affect invasion. The transfection of Caki‑1 cells with human B‑FABP cDNA generated an increment of B‑FABP mRNA but the protein was not detectable. Transfected Caki‑1 cells developed a faster proliferation compared to untreated cells. An effect on the process of invasion was not observed. Our data suggest that B‑FABP is involved in cell proliferation and migration of human renal carcinoma cells. The detailed molecular mechanisms remain to be elucidated.

  9. Cell type-dependent Erk-Akt pathway crosstalk regulates the proliferation of fetal neural progenitor cells

    PubMed Central

    Rhim, Ji heon; Luo, Xiangjian; Gao, Dongbing; Xu, Xiaoyun; Zhou, Tieling; Li, Fuhai; Wang, Ping; Wong, Stephen T. C.; Xia, Xiaofeng

    2016-01-01

    Neural progenitor (NP) cells are the multipotent cells that produce neurons and glia in the central nervous system. Compounds regulating their proliferation are key to both understanding brain development and unlocking their potential in regenerative repair. We discuss a chemical screen that unexpectedly identified inhibitors of Erk signaling potently promoting the self-renewing divisions of fetal NP cells. This occurred through crosstalk between Erk and Akt signaling cascades. The crosstalk mechanism is cell type-specific, and is not detected in adult NP cells as well as brain tumor cells. The mechanism was also shown to be independent from the GSK-3 signaling pathway, which has been reported to be a major regulator of NP cell homeostasis and inhibitors to which were also identified in the screen. In vitro Erk inhibition led to the prolonged rapid expansion of fetal NP cells while retaining their multipotency. In vivo inhibitor administration significantly inhibited the neuronal differentiation, and resulted in increased proliferative progenitor cells in the ventricular/subventricular zone (VZ/SVZ) of the embryonic cortex. Our results uncovered a novel regulating pathway for NP cell proliferation in the developing brain. The discovery provides a pharmacological basis for in vitro expansion and in vivo manipulation of NP cells. PMID:27211495

  10. The type III secretion system of Vibrio alginolyticus induces rapid apoptosis, cell rounding and osmotic lysis of fish cells.

    PubMed

    Zhao, Zhe; Chen, Chang; Hu, Chao-Qun; Ren, Chun-Hua; Zhao, Jing-Jing; Zhang, Lv-Ping; Jiang, Xiao; Luo, Peng; Wang, Qing-Bai

    2010-09-01

    Vibrio alginolyticus is a Gram-negative bacterium and has been recognized as an opportunistic pathogen in humans as well as marine animals. However, the virulence mechanisms for this species of Vibrio have not been elucidated. This study characterized multiple mechanisms that induce cell death in fish cells upon infection with a V. alginolyticus strain, ZJO. The bacterium required its type III secretion system (T3SS) to cause rapid death of infected fish cells. Dying cells exhibited some features of apoptotic cells, such as membrane blebbing, nuclear condensation and DNA fragmentation. Further studies showed that caspase-3 was activated by the T3SS of the ZJO strain, confirming that infection with V. alginolyticus rapidly induces T3SS-dependent apoptosis in fish cells. Infection with the ZJO strain also led to membrane pore formation and release of cellular contents from infected fish cells, as evidenced by lactate dehydrogenase release and the uptake of a membrane-impermeable dye. Importantly, inhibition of apoptosis did not prevent ZJO-infected cells from releasing cellular contents and did not block cell rounding. Taken together, these data demonstrate that infection with V. alginolyticus may promote at least three different T3SS-dependent events, which lead to the death of fish cells. This study provides an important insight into the mechanism used by Vibrio species to cause host-cell death.

  11. Protective effect of catechin in type I Gaucher disease cells by reducing endoplasmic reticulum stress

    SciTech Connect

    Lee, Yea-Jin; Kim, Sung-Jo; Heo, Tae-Hwe

    2011-09-23

    Highlights: {yields} Catechin reduces the expression level of ER stress marker protein in type I Gaucher disease cells. {yields} Catechin induces the proliferation rate of GD cells similar levels to normal cells. {yields} Catechin improves wound healing activity. {yields} Catechin-mediated reductions in ER stress may be associated with enhanced cell survival. {yields} We identified catechin as a protective agent against ER stress in GD cells. -- Abstract: Gaucher disease (GD) is the most common lysosomal storage disorder (LSD) and is divided into three phenotypes, I, II, and III. Type I is the most prevalent form and has its onset in adulthood. The degree of endoplasmic reticulum (ER) stress is one of the factors that determine GD severity. It has recently been reported that antioxidants reduce ER stress and apoptosis by scavenging the oxidants that cause oxidative stress. For this report, we investigated the possibility that catechin can act on type I GD patient cells to alleviate the pathogenic conditions of GD. We treated GD cells with catechin and examined the expression level of GRP78/BiP (an ER stress marker) by western blots and fluorescence microscopy, the proliferation rate of GD cells, and scratch-induced wound healing activity. Our results show that catechin reduces the expression level of GRP78/BiP, leads to cell proliferation rates of GD cells similar levels to normal cells, and improves wound healing activity. We conclude that catechin protects against ER stress in GD cells and catechin-mediated reductions in ER stress may be associated with enhanced cell survival.

  12. Cutting edge: pulmonary Legionella pneumophila is controlled by plasmacytoid dendritic cells but not type I IFN.

    PubMed

    Ang, Desmond K Y; Oates, Clare V L; Schuelein, Ralf; Kelly, Michelle; Sansom, Fiona M; Bourges, Dorothée; Boon, Louis; Hertzog, Paul J; Hartland, Elizabeth L; van Driel, Ian R

    2010-05-15

    Plasmacytoid dendritic cells (pDCs) are well known as the major cell type that secretes type I IFN in response to viral infections. Their role in combating other classes of infectious organisms, including bacteria, and their mechanisms of action are poorly understood. We have found that pDCs play a significant role in the acute response to the intracellular bacterial pathogen Legionella pneumophila. pDCs were rapidly recruited to the lungs of L. pneumophila-infected mice, and depletion of pDCs resulted in increased bacterial load. The ability of pDCs to combat infection did not require type I IFN. This study points to an unappreciated role for pDCs in combating bacterial infections and indicates a novel mechanism of action for this cell type.

  13. Low-cost zinc-plated photoanode for fabric-type dye-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Kong, Lingfeng; Bao, Yunna; Guo, Wanwan; Cheng, Li; Du, Jun; Liu, Renlong; Wang, Yundong; Fan, Xing; Tao, Changyuan

    2016-02-01

    Fabric-type flexible solar cells have been recently proposed as a very promising power source for wearable electronics. To increase the photocurrent of fabric-type flexible solar cells, low-cost zinc-plated wire and mesh photoanodes are assembled for the first time through a mild wet process. Given the protection of the compact protection layer, the DSSC device could benefit from the low work function of Zn and self-repairing behavior on the Zn/ZnO interface. An evident current increase by ∼6 mA/cm2 could be observed after coating a layer of metal Zn on various metal substrates, such as traditional stainless steel wire. Given the self-repairing behavior on Zn/ZnO interface, the Zn layer can help to improve the interfacial carrier transfer, leading to better photovoltaic performance, for both liquid-type and solid-type cells.

  14. The genome of the ctenophore Mnemiopsis leidyi and its implications for cell type evolution.

    PubMed

    Ryan, Joseph F; Pang, Kevin; Schnitzler, Christine E; Nguyen, Anh-Dao; Moreland, R Travis; Simmons, David K; Koch, Bernard J; Francis, Warren R; Havlak, Paul; Smith, Stephen A; Putnam, Nicholas H; Haddock, Steven H D; Dunn, Casey W; Wolfsberg, Tyra G; Mullikin, James C; Martindale, Mark Q; Baxevanis, Andreas D

    2013-12-13

    An understanding of ctenophore biology is critical for reconstructing events that occurred early in animal evolution. Toward this goal, we have sequenced, assembled, and annotated the genome of the ctenophore Mnemiopsis leidyi. Our phylogenomic analyses of both amino acid positions and gene content suggest that ctenophores rather than sponges are the sister lineage to all other animals. Mnemiopsis lacks many of the genes found in bilaterian mesodermal cell types, suggesting that these cell types evolved independently. The set of neural genes in Mnemiopsis is similar to that of sponges, indicating that sponges may have lost a nervous system. These results present a newly supported view of early animal evolution that accounts for major losses and/or gains of sophisticated cell types, including nerve and muscle cells. PMID:24337300

  15. The genome of the ctenophore Mnemiopsis leidyi and its implications for cell type evolution

    PubMed Central

    Ryan, Joseph F.; Pang, Kevin; Schnitzler, Christine E.; Nguyen, Anh-Dao; Moreland, R. Travis; Simmons, David K.; Koch, Bernard J.; Francis, Warren R.; Havlak, Paul; Smith, Stephen A.; Putnam, Nicholas H.; Haddock, Steven H. D.; Dunn, Casey W.; Wolfsberg, Tyra G.; Mullikin, James C.; Martindale, Mark Q.; Baxevanis, Andreas D.

    2014-01-01

    An understanding of ctenophore biology is critical for reconstructing events that occurred early in animal evolution. Towards this goal, we have sequenced, assembled, and annotated the genome of the ctenophore Mnemiopsis leidyi. Our phylogenomic analyses of both amino acid positions and gene content suggests that ctenophores rather than sponges are the sister lineage to all other animals. Mnemiopsis lacks many of the genes found in bilaterian mesodermal cell types, suggesting that these cell types evolved independently. The set of neural genes in Mnemiopsis is similar to that of sponges, indicating that sponges may have lost a nervous system. These results present a new view of early animal evolution that accounts for major losses and/or gains of sophisticated cell types, including nerve and muscle cells. PMID:24337300

  16. The genome of the ctenophore Mnemiopsis leidyi and its implications for cell type evolution.

    PubMed

    Ryan, Joseph F; Pang, Kevin; Schnitzler, Christine E; Nguyen, Anh-Dao; Moreland, R Travis; Simmons, David K; Koch, Bernard J; Francis, Warren R; Havlak, Paul; Smith, Stephen A; Putnam, Nicholas H; Haddock, Steven H D; Dunn, Casey W; Wolfsberg, Tyra G; Mullikin, James C; Martindale, Mark Q; Baxevanis, Andreas D

    2013-12-13

    An understanding of ctenophore biology is critical for reconstructing events that occurred early in animal evolution. Toward this goal, we have sequenced, assembled, and annotated the genome of the ctenophore Mnemiopsis leidyi. Our phylogenomic analyses of both amino acid positions and gene content suggest that ctenophores rather than sponges are the sister lineage to all other animals. Mnemiopsis lacks many of the genes found in bilaterian mesodermal cell types, suggesting that these cell types evolved independently. The set of neural genes in Mnemiopsis is similar to that of sponges, indicating that sponges may have lost a nervous system. These results present a newly supported view of early animal evolution that accounts for major losses and/or gains of sophisticated cell types, including nerve and muscle cells.

  17. Gene expression profiles and neural activities of Kenyon cell subtypes in the honeybee brain: identification of novel 'middle-type' Kenyon cells.

    PubMed

    Kaneko, Kumi; Suenami, Shota; Kubo, Takeo

    2016-01-01

    In the honeybee (Apis mellifera L.), it has long been thought that the mushroom bodies, a higher-order center in the insect brain, comprise three distinct subtypes of intrinsic neurons called Kenyon cells. In class-I large-type Kenyon cells and class-I small-type Kenyon cells, the somata are localized at the edges and in the inner core of the mushroom body calyces, respectively. In class-II Kenyon cells, the somata are localized at the outer surface of the mushroom body calyces. The gene expression profiles of the large- and small-type Kenyon cells are distinct, suggesting that each exhibits distinct cellular characteristics. We recently identified a novel gene, mKast (middle-type Kenyon cell-preferential arrestin-related gene-1), which has a distinctive expression pattern in the Kenyon cells. Detailed expression analyses of mKast led to the discovery of novel 'middle-type' Kenyon cells characterized by their preferential mKast-expression in the mushroom bodies. The somata of the middle-type Kenyon cells are localized between the large- and small-type Kenyon cells, and the size of the middle-type Kenyon cell somata is intermediate between that of large- and small-type Kenyon cells. Middle-type Kenyon cells appear to differentiate from the large- and/or small-type Kenyon cell lineage(s). Neural activity mapping using an immediate early gene, kakusei, suggests that the small-type and some middle-type Kenyon cells are prominently active in the forager brain, suggesting a potential role in processing information during foraging flight. Our findings indicate that honeybee mushroom bodies in fact comprise four types of Kenyon cells with different molecular and cellular characteristics: the previously known class-I large- and small-type Kenyon cells, class-II Kenyon cells, and the newly identified middle-type Kenyon cells described in this review. As the cellular characteristics of the middle-type Kenyon cells are distinct from those of the large- and small-type Kenyon

  18. Proinflammatory cytokines contribute to development and function of regulatory T cells in type 1 diabetes.

    PubMed

    Thomas, Helen E; Graham, Kate L; Chee, Jonathan; Thomas, Ranjeny; Kay, Thomas W; Krishnamurthy, Balasubramanian

    2013-04-01

    Type 1 diabetes is caused by immune-mediated loss of pancreatic beta cells. It has been proposed that inflammatory cytokines play a role in killing beta cells. Expression of interleukin (IL)-1 and tumor necrosis factor (TNF-α) has been detected in islets from patients with type 1 diabetes, and these cytokines can induce beta cell death in vitro. We produced nonobese diabetic (NOD) mice lacking receptors for these cytokines. Islets from mice lacking IL-1RI or TNFR1 were killed when transplanted into wild-type NOD mice, suggesting that cytokine action on beta cells is not required for killing. Mice lacking TNFR1 did not develop diabetes, and mice lacking IL-1R had delayed onset of diabetes, indicating a role for these cytokines in disease development. TNFR1-deficient mice had an increased number of CD4(+) CD25(+) FoxP3(+) regulatory T cells with enhanced suppressive capacity. IL-1 was produced at higher levels in NOD mice and resulted in dilution of suppressor function of CD4(+) CD25(+) FoxP3(+) regulatory T cells. Our data suggest that blocking inflammatory cytokines may increase the capacity of the immune system to suppress type 1 diabetes through regulatory T cells.

  19. Potential role of stem cell therapy in type 1 diabetes mellitus.

    PubMed

    Couri, Carlos Eduardo Barra; Voltarelli, Júlio César

    2008-03-01

    Type 1 diabetes mellitus is the result of the autoimmune response against pancreatic beta-cell(s). At the time of clinical diagnosis near 70% of beta-cell mass is been destroyed as a consequence of the auto-destruction that begins months or even years before the clinical diagnosis. Although marked reduction of chronic complications was seen after development and progression of insulin therapy over the years for type 1 diabetic population, associated risks of chronic end-organ damage and hypoglycemia still remain. Besides tight glucose control, beta-cell mass preservation and/or increase are known to be other important targets in management of type 1 diabetes as long as it reduces chronic microvascular complications in the eyes, kidneys and nerves. Moreover, the larger the beta-cell mass, the lower the incidence of hypoglycemic events. In this article, we discuss some insights about beta-cell regeneration, the importance of regulation of the autoimmune process and what is being employed in human type 1 diabetes in regard to stem cell repertoire to promote regeneration and/or preservation of beta-cell mass.

  20. AMPA-type glutamate receptor subunits are expressed in the avian cochlear hair cells and ganglion cells.

    PubMed

    Reng, D; Hack, I; Müller, M; Smolders, J W

    1999-07-13

    The cellular localization of AMPA-type glutamate receptor subunits was examined in the pigeon inner ear using subunit specific polyclonal antibodies (GluR1-4). In the auditory ganglion cell bodies immunoreactivity for the subunits GluR2/3 and GluR4, but not for GluR1 was detected. The hair cells showed diffuse immunoreactivity for GluR4. Additionally, immunostaining for the subunits GluR2/3 and GluR4 was present below the hair cells. These results indicate that the AMPA type glutamate receptors play a role in neurotransmission at the hair cell afferent synapse in the avian auditory system.

  1. Differential mechanisms of memory CD8 T cell maintenance by individual myeloid cell types

    PubMed Central

    Frasca, Loredana; Stonier, Spencer W.; Overwijk, Willem W.; Schluns, Kimberly S.

    2010-01-01

    This study tested the hypothesis that individual myeloid subsets have a differential ability to maintain memory CD8 T cells via IL-15. Although DCs support IL-15-mediated homeostasis of memory CD8 T cells in vivo, whether various DC subsets and other myeloid cells similarly mediate homeostasis is unknown. Therefore, we studied the ability of different myeloid cells to maintain memory CD8 T cells in vitro. Using an in vitro cocoulture system that recapitulated known roles of DCs and IL-15 on memory CD8 T cells, all in vitro-derived or ex vivo-isolated DCs maintained CD8 T cells better than rIL-15 alone, and FLT-3L-DCs are the most efficient compared with GM-DCs, BM-derived macrophages, or freshly isolated DCs. Although FLT-3L-DCs were the least effective at inducing CD8 T cell proliferation, FLT-3L-DCs promoted better CD8 T cell survival and increased Bcl-2 and MCL-2 expression in CD8 T cells. T cell maintenance correlated only partially with DC expression of IL-15Rα and IL-15, suggesting that DCs provided additional support signals. Indeed, in the absence of IL-15 signals, CD70/CD27 further supported CD8 T cell maintenance. IFN-α enhanced CD70 expression by DCs, resulting in increased proliferation of CD8 T cells. Overall, this study supports our hypothesis by demonstrating that specific DC subtypes had a greater capacity to support memory CD8 T cell maintenance and did so through different mechanisms. Furthermore, this study shows that IL-15 trans-presentation can work in conjunction with other signals, such as CD70/CD27 interactions, to mediate CD8 T cell homeostasis efficiently. PMID:20354106

  2. Laser Microdissection Unravels Cell-Type-Specific Transcription in Arbuscular Mycorrhizal Roots, Including CAAT-Box Transcription Factor Gene Expression Correlating with Fungal Contact and Spread1[W

    PubMed Central

    Hogekamp, Claudia; Arndt, Damaris; Pereira, Patrícia A.; Becker, Jörg D.; Hohnjec, Natalija; Küster, Helge

    2011-01-01

    Arbuscular mycorrhizae (AM) are the most widespread symbioses on Earth, promoting nutrient supply of most terrestrial plant species. To unravel gene expression in defined stages of Medicago truncatula root colonization by AM fungi, we here combined genome-wide transcriptome profiling based on whole mycorrhizal roots with real-time reverse transcription-PCR experiments that relied on characteristic cell types obtained via laser microdissection. Our genome-wide approach delivered a core set of 512 genes significantly activated by the two mycorrhizal fungi Glomus intraradices and Glomus mossae. Focusing on 62 of these genes being related to membrane transport, signaling, and transcriptional regulation, we distinguished whether they are activated in arbuscule-containing or the neighboring cortical cells harboring fungal hyphae. In addition, cortical cells from nonmycorrhizal roots served as a reference for gene expression under noncolonized conditions. Our analysis identified 25 novel arbuscule-specific genes and 37 genes expressed both in the arbuscule-containing and the adjacent cortical cells colonized by fungal hyphae. Among the AM-induced genes specifying transcriptional regulators were two members encoding CAAT-box binding transcription factors (CBFs), designated MtCbf1 and MtCbf2. Promoter analyses demonstrated that both genes were already activated by the first physical contact between the symbionts. Subsequently, and corresponding to our cell-type expression patterns, they were progressively up-regulated in those cortical areas colonized by fungal hyphae, including the arbuscule-containing cells. The encoded CBFs thus represent excellent candidates for regulators that mediate a sequential reprogramming of root tissues during the establishment of an AM symbiosis. PMID:22034628

  3. Tolerization of a type I allergic immune response through transplantation of genetically modified hematopoietic stem cells.

    PubMed

    Baranyi, Ulrike; Linhart, Birgit; Pilat, Nina; Gattringer, Martina; Bagley, Jessamyn; Muehlbacher, Ferdinand; Iacomini, John; Valenta, Rudolf; Wekerle, Thomas

    2008-06-15

    Allergy represents a hypersensitivity disease that affects >25% of the population in industrialized countries. The underlying type I allergic immune reaction occurs in predisposed atopic individuals in response to otherwise harmless Ags (i.e., allergens) and is characterized by the production of allergen-specific IgE, an allergen-specific T cell response, and the release of biologically active mediators such as histamine from mast cells and basophils. Regimens permanently tolerizing an allergic immune response still need to be developed. We therefore retrovirally transduced murine hematopoietic stem cells to express the major grass pollen allergen Phl p 5 on their cell membrane. Transplantation of these genetically modified hematopoietic stem cells led to durable multilineage molecular chimerism and permanent immunological tolerance toward the introduced allergen at the B cell, T cell, and effector cell levels. Notably, Phl p 5-specific serum IgE and IgG remained undetectable, and T cell nonresponsiveness persisted throughout follow-up (40 wk). Besides, mediator release was specifically absent in in vitro and in vivo assays. B cell, T cell, and effector cell responses to an unrelated control allergen (Bet v 1) were unperturbed, demonstrating specificity of this tolerance protocol. We thus describe a novel cell-based strategy for the prevention of allergy.

  4. Small Molecule Screening in Human Induced Pluripotent Stem Cell-derived Terminal Cell Types*

    PubMed Central

    Engle, Sandra J.; Vincent, Fabien

    2014-01-01

    A need for better clinical outcomes has heightened interest in the use of physiologically relevant human cells in the drug discovery process. Patient-specific human induced pluripotent stem cells may offer a relevant, robust, scalable, and cost-effective model of human disease physiology. Small molecule high throughput screening in human induced pluripotent stem cell-derived cells with the intent of identifying novel therapeutic compounds is starting to influence the drug discovery process; however, the use of these cells presents many high throughput screening development challenges. This technology has the potential to transform the way drug discovery is performed. PMID:24362033

  5. Laser-assisted Microdissection (LAM) as a Tool for Transcriptional Profiling of Individual Cell Types.

    PubMed

    Florez Rueda, Ana Marcela; Grossniklaus, Ueli; Schmidt, Anja

    2016-01-01

    The understanding of developmental processes at the molecular level requires insights into transcriptional regulation, and thus the transcriptome, at the level of individual cell types. While the methods described here are generally applicable to a wide range of species and cell types, our research focuses on plant reproduction. Plant cultivation and seed production is of crucial importance for human and animal nutrition. A detailed understanding of the regulatory networks that govern the formation of the reproductive lineage (germline) and ultimately of seeds is a precondition for the targeted manipulation of plant reproduction. In particular, the engineering of apomixis (asexual reproduction through seeds) into crop plants promises great improvements, as it leads to the formation of clonal seeds that are genetically identical to the mother plant. Consequently, the cell types of the female germline are of major importance for the understanding and engineering of apomixis. However, as the corresponding cells are deeply embedded within the floral tissues, they are very difficult to access for experimental analyses, including cell-type specific transcriptomics. To overcome this limitation, sections of individual cells can be isolated by laser-assisted microdissection (LAM). While LAM in combination with transcriptional profiling allows the identification of genes and pathways active in any cell type with high specificity, establishing a suitable protocol can be challenging. Specifically, the quality of RNA obtained after LAM can be compromised, especially when small, single cells are targeted. To circumvent this problem, we have established a workflow for LAM that reproducibly results in high RNA quality that is well suitable for transcriptomics, as exemplified here by the isolation of cells of the female germline in apomictic Boechera. In this protocol, procedures are described for tissue preparation and LAM, also with regard to RNA extraction and quality control

  6. Cell- and stimulus type-specific intracellular free Ca2+ signals in Arabidopsis.

    PubMed

    Martí, María C; Stancombe, Matthew A; Webb, Alex A R

    2013-10-01

    Appropriate stimulus-response coupling requires that each signal induces a characteristic response, distinct from that induced by other signals, and that there is the potential for individual signals to initiate different downstream responses dependent on cell type. How such specificity is encoded in plant signaling is not known. One possibility is that information is encoded in signal transduction pathways to ensure stimulus- and cell type-specific responses. The calcium ion acts as a second messenger in response to mechanical stimulation, hydrogen peroxide, NaCl, and cold in plants and also in circadian timing. We use GAL4 transactivation of aequorin in enhancer trap lines of Arabidopsis (Arabidopsis thaliana) to test the hypothesis that stimulus- and cell-specific information can be encoded in the pattern of dynamic alterations in the concentration of intracellular free Ca(2+) ([Ca(2+)]i). We demonstrate that mechanically induced increases in [Ca(2+)]i are largely restricted to the epidermal pavement cells of leaves, that NaCl induces oscillatory [Ca(2+)]i signals in spongy mesophyll and vascular bundle cells, but not other cell types, and detect circadian rhythms of [Ca(2+)]i only in the spongy mesophyll. We demonstrate stimulus-specific [Ca(2+)]i dynamics in response to touch, cold, and hydrogen peroxide, which in the case of the latter two signals are common to all cell types tested. GAL4 transactivation of aequorin in specific leaf cell types has allowed us to bypass the technical limitations associated with fluorescent Ca(2+) reporter dyes in chlorophyll-containing tissues to identify the cell- and stimulus-specific complexity of [Ca(2+)]i dynamics in leaves of Arabidopsis and to determine from which tissues stress- and circadian-regulated [Ca(2+)]i signals arise.

  7. Plasmolysis and recovery of different cell types in cryoprotected shoot tips of Mentha X piperita.

    PubMed

    Volk, Gayle M; Caspersen, Ann M

    2007-01-01

    Successful cryopreservation of plant shoot tips is dependent upon effective desiccation through osmotic or physical processes. Microscopy techniques were used to determine the extent of cellular damage and plasmolysis that occurs in peppermint (Mentha x piperita) shoot tips during the process of cryopreservation, using the cryoprotectant plant vitrification solution 2 (PVS2) (30% glycerol, 15% dimethyl sulfoxide, 15% ethylene glycol, 0.4 M sucrose) prior to liquid-nitrogen exposure. The meristem cells were the smallest and least plasmolyzed cell type of the shoot tips, while the large, older leaf and lower cortex cells were the most damaged. When treated with cryoprotectant solutions, meristem cells exhibited concave plasmolysis, suggesting that this cell type has a highly viscous protoplasm, and protoplasts have many cell wall attachment sites. Shoot tip cells were most severely plasmolyzed after PVS2 treatment, liquid-nitrogen exposure, and warming in 1.2 M sucrose. Successful recovery may be dependent upon surviving the plasmolytic conditions induced by warming and diluting treated shoot tips in 1.2 M sucrose solutions. In peppermint shoot tips, clumps of young meristem or young leaf cells survive the cryopreservation process and regenerate plants containing many shoots. Cryoprotective treatments that favor survival of small, meristematic cells and young leaf cells are most likely to produce high survival rates after liquid-nitrogen exposure.

  8. Therapeutic potential of umbilical cord blood cells for type 1 diabetes mellitus.

    PubMed

    He, Binbin; Li, Xia; Yu, Haibo; Zhou, Zhiguang

    2015-11-01

    Type 1 diabetes mellitus (T1DM) is a chronic disorder that results from autoimmune-mediated destruction of pancreatic islet β-cells. However, to date, no conventional intervention has successfully treated the disease. The optimal therapeutic method for T1DM should effectively control the autoimmunity, restore immune homeostasis, preserve residual β-cells, reverse β-cell destruction, and protect the regenerated insulin-producing cells against re-attack. Umbilical cord blood is rich in regulatory T (T(reg)) cells and multiple types of stem cells that exhibit immunomodulating potential and hold promise in their ability to restore peripheral tolerance towards pancreatic islet β-cells through remodeling of immune responses and suppression of autoreactive T cells. Recently, reinfusion of autologous umbilical cord blood or immune cells from cord blood has been proposed as a novel therapy for T1DM, with the advantages of no risk to the donors, minimal ethical concerns, a low incidence of graft-versus-host disease and easy accessibility. In this review, we revisit the role of autologous umbilical cord blood or immune cells from cord blood-based applications for the treatment of T1DM. PMID:25799887

  9. Therapeutic potential of umbilical cord blood cells for type 1 diabetes mellitus.

    PubMed

    He, Binbin; Li, Xia; Yu, Haibo; Zhou, Zhiguang

    2015-11-01

    Type 1 diabetes mellitus (T1DM) is a chronic disorder that results from autoimmune-mediated destruction of pancreatic islet β-cells. However, to date, no conventional intervention has successfully treated the disease. The optimal therapeutic method for T1DM should effectively control the autoimmunity, restore immune homeostasis, preserve residual β-cells, reverse β-cell destruction, and protect the regenerated insulin-producing cells against re-attack. Umbilical cord blood is rich in regulatory T (T(reg)) cells and multiple types of stem cells that exhibit immunomodulating potential and hold promise in their ability to restore peripheral tolerance towards pancreatic islet β-cells through remodeling of immune responses and suppression of autoreactive T cells. Recently, reinfusion of autologous umbilical cord blood or immune cells from cord blood has been proposed as a novel therapy for T1DM, with the advantages of no risk to the donors, minimal ethical concerns, a low incidence of graft-versus-host disease and easy accessibility. In this review, we revisit the role of autologous umbilical cord blood or immune cells from cord blood-based applications for the treatment of T1DM.

  10. The importance of detailed epigenomic profiling of different cell types within organs.

    PubMed

    Stueve, Theresa Ryan; Marconett, Crystal N; Zhou, Beiyun; Borok, Zea; Laird-Offringa, Ite A

    2016-06-01

    The human body consists of hundreds of kinds of cells specified from a single genome overlaid with cell type-specific epigenetic information. Comprehensively profiling the body's distinct epigenetic landscapes will allow researchers to verify cell types used in regenerative medicine and to determine the epigenetic effects of disease, environmental exposures and genetic variation. Key marks/factors that should be investigated include regions of nucleosome-free DNA accessible to regulatory factors, histone marks defining active enhancers and promoters, DNA methylation levels, regulatory RNAs, and factors controlling the three-dimensional conformation of the genome. Here we use the lung to illustrate the importance of investigating an organ's purified cell epigenomes, and outline the challenges and promise of realizing a comprehensive catalog of primary cell epigenomes. PMID:27305639

  11. Quantification of HIV-based lentiviral vectors: influence of several cell type parameters on vector infectivity.

    PubMed

    Gay, Virginie; Moreau, Karen; Hong, Saw-See; Ronfort, Corinne

    2012-02-01

    A human immunodeficiency virus type (HIV-1)-based lentiviral vector pseudotyped with the vesicular stomatitis virus envelope glycoprotein and encoding the GFP reporter gene was used to evaluate different methods of lentiviral vector titration. GFP expression, viral DNA quantification and the efficiency of vector DNA integration were assayed after infection of conventional HIV-1-permissive cell lines and human primary adult fibroblasts with the vector. We found that vector titers based on GFP expression determined by flow cytometry may vary by more than 50-fold depending on the cell type and the promoter-cell combination used. Interestingly, we observed that the viral integration process in primary HDFa cells was significantly more efficient compared to that in SupT1 or 293T cells. We propose that determination of the amount of integrated viral DNA by quantitative PCR be used in combination with the reporter gene expression assay.

  12. Ultra-Deep Bisulfite Sequencing to Detect Specific DNA Methylation Patterns of Minor Cell Types in Heterogeneous Cell Populations: An Example of the Pituitary Tissue.

    PubMed

    Arai, Yoshikazu; Fukukawa, Hisho; Atozi, Takanori; Matsumoto, Shoma; Hanazono, Yutaka; Nagashima, Hiroshi; Ohgane, Jun

    2016-01-01

    DNA methylation is an epigenetic modification important for cell fate determination and cell type-specific gene expression. Transcriptional regulatory regions of the mammalian genome contain a large number of tissue/cell type-dependent differentially methylated regions (T-DMRs) with DNA methylation patterns crucial for transcription of the corresponding genes. In general, tissues consist of multiple cell types in various proportions, making it difficult to detect T-DMRs of minor cell types in tissues. The present study attempts to detect T-DMRs of minor cell types in tissues by ultra-deep bisulfite sequencing of cell type-restricted genes and to assume proportions of minor cell types based on DNA methylation patterns of sequenced reads. For this purpose, we focused on transcriptionally active hypomethylated alleles (Hypo-alleles), which can be recognized by the high ratio of unmethylated CpGs in each sequenced read (allele). The pituitary gland contains multiple cell types including five hormone-expressing cell types and stem/progenitor cells, each of which is a minor cell type in the pituitary tissue. By ultra-deep sequencing of more than 100 reads for detection of Hypo-alleles in pituitary cell type-specific genes, we identified T-DMRs specific to hormone-expressing cells and stem/progenitor cells and used them to estimate the proportions of each cell type based on the Hypo-allele ratio in pituitary tissue. Therefore, introduction of the novel Hypo-allele concept enabled us to detect T-DMRs of minor cell types with estimation of their proportions in the tissue by ultra-deep bisulfite sequencing.

  13. Replication of bovine herpesvirus type 4 in human cells in vitro.

    PubMed

    Egyed, L

    1998-07-01

    A reference strain (Movár 33/63) of bovine herpesvirus type 4 (BHV-4) was inoculated into 14 different human cell lines and five primary cell cultures representing various human tissues. BHV-4 replicated in two embryonic lung cell lines, MRC-5 and Wistar-38, and in a giant-cell glioblastoma cell culture. Cytopathic effect and intranuclear inclusion bodies were observed in these cells. PCR detected a 10,000-times-higher level of BHV-4 DNA. Titration of the supernatant indicated a 100-fold increase of infectious particles. Since this is the first bovine (human herpesvirus 8 and Epstein-Barr virus related) herpesvirus which replicates on human cells in vitro, the danger of possible human BHV-4 infection should not be ignored.

  14. Surface-Engineered Viral Vectors for Selective and Cell Type-Specific Gene Delivery.

    PubMed

    Buchholz, Christian J; Friedel, Thorsten; Büning, Hildegard

    2015-12-01

    Recent progress in gene transfer technology enables the delivery of genes precisely to the application-relevant cell type ex vivo on cultivated primary cells or in vivo on local or systemic administration. Gene vectors based on lentiviruses or adeno-associated viruses can be engineered such that they use a cell surface marker of choice for cell entry instead of their natural receptors. Binding to the surface marker is mediated by a targeting ligand displayed on the vector particle surface, which can be a peptide, single-chain antibody, or designed ankyrin repeat protein. Examples include vectors that deliver genes to specialized endothelial cells or lymphocytes, tumor cells, or particular cells of the nervous system with potential applications in gene function studies and molecular medicine.

  15. Cell-based interventions to halt autoimmunity in type 1 diabetes mellitus

    PubMed Central

    Barcala Tabarrozzi, A E; Castro, C N; Dewey, R A; Sogayar, M C; Labriola, L; Perone, M J

    2013-01-01

    Type 1 diabetes mellitus (T1DM) results from death of insulin-secreting β cells mediated by self-immune cells, and the consequent inability of the body to maintain insulin levels for appropriate glucose homeostasis. Probably initiated by environmental factors, this disease takes place in genetically predisposed individuals. Given the autoimmune nature of T1DM, therapeutics targeting immune cells involved in disease progress have been explored over the last decade. Several high-cost trials have been attempted to prevent and/or reverse T1DM. Although a definitive solution to cure T1DM is not yet available, a large amount of information about its nature and development has contributed greatly to both the improvement of patient's health care and design of new treatments. In this study, we discuss the role of different types of immune cells involved in T1DM pathogenesis and their therapeutic potential as targets and/or modified tools to treat patients. Recently, encouraging results and new approaches to sustain remnant β cell mass and to increase β cell proliferation by different cell-based means have emerged. Results coming from ongoing clinical trials employing cell therapy designed to arrest T1DM will probably proliferate in the next few years. Strategies under consideration include infusion of several types of stem cells, dendritic cells and regulatory T cells, either manipulated genetically ex vivo or non-manipulated. Their use in combination approaches is another therapeutic alternative. Cell-based interventions, without undesirable side effects, directed to block the uncontrollable autoimmune response may become a clinical reality in the next few years for the treatment of patients with T1DM. PMID:23286940

  16. Cell-based interventions to halt autoimmunity in type 1 diabet