Polyploidization in liver tissue.

PubMed

Gentric, Géraldine; Desdouets, Chantal

2014-02-01

Polyploidy (alias whole genome amplification) refers to organisms containing more than two basic sets of chromosomes. Polyploidy was first observed in plants more than a century ago, and it is known that such processes occur in many eukaryotes under a variety of circumstances. In mammals, the development of polyploid cells can contribute to tissue differentiation and, therefore, possibly a gain of function; alternately, it can be associated with development of disease, such as cancer. Polyploidy can occur because of cell fusion or abnormal cell division (endoreplication, mitotic slippage, or cytokinesis failure). Polyploidy is a common characteristic of the mammalian liver. Polyploidization occurs mainly during liver development, but also in adults with increasing age or because of cellular stress (eg, surgical resection, toxic exposure, or viral infections). This review will explore the mechanisms that lead to the development of polyploid cells, our current state of understanding of how polyploidization is regulated during liver growth, and its consequence on liver function. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Repair of DNA Damage Induced by the Cytidine Analog Zebularine Requires ATR and ATM in Arabidopsis[OPEN

PubMed Central

Liu, Chun-Hsin; Finke, Andreas; Díaz, Mariana; Rozhon, Wilfried; Poppenberger, Brigitte; Baubec, Tuncay; Pecinka, Ales

2015-01-01

DNA damage repair is an essential cellular mechanism that maintains genome stability. Here, we show that the nonmethylable cytidine analog zebularine induces a DNA damage response in Arabidopsis thaliana, independent of changes in DNA methylation. In contrast to genotoxic agents that induce damage in a cell cycle stage-independent manner, zebularine induces damage specifically during strand synthesis in DNA replication. The signaling of this damage is mediated by additive activity of ATAXIA TELANGIECTASIA MUTATED AND RAD3-RELATED and ATAXIA TELANGIECTASIA MUTATED kinases, which cause postreplicative cell cycle arrest and increased endoreplication. The repair requires a functional STRUCTURAL MAINTENANCE OF CHROMOSOMES5 (SMC5)-SMC6 complex and is accomplished predominantly by synthesis-dependent strand-annealing homologous recombination. Here, we provide insight into the response mechanism for coping with the genotoxic effects of zebularine and identify several components of the zebularine-induced DNA damage repair pathway. PMID:26023162

Regulating DNA Replication in Plants

PubMed Central

Sanchez, Maria de la Paz; Costas, Celina; Sequeira-Mendes, Joana; Gutierrez, Crisanto

2012-01-01

Chromosomal DNA replication in plants has requirements and constraints similar to those in other eukaryotes. However, some aspects are plant-specific. Studies of DNA replication control in plants, which have unique developmental strategies, can offer unparalleled opportunities of comparing regulatory processes with yeast and, particularly, metazoa to identify common trends and basic rules. In addition to the comparative molecular and biochemical studies, genomic studies in plants that started with Arabidopsis thaliana in the year 2000 have now expanded to several dozens of species. This, together with the applicability of genomic approaches and the availability of a large collection of mutants, underscores the enormous potential to study DNA replication control in a whole developing organism. Recent advances in this field with particular focus on the DNA replication proteins, the nature of replication origins and their epigenetic landscape, and the control of endoreplication will be reviewed. PMID:23209151

Entrainment of the Mammalian Cell Cycle by the Circadian Clock: Modeling Two Coupled Cellular Rhythms

PubMed Central

Gérard, Claude; Goldbeter, Albert

2012-01-01

The cell division cycle and the circadian clock represent two major cellular rhythms. These two periodic processes are coupled in multiple ways, given that several molecular components of the cell cycle network are controlled in a circadian manner. For example, in the network of cyclin-dependent kinases (Cdks) that governs progression along the successive phases of the cell cycle, the synthesis of the kinase Wee1, which inhibits the G2/M transition, is enhanced by the complex CLOCK-BMAL1 that plays a central role in the circadian clock network. Another component of the latter network, REV-ERBα, inhibits the synthesis of the Cdk inhibitor p21. Moreover, the synthesis of the oncogene c-Myc, which promotes G1 cyclin synthesis, is repressed by CLOCK-BMAL1. Using detailed computational models for the two networks we investigate the conditions in which the mammalian cell cycle can be entrained by the circadian clock. We show that the cell cycle can be brought to oscillate at a period of 24 h or 48 h when its autonomous period prior to coupling is in an appropriate range. The model indicates that the combination of multiple modes of coupling does not necessarily facilitate entrainment of the cell cycle by the circadian clock. Entrainment can also occur as a result of circadian variations in the level of a growth factor controlling entry into G1. Outside the range of entrainment, the coupling to the circadian clock may lead to disconnected oscillations in the cell cycle and the circadian system, or to complex oscillatory dynamics of the cell cycle in the form of endoreplication, complex periodic oscillations or chaos. The model predicts that the transition from entrainment to 24 h or 48 h might occur when the strength of coupling to the circadian clock or the level of growth factor decrease below critical values. PMID:22693436

Regulation of CTP Synthase Filament Formation During DNA Endoreplication in Drosophila.

PubMed

Wang, Pei-Yu; Lin, Wei-Cheng; Tsai, Yi-Cheng; Cheng, Mei-Ling; Lin, Yu-Hung; Tseng, Shu-Heng; Chakraborty, Archan; Pai, Li-Mei

2015-12-01

CTP synthase (CTPsyn) plays an essential role in DNA, RNA, and lipid synthesis. Recent studies in bacteria, yeast, and Drosophila all reveal a polymeric CTPsyn structure, which dynamically regulates its enzymatic activity. However, the molecular mechanism underlying the formation of CTPsyn polymers is not completely understood. In this study, we found that reversible ubiquitination regulates the dynamic assembly of the filamentous structures of Drosophila CTPsyn. We further determined that the proto-oncogene Cbl, an E3 ubiquitin ligase, controls CTPsyn filament formation in endocycles. While the E3 ligase activity of Cbl is required for CTPsyn filament formation, Cbl does not affect the protein levels of CTPsyn. It remains unclear whether the regulation of CTPsyn filaments by Cbl is through direct ubiquitination of CTPsyn. In the absence of Cbl or with knockdown of CTPsyn, the progression of the endocycle-associated S phase was impaired. Furthermore, overexpression of wild-type, but not enzymatically inactive CTPsyn, rescued the endocycle defect in Cbl mutant cells. Together, these results suggest that Cbl influences the nucleotide pool balance and controls CTPsyn filament formation in endocycles. This study links Cbl-mediated ubiquitination to the polymerization of a metabolic enzyme and reveals a role for Cbl in endocycles during Drosophila development. Copyright © 2015 by the Genetics Society of America.

Cell fusion in the liver, revisited

PubMed Central

Lizier, Michela; Castelli, Alessandra; Montagna, Cristina; Lucchini, Franco; Vezzoni, Paolo; Faggioli, Francesca

2018-01-01

There is wide agreement that cell fusion is a physiological process in cells in mammalian bone, muscle and placenta. In other organs, such as the cerebellum, cell fusion is controversial. The liver contains a considerable number of polyploid cells: They are commonly believed to originate by genome endoreplication, although the contribution of cell fusion to polyploidization has not been excluded. Here, we address the topic of cell fusion in the liver from a historical point of view. We discuss experimental evidence clearly supporting the hypothesis that cell fusion occurs in the liver, specifically when bone marrow cells were injected into mice and shown to rescue genetic hepatic degenerative defects. Those experiments-carried out in the latter half of the last century-were initially interpreted to show “transdifferentiation”, but are now believed to demonstrate fusion between donor macrophages and host hepatocytes, raising the possibility that physiologically polyploid cells, such as hepatocytes, could originate, at least partially, through homotypic cell fusion. In support of the homotypic cell fusion hypothesis, we present new data generated using a chimera-based model, a much simpler model than those previously used. Cell fusion as a road to polyploidization in the liver has not been extensively investigated, and its contribution to a variety of conditions, such as viral infections, carcinogenesis and aging, remains unclear. PMID:29527257

Characterisation of cell cycle arrest and terminal differentiation in a maximally proliferative human epithelial tissue: Lessons from the human hair follicle matrix.

PubMed

Purba, Talveen S; Brunken, Lars; Peake, Michael; Shahmalak, Asim; Chaves, Asuncion; Poblet, Enrique; Ceballos, Laura; Gandarillas, Alberto; Paus, Ralf

2017-09-01

Human hair follicle (HF) growth and hair shaft formation require terminal differentiation-associated cell cycle arrest of highly proliferative matrix keratinocytes. However, the regulation of this complex event remains unknown. CIP/KIP family member proteins (p21 CIP1 , p27 KIP1 and p57 KIP2 ) regulate cell cycle progression/arrest, endoreplication, differentiation and apoptosis. Since they have not yet been adequately characterized in the human HF, we asked whether and where CIP/KIP proteins localise in the human hair matrix and pre-cortex in relation to cell cycle activity and HF-specific epithelial cell differentiation that is marked by keratin 85 (K85) protein expression. K85 expression coincided with loss or reduction in cell cycle activity markers, including in situ DNA synthesis (EdU incorporation), Ki-67, phospho-histone H3 and cyclins A and B1, affirming a post-mitotic state of pre-cortical HF keratinocytes. Expression of CIP/KIP proteins was found abundantly within the proliferative hair matrix, concomitant with a role in cell cycle checkpoint control. p21 CIP1 , p27 KIP1 and cyclin E persisted within post-mitotic keratinocytes of the pre-cortex, whereas p57 KIP2 protein decreased but became nuclear. These data imply a supportive role for CIP/KIP proteins in maintaining proliferative arrest, differentiation and anti-apoptotic pathways, promoting continuous hair bulb growth and hair shaft formation in anagen VI. Moreover, post-mitotic hair matrix regions contained cells with enlarged nuclei, and DNA in situ hybridisation showed cells that were >2N in the pre-cortex. This suggests that CIP/KIP proteins might counterbalance cyclin E to control further rounds of DNA replication in a cell population that has a propensity to become tetraploid. These data shed new light on the in situ-biography of human hair matrix keratinocytes on their path of active cell cycling, arrest and terminal differentiation, and showcase the human HF as an excellent, clinically relevant model system for cell cycle physiology research of human epithelial cells within their natural tissue habitat. Crown Copyright © 2017. Published by Elsevier GmbH. All rights reserved.

Polyploidization without mitosis improves in vivo liver transduction with lentiviral vectors.

PubMed

Pichard, Virginie; Couton, Dominique; Desdouets, Chantal; Ferry, Nicolas

2013-02-01

Lentiviral vectors are efficient gene delivery vehicles for therapeutic and research applications. In contrast to oncoretroviral vectors, they are able to infect most nonproliferating cells. In the liver, induction of cell proliferation dramatically improved hepatocyte transduction using all types of retroviral vectors. However, the precise relationship between hepatocyte division and transduction efficiency has not been determined yet. Here we compared gene transfer efficiency in the liver after in vivo injection of recombinant lentiviral or Moloney murine leukemia viral (MoMuLV) vectors in hepatectomized rats treated or not with retrorsine, an alkaloid that blocks hepatocyte division and induces megalocytosis. Partial hepatectomy alone resulted in a similar increase in hepatocyte transduction using either vector. In retrorsine-treated and partially hepatectomized rats, transduction with MoMuLV vectors dropped dramatically. In contrast, we observed that retrorsine treatment combined with partial hepatectomy increased lentiviral transduction to higher levels than hepatectomy alone. Analysis of nuclear ploidy in single cells showed that a high level of transduction was associated with polyploidization. In conclusion, endoreplication could be exploited to improve the efficiency of liver-directed lentiviral gene therapy.

Polyploidization Without Mitosis Improves In Vivo Liver Transduction With Lentiviral Vectors

PubMed Central

Couton, Dominique; Desdouets, Chantal; Ferry, Nicolas

2013-01-01

Abstract Lentiviral vectors are efficient gene delivery vehicles for therapeutic and research applications. In contrast to oncoretroviral vectors, they are able to infect most nonproliferating cells. In the liver, induction of cell proliferation dramatically improved hepatocyte transduction using all types of retroviral vectors. However, the precise relationship between hepatocyte division and transduction efficiency has not been determined yet. Here we compared gene transfer efficiency in the liver after in vivo injection of recombinant lentiviral or Moloney murine leukemia viral (MoMuLV) vectors in hepatectomized rats treated or not with retrorsine, an alkaloid that blocks hepatocyte division and induces megalocytosis. Partial hepatectomy alone resulted in a similar increase in hepatocyte transduction using either vector. In retrorsine-treated and partially hepatectomized rats, transduction with MoMuLV vectors dropped dramatically. In contrast, we observed that retrorsine treatment combined with partial hepatectomy increased lentiviral transduction to higher levels than hepatectomy alone. Analysis of nuclear ploidy in single cells showed that a high level of transduction was associated with polyploidization. In conclusion, endoreplication could be exploited to improve the efficiency of liver-directed lentiviral gene therapy. PMID:23249390

A mitosis block links active cell cycle with human epidermal differentiation and results in endoreplication.

PubMed

Zanet, Jennifer; Freije, Ana; Ruiz, María; Coulon, Vincent; Sanz, J Ramón; Chiesa, Jean; Gandarillas, Alberto

2010-12-20

How human self-renewal tissues co-ordinate proliferation with differentiation is unclear. Human epidermis undergoes continuous cell growth and differentiation and is permanently exposed to mutagenic hazard. Keratinocytes are thought to arrest cell growth and cell cycle prior to terminal differentiation. However, a growing body of evidence does not satisfy this model. For instance, it does not explain how skin maintains tissue structure in hyperproliferative benign lesions. We have developed and applied novel cell cycle techniques to human skin in situ and determined the dynamics of key cell cycle regulators of DNA replication or mitosis, such as cyclins E, A and B, or members of the anaphase promoting complex pathway: cdc14A, Ndc80/Hec1 and Aurora kinase B. The results show that actively cycling keratinocytes initiate terminal differentiation, arrest in mitosis, continue DNA replication in a special G2/M state, and become polyploid by mitotic slippage. They unambiguously demonstrate that cell cycle progression coexists with terminal differentiation, thus explaining how differentiating cells increase in size. Epidermal differentiating cells arrest in mitosis and a genotoxic-induced mitosis block rapidly pushes epidermal basal cells into differentiation and polyploidy. These observations unravel a novel mitosis-differentiation link that provides new insight into skin homeostasis and cancer. It might constitute a self-defence mechanism against oncogenic alterations such as Myc deregulation.

A Mitosis Block Links Active Cell Cycle with Human Epidermal Differentiation and Results in Endoreplication

PubMed Central

Zanet, Jennifer; Freije, Ana; Ruiz, María; Coulon, Vincent; Sanz, J. Ramón; Chiesa, Jean; Gandarillas, Alberto

2010-01-01

How human self-renewal tissues co-ordinate proliferation with differentiation is unclear. Human epidermis undergoes continuous cell growth and differentiation and is permanently exposed to mutagenic hazard. Keratinocytes are thought to arrest cell growth and cell cycle prior to terminal differentiation. However, a growing body of evidence does not satisfy this model. For instance, it does not explain how skin maintains tissue structure in hyperproliferative benign lesions. We have developed and applied novel cell cycle techniques to human skin in situ and determined the dynamics of key cell cycle regulators of DNA replication or mitosis, such as cyclins E, A and B, or members of the anaphase promoting complex pathway: cdc14A, Ndc80/Hec1 and Aurora kinase B. The results show that actively cycling keratinocytes initiate terminal differentiation, arrest in mitosis, continue DNA replication in a special G2/M state, and become polyploid by mitotic slippage. They unambiguously demonstrate that cell cycle progression coexists with terminal differentiation, thus explaining how differentiating cells increase in size. Epidermal differentiating cells arrest in mitosis and a genotoxic-induced mitosis block rapidly pushes epidermal basal cells into differentiation and polyploidy. These observations unravel a novel mitosis-differentiation link that provides new insight into skin homeostasis and cancer. It might constitute a self-defence mechanism against oncogenic alterations such as Myc deregulation. PMID:21187932

Long Term Ex Vivo Culture and Live Imaging of Drosophila Larval Imaginal Discs.

PubMed

Tsao, Chia-Kang; Ku, Hui-Yu; Lee, Yuan-Ming; Huang, Yu-Fen; Sun, Yi Henry

Continuous imaging of live tissues provides clear temporal sequence of biological events. The Drosophila imaginal discs have been popular experimental subjects for the study of a wide variety of biological phenomena, but long term culture that allows normal development has not been satisfactory. Here we report a culture method that can sustain normal development for 18 hours and allows live imaging. The method is validated in multiple discs and for cell proliferation, differentiation and migration. However, it does not support disc growth and cannot support cell proliferation for more than 7 to 12 hr. We monitored the cellular behavior of retinal basal glia in the developing eye disc and found that distinct glia type has distinct properties of proliferation and migration. The live imaging provided direct proof that wrapping glia differentiated from existing glia after migrating to the anterior front, and unexpectedly found that they undergo endoreplication before wrapping axons, and their nuclei migrate up and down along the axons. UV-induced specific labeling of a single carpet glia also showed that the two carpet glia membrane do not overlap and suggests a tiling or repulsion mechanism between the two cells. These findings demonstrated the usefulness of an ex vivo culture method and live imaging.

Janus face-like effects of Aurora B inhibition: antitumoral mode of action versus induction of aneuploid progeny.

PubMed

Wiedemuth, Ralf; Klink, Barbara; Fujiwara, Mamoru; Schröck, Evelin; Tatsuka, Masaaki; Schackert, Gabriele; Temme, Achim

2016-10-01

The mitotic Aurora B kinase is overexpressed in tumors and various inhibitors for Aurora B are currently under clinical assessments. However, when considering Aurora B kinase inhibitors as anticancer drugs, their mode of action and the role of p53 status as a possible predictive factor for response still needs to be investigated. In this study, we analyzed the effects of selective Aurora B inhibition using AZD1152-HQPA/Barasertib (AZD1152) on HCT116 cells, U87-MG, corresponding isogenic p53-deficient cells and a primary glioblastoma cell line. AZD1152 treatment caused polyploidy and non-apoptotic cell death in all cell lines irrespective of p53 status and was accompanied by poly-merotelic kinetochore-microtubule attachments and DNA damage. In p53 wild-type cells a DNA damage response induced an inefficient pseudo-G1 cell cycle arrest, which was not able to halt ongoing endoreplication of cells. Of note, release of tumor cells from AZD1152 resulted in recovery of aneuploid progenies bearing numerical and structural chromosomal aberrations. Yet, AZD1152 treatment enhanced death receptor TRAIL-R2 levels in all tumor cell lines investigated. A concomitant increase of the activating natural killer (NK) cell ligand MIC A/B in p53-deficient cells and an induction of FAS/CD95 in cells containing p53 rendered AZD1152-treated cells more susceptible for NK-cell-mediated lysis. Our study mechanistically explains a p53-independent mode of action of a chemical Aurora B inhibitor and suggests a potential triggering of antitumoral immune responses, following polyploidization of tumor cells, which might constrain recovery of aneuploid tumor cells. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Drosophila Symplekin localizes dynamically to the histone locus body and tricellular junctions.

PubMed

Tatomer, Deirdre C; Rizzardi, Lindsay F; Curry, Kaitlin P; Witkowski, Alison M; Marzluff, William F; Duronio, Robert J

2014-01-01

The scaffolding protein Symplekin is part of multiple complexes involved in generating and modifying the 3' end of mRNAs, including cleavage-polyadenylation, histone pre-mRNA processing and cytoplasmic polyadenylation. To study these functions in vivo, we examined the localization of Symplekin during development and generated mutations of the Drosophila Symplekin gene. Mutations in Symplekin that reduce Symplekin protein levels alter the efficiency of both poly A(+) and histone mRNA 3' end formation resulting in lethality or sterility. Histone mRNA synthesis takes place at the histone locus body (HLB) and requires a complex composed of Symplekin and several polyadenylation factors that associates with the U7 snRNP. Symplekin is present in the HLB in the early embryo when Cyclin E/Cdk2 is active and histone genes are expressed and is absent from the HLB in cells that have exited the cell cycle. During oogenesis, Symplekin is preferentially localized to HLBs during S-phase in endoreduplicating follicle cells when histone mRNA is synthesized. After the completion of endoreplication, Symplekin accumulates in the cytoplasm, in addition to the nucleoplasm, and localizes to tricellular junctions of the follicle cell epithelium. This localization depends on the RNA binding protein ypsilon schachtel. CPSF-73 and a number of mRNAs are localized at this same site, suggesting that Symplekin participates in cytoplasmic polyadenylation at tricellular junctions.

Drug-Free Approach To Study the Unusual Cell Cycle of Giardia intestinalis

PubMed Central

Horlock-Roberts, Kathleen; Reaume, Chase; Dayer, Guillem; Ouellet, Christine; Cook, Nicholas

2017-01-01

ABSTRACT Giardia intestinalis is a protozoan parasite that causes giardiasis, a form of severe and infectious diarrhea. Despite the importance of the cell cycle in the control of proliferation and differentiation during a giardia infection, it has been difficult to study this process due to the absence of a synchronization procedure that would not induce cellular damage resulting in artifacts. We utilized counterflow centrifugal elutriation (CCE), a size-based separation technique, to successfully obtain fractions of giardia cultures enriched in G1, S, and G2. Unlike drug-induced synchronization of giardia cultures, CCE did not induce double-stranded DNA damage or endoreplication. We observed increases in the appearance and size of the median body in the cells from elutriation fractions corresponding to the progression of the cell cycle from early G1 to late G2. Consequently, CCE could be used to examine the dynamics of the median body and other structures and organelles in the giardia cell cycle. For the cell cycle gene expression studies, the actin-related gene was identified by the program geNorm as the most suitable normalizer for reverse transcription-quantitative PCR (RT-qPCR) analysis of the CCE samples. Ten of 11 suspected cell cycle-regulated genes in the CCE fractions have expression profiles in giardia that resemble those of higher eukaryotes. However, the RNA levels of these genes during the cell cycle differ less than 4-fold to 5-fold, which might indicate that large changes in gene expression are not required by giardia to regulate the cell cycle. IMPORTANCE Giardias are among the most commonly reported intestinal protozoa in the world, with infections seen in humans and over 40 species of animals. The life cycle of giardia alternates between the motile trophozoite and the infectious cyst. The regulation of the cell cycle controls the proliferation of giardia trophozoites during an active infection and contains the restriction point for the differentiation of trophozoite to cyst. Here, we developed counterflow centrifugal elutriation as a drug-free method to obtain fractions of giardia cultures enriched in cells from the G1, S, and G2 stages of the cell cycle. Analysis of these fractions showed that the cells do not show side effects associated with the drugs used for synchronization of giardia cultures. Therefore, counterflow centrifugal elutriation would advance studies on key regulatory events during the giardia cell cycle and identify potential drug targets to block giardia proliferation and transmission. PMID:28959734

Diversity and dynamics of plant genome size: an example of polysomaty from a cytogenetic study of Tahitian vanilla (Vanilla xtahitensis, Orchidaceae).

PubMed

Lepers-Andrzejewski, Sandra; Siljak-Yakovlev, Sonja; Brown, Spencer C; Wong, Maurice; Dron, Michel

2011-06-01

Abnormal mitotic behavior with somatic aneuploidy and partial endoreplication were previously reported for the first time in the plant kingdom in Vanilla planifolia. Because vanilla plants are vegetatively propagated, such abnormalities have been transmitted. This study aimed to determine whether mitotic abnormalities also occur in Vanilla hybrid or are suppressed by sexual reproduction. Twenty-eight accessions of Vanilla ×tahitensis, one V. planifolia, and hybrid V. planifolia × V. ×tahitensis were analyzed by chromosome counts, cytometry, and fluorescent in situ hybridization of 18S-5.8S-26S rDNA. In a single root meristem of V. ×tahitensis, chromosome number varied from 22 to 31 in diploids (mean 2C = 5.23 pg), 31 to 41 in triploids (2C = 7.82 pg) and 43 to 60 in tetraploids (2C = 10.27 pg). Morphological diversity is apparently related to ploidy changes. Aneuploidy and partial (asymmetrical) endoreduplication were observed in root meristems of both V. ×tahitensis and the hybrid V. planifolia × V. ×tahitensis, but pollen grains had the euploid chromosome number (n = 15 in diploids). Genome irregularities may be transmitted not only during vegetative propagation but also by sexual reproduction in Vanilla. However, there must be a complex regulation of genome size and organization between the aneuploidy in somatic tissues and subsequently euploid gametic tissue. This is a novel example of polysomaty with developmentally regulated partial endoreplication.

Multi-nucleate retinal pigment epithelium cells of the human macula exhibit a characteristic and highly specific distribution.

PubMed

Starnes, Austin C; Huisingh, Carrie; McGwin, Gerald; Sloan, Kenneth R; Ablonczy, Zsolt; Smith, R Theodore; Curcio, Christine A; Ach, Thomas

2016-01-01

The human retinal pigment epithelium (RPE) is reportedly 3% bi-nucleated. The importance to human vision of multi-nucleated (MN)-RPE cells could be clarified with more data about their distribution in central retina. Nineteen human RPE-flatmounts (9 ≤ 51 years, 10 > 80 years) were imaged at 12 locations: 3 eccentricities (fovea, perifovea, near periphery) in 4 quadrants (superior, inferior, temporal, nasal). Image stacks of lipofuscin-attributable autofluorescence and phalloidin labeled F-actin cytoskeleton were obtained using a confocal fluorescence microscope. Nuclei were devoid of autofluorescence and were marked using morphometric software. Cell areas were approximated by Voronoi regions. Mean number of nuclei per cell among eccentricity/quadrant groups and by age were compared using Poisson and binominal regression models. A total of 11,403 RPE cells at 200 locations were analyzed: 94.66% mono-, 5.31% bi-, 0.02% tri-nucleate, and 0.01% with 5 nuclei. Age had no effect on number of nuclei. There were significant regional differences: highest frequencies of MN-cells were found at the perifovea (9.9%) and near periphery (6.8%). The fovea lacked MN-cells almost entirely. The nasal quadrant had significantly more MN-cells compared to other quadrants, at all eccentricities. This study demonstrates MN-RPE cells in human macula. MN-cells may arise due to endoreplication, cell fusion, or incomplete cell division. The topography of MN-RPE cells follows the topography of photoreceptors; with near-absence at the fovea (cones only) and high frequency at perifovea (highest rod density). This distribution might reflect specific requirements of retinal metabolism or other mechanisms addressable in further studies.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Era, Saho; Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501; Abe, Takuya

Highlights: Black-Right-Pointing-Pointer SENP1 knockout chicken DT40 cells are hypersensitive to spindle poisons. Black-Right-Pointing-Pointer Spindle poison treatment of SENP1{sup -/-} cells leads to increased mitotic slippage. Black-Right-Pointing-Pointer Mitotic slippage in SENP1{sup -/-} cells associates with apoptosis and endoreplication. Black-Right-Pointing-Pointer SENP1 counteracts sister chromatid separation during mitotic arrest. Black-Right-Pointing-Pointer Plk1-mediated cohesion down-regulation is involved in colcemid cytotoxicity. -- Abstract: SUMO conjugation is a reversible posttranslational modification that regulates protein function. SENP1 is one of the six SUMO-specific proteases present in vertebrate cells and its altered expression is observed in several carcinomas. To characterize SENP1 role in genome integrity, we generated Senp1 knockoutmore » chicken DT40 cells. SENP1{sup -/-} cells show normal proliferation, but are sensitive to spindle poisons. This hypersensitivity correlates with increased sister chromatid separation, mitotic slippage, and apoptosis. To test whether the cohesion defect had a causal relationship with the observed mitotic events, we restored the cohesive status of sister chromatids by introducing the TOP2{alpha}{sup +/-} mutation, which leads to increased catenation, or by inhibiting Plk1 and Aurora B kinases that promote cohesin release from chromosomes during prolonged mitotic arrest. Although TOP2{alpha} is SUMOylated during mitosis, the TOP2{alpha}{sup +/-} mutation had no obvious effect. By contrast, inhibition of Plk1 or Aurora B rescued the hypersensitivity of SENP1{sup -/-} cells to colcemid. In conclusion, we identify SENP1 as a novel factor required for mitotic arrest and cohesion maintenance during prolonged mitotic arrest induced by spindle poisons.« less

Natural polyploidy in Vanilla planifolia (Orchidaceae).

PubMed

Bory, Séverine; Catrice, Olivier; Brown, Spencer; Leitch, Ilia J; Gigant, Rodolphe; Chiroleu, Frédéric; Grisoni, Michel; Duval, Marie-France; Besse, Pascale

2008-10-01

Vanilla planifolia accessions cultivated in Reunion Island display important phenotypic variation, but little genetic diversity is demonstrated by AFLP and SSR markers. This study, based on analyses of flow cytometry data, Feulgen microdensitometry data, chromosome counts, and stomatal length measurements, was performed to determine whether polyploidy could be responsible for some of the intraspecific phenotypic variation observed. Vanilla planifolia exhibited an important variation in somatic chromosome number in root cells, as well as endoreplication as revealed by flow cytometry. Nevertheless, the 2C-values of the 50 accessions studied segregated into three distinct groups averaging 5.03 pg (for most accessions), 7.67 pg (for the 'Stérile' phenotypes), and 10.00 pg (for the 'Grosse Vanille' phenotypes). For the three groups, chromosome numbers varied from 16 to 32, 16 to 38, and 22 to 54 chromosomes per cell, respectively. The stomatal length showed a significant variation from 37.75 microm to 48.25 microm. Given that 2C-values, mean chromosome numbers, and stomatal lengths were positively correlated and that 'Stérile' and 'Grosse Vanille' accessions were indistinguishable from 'Classique' accessions using molecular markers, the occurrence of recent autotriploid and autotetraploid types in Reunion Island is supported. This is the first report showing evidence of a recent autopolyploidy in V. planifolia contributing to the phenotypic variation observed in this species.

Signalling requirements for Erwinia amylovora-induced disease resistance, callose deposition and cell growth in the non-host Arabidopsis thaliana.

PubMed

Hamdoun, Safae; Gao, Min; Gill, Manroop; Kwon, Ashley; Norelli, John L; Lu, Hua

2018-05-01

Erwinia amylovora is the causal agent of the fire blight disease in some plants of the Rosaceae family. The non-host plant Arabidopsis serves as a powerful system for the dissection of mechanisms of resistance to E. amylovora. Although not yet known to mount gene-for-gene resistance to E. amylovora, we found that Arabidopsis activated strong defence signalling mediated by salicylic acid (SA), with kinetics and amplitude similar to that induced by the recognition of the bacterial effector avrRpm1 by the resistance protein RPM1. Genetic analysis further revealed that SA signalling, but not signalling mediated by ethylene (ET) and jasmonic acid (JA), is required for E. amylovora resistance. Erwinia amylovora induces massive callose deposition on infected leaves, which is independent of SA, ET and JA signalling and is necessary for E. amylovora resistance in Arabidopsis. We also observed tumour-like growths on E. amylovora-infected Arabidopsis leaves, which contain enlarged mesophyll cells with increased DNA content and are probably a result of endoreplication. The formation of such growths is largely independent of SA signalling and some E. amylovora effectors. Together, our data reveal signalling requirements for E. amylovora-induced disease resistance, callose deposition and cell fate change in the non-host plant Arabidopsis. Knowledge from this study could facilitate a better understanding of the mechanisms of host defence against E. amylovora and eventually improve host resistance to the pathogen. © 2017 BSPP AND JOHN WILEY & SONS LTD.

Induction of polyploidy by nuclear fusion mechanism upon decreased expression of the nuclear envelope protein LAP2β in the human osteosarcoma cell line U2OS.

PubMed

Ben-Shoshan, Shirley Oren; Simon, Amos J; Jacob-Hirsch, Jasmine; Shaklai, Sigal; Paz-Yaacov, Nurit; Amariglio, Ninette; Rechavi, Gideon; Trakhtenbrot, Luba

2014-01-28

Polyploidy has been recognized for many years as an important hallmark of cancer cells. Polyploid cells can arise through cell fusion, endoreplication and abortive cell cycle. The inner nuclear membrane protein LAP2β plays key roles in nuclear envelope breakdown and reassembly during mitosis, initiation of replication and transcriptional repression. Here we studied the function of LAP2β in the maintenance of cell ploidy state, a role which has not yet been assigned to this protein. By knocking down the expression of LAP2β, using both viral and non-viral RNAi approaches in osteosarcoma derived U2OS cells, we detected enlarged nuclear size, nearly doubling of DNA content and chromosomal duplications, as analyzed by fluorescent in situ hybridization and spectral karyotyping methodologies. Spectral karyotyping analyses revealed that near-hexaploid karyotypes of LAP2β knocked down cells consisted of not only seven duplicated chromosomal markers, as could be anticipated by genome duplication mechanism, but also of four single chromosomal markers. Furthermore, spectral karyotyping analysis revealed that both of two near-triploid U2OS sub-clones contained the seven markers that were duplicated in LAP2β knocked down cells, whereas the four single chromosomal markers were detected only in one of them. Gene expression profiling of LAP2β knocked down cells revealed that up to a third of the genes exhibiting significant changes in their expression are involved in cancer progression. Our results suggest that nuclear fusion mechanism underlies the polyploidization induction upon LAP2β reduced expression. Our study implies on a novel role of LAP2β in the maintenance of cell ploidy status. LAP2β depleted U2OS cells can serve as a model to investigate polyploidy and aneuploidy formation by nuclear fusion mechanism and its involvement in cancerogenesis.

Effects of selective inhibitors of Aurora kinases on anaplastic thyroid carcinoma cell lines.

PubMed

Baldini, Enke; Tuccilli, Chiara; Prinzi, Natalie; Sorrenti, Salvatore; Antonelli, Alessandro; Gnessi, Lucio; Morrone, Stefania; Moretti, Costanzo; Bononi, Marco; Arlot-Bonnemains, Yannick; D'Armiento, Massimino; Ulisse, Salvatore

2014-10-01

Aurora kinases are serine/threonine kinases that play an essential role in cell division. Their aberrant expression and/or function induce severe mitotic abnormalities, resulting in either cell death or aneuploidy. Overexpression of Aurora kinases is often found in several malignancies, among which is anaplastic thyroid carcinoma (ATC). We have previously demonstrated the in vitro efficacy of Aurora kinase inhibitors in restraining cell growth and survival of different ATC cell lines. In this study, we sought to establish which Aurora might represent the preferential drug target for ATC. To this end, the effects of two selective inhibitors of Aurora-A (MLN8237) and Aurora-B (AZD1152) on four human ATC cell lines (CAL-62, BHT-101, 8305C, and 8505C) were analysed. Both inhibitors reduced cell proliferation in a time- and dose-dependent manner, with IC50 ranges of 44.3-134.2 nM for MLN8237 and of 9.2-461.3 nM for AZD1152. Immunofluorescence experiments and time-lapse videomicroscopy yielded evidence that each inhibitor induced distinct mitotic phenotypes, but both of them prevented the completion of cytokinesis. As a result, poliploidy increased in all AZD1152-treated cells, and in two out of four cell lines treated with MLN8237. Apoptosis was induced in all the cells by MLN8237, and in BHT-101, 8305C, and 8505C by AZD1152, while CAL-62 exposed to AZD1152 died through necrosis after multiple rounds of endoreplication. Both inhibitors were capable of blocking anchorage-independent cell growth. In conclusion, we demonstrated that either Aurora-A or Aurora-B might represent therapeutic targets for the ATC treatment, but inhibition of Aurora-A appears more effective for suppressing ATC cell proliferation and for inducing the apoptotic pathway. © 2014 Society for Endocrinology.

Two inhibitory systems and CKIs regulate cell cycle exit of mammalian cardiomyocytes after birth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tane, Shoji; Okayama, Hitomi; Ikenishi, Aiko

Mammalian cardiomyocytes actively proliferate during embryonic stages, following which they exit their cell cycle after birth, and the exit is maintained. Previously, we showed that two inhibitory systems (the G1-phase inhibitory system: repression of cyclin D1 expression; the M-phase inhibitory system: inhibition of CDK1 activation) maintain the cell cycle exit of mouse adult cardiomyocytes. We also showed that two CDK inhibitors (CKIs), p21{sup Cip1} and p27{sup Kip1}, regulate the cell cycle exit in a portion of postnatal cardiomyocytes. It remains unknown whether the two inhibitory systems are involved in the cell cycle exit of postnatal cardiomyocytes and whether p21{sup Cip1}more » and p27{sup Kip1} also inhibit entry to M-phase. Here, we showed that more than 40% of cardiomyocytes entered an additional cell cycle by induction of cyclin D1 expression at postnatal stages, but M-phase entry was inhibited in the majority of cardiomyocytes. Marked cell cycle progression and endoreplication were observed in cardiomyocytes of p21{sup Cip1} knockout mice at 4 weeks of age. In addition, tri- and tetranucleated cardiomyocytes increased significantly in p21{sup Cip1} knockout mice. These data showed that the G1-phase inhibitory system and two CKIs (p21{sup Cip1} and p27{sup Kip1}) inhibit entry to an additional cell cycle in postnatal cardiomyocytes, and that the M-phase inhibitory system and p21{sup Cip1} inhibit M-phase entry of cardiomyocytes which have entered the additional cell cycle. - Highlights: • Many postnatal cardiomyocytes entered an additional cell cycle by cyclin D1 induction. • The majority of cardiomyocytes could not enter M-phase after cyclin D1 induction. • Cell cycle progressed markedly in p21{sup Cip1} knockout mice after postnatal day 14. • Tri- and tetranucleated cardiomyocytes increased in p21{sup Cip1} knockout mice.« less

CDK inhibitors, p21{sup Cip1} and p27{sup Kip1}, participate in cell cycle exit of mammalian cardiomyocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tane, Shoji; Ikenishi, Aiko; Okayama, Hitomi

2014-01-17

Highlights: •Expression of p21 and p27 in the hearts showed a peak during postnatal stages. •p21 and p27 bound to cyclin E, cyclin A and CDK2 in the hearts at postnatal stages. •Cardiomyocytes in both KO mice showed failure in the cell cycle exit at G1-phase. •These data show the first apparent phenotypes in the hearts of Cip/Kip KO mice. -- Abstract: Mammalian cardiomyocytes actively proliferate during embryonic stages, following which cardiomyocytes exit their cell cycle after birth. The irreversible cell cycle exit inhibits cardiac regeneration by the proliferation of pre-existing cardiomyocytes. Exactly how the cell cycle exit occurs remainsmore » largely unknown. Previously, we showed that cyclin E- and cyclin A-CDK activities are inhibited before the CDKs levels decrease in postnatal stages. This result suggests that factors such as CDK inhibitors (CKIs) inhibit CDK activities, and contribute to the cell cycle exit. In the present study, we focused on a Cip/Kip family, which can inhibit cyclin E- and cyclin A-CDK activities. Expression of p21{sup Cip1} and p27{sup Kip1} but not p57{sup Kip2} showed a peak around postnatal day 5, when cyclin E- and cyclin A-CDK activities start to decrease. p21{sup Cip1} and p27{sup Kip1} bound to cyclin E, cyclin A and CDK2 at postnatal stages. Cell cycle distribution patterns of postnatal cardiomyocytes in p21{sup Cip1} and p27{sup Kip1} knockout mice showed failure in the cell cycle exit at G1-phase, and endoreplication. These results indicate that p21{sup Cip1} and p27{sup Kip} play important roles in the cell cycle exit of postnatal cardiomyocytes.« less

PRM1 and KAR5 function in cell-cell fusion and karyogamy to drive distinct bisexual and unisexual cycles in the Cryptococcus pathogenic species complex

PubMed Central

Fu, Ci; Heitman, Joseph

2017-01-01

Sexual reproduction is critical for successful evolution of eukaryotic organisms in adaptation to changing environments. In the opportunistic human fungal pathogens, the Cryptococcus pathogenic species complex, C. neoformans primarily undergoes bisexual reproduction, while C. deneoformans undergoes both unisexual and bisexual reproduction. During both unisexual and bisexual cycles, a common set of genetic circuits regulates a yeast-to-hyphal morphological transition, that produces either monokaryotic or dikaryotic hyphae. As such, both the unisexual and bisexual cycles can generate genotypic and phenotypic diversity de novo. Despite the similarities between these two cycles, genetic and morphological differences exist, such as the absence of an opposite mating-type partner and monokaryotic instead of dikaryotic hyphae during C. deneoformans unisexual cycle. To better understand the similarities and differences between these modes of sexual reproduction, we focused on two cellular processes involved in sexual reproduction: cell-cell fusion and karyogamy. We identified orthologs of the plasma membrane fusion protein Prm1 and the nuclear membrane fusion protein Kar5 in both Cryptococcus species, and demonstrated their conserved roles in cell fusion and karyogamy during C. deneoformans α-α unisexual reproduction and C. deneoformans and C. neoformans a-α bisexual reproduction. Notably, karyogamy occurs inside the basidum during bisexual reproduction in C. neoformans, but often occurs earlier following cell fusion during bisexual reproduction in C. deneoformans. Characterization of these two genes also showed that cell fusion is dispensable for solo unisexual reproduction in C. deneoformans. The blastospores produced along hyphae during C. deneoformans unisexual reproduction are diploid, suggesting that diploidization occurs early during hyphal development, possibly through either an endoreplication pathway or cell fusion-independent karyogamy events. Taken together, our findings suggest distinct mating mechanisms for unisexual and bisexual reproduction in Cryptococcus, exemplifying distinct evolutionary trajectories within this pathogenic species complex. PMID:29176784

The MADS-box XAANTAL1 increases proliferation at the Arabidopsis root stem-cell niche and participates in transition to differentiation by regulating cell-cycle components

PubMed Central

García-Cruz, Karla V.; García-Ponce, Berenice; Garay-Arroyo, Adriana; Sanchez, María De La Paz; Ugartechea-Chirino, Yamel; Desvoyes, Bénédicte; Pacheco-Escobedo, Mario A.; Tapia-López, Rosalinda; Ransom-Rodríguez, Ivan; Gutierrez, Crisanto; Alvarez-Buylla, Elena R.

2016-01-01

Background Morphogenesis depends on the concerted modulation of cell proliferation and differentiation. Such modulation is dynamically adjusted in response to various external and internal signals via complex transcriptional regulatory networks that mediate between such signals and regulation of cell-cycle and cellular responses (proliferation, growth, differentiation). In plants, which are sessile, the proliferation/differentiation balance is plastically adjusted during their life cycle and transcriptional networks are important in this process. MADS-box genes are key developmental regulators in eukaryotes, but their role in cell proliferation and differentiation modulation in plants remains poorly studied. Methods We characterize the XAL1 loss-of-function xal1-2 allele and overexpression lines using quantitative cellular and cytometry analyses to explore its role in cell cycle, proliferation, stem-cell patterning and transition to differentiation. We used quantitative PCR and cellular markers to explore if XAL1 regulates cell-cycle components and PLETHORA1 (PLT1) gene expression, as well as confocal microscopy to analyse stem-cell niche organization. Key Results We previously showed that XAANTAL1 (XAL1/AGL12) is necessary for Arabidopsis root development as a promoter of cell proliferation in the root apical meristem. Here, we demonstrate that XAL1 positively regulates the expression of PLT1 and important components of the cell cycle: CYCD3;1, CYCA2;3, CYCB1;1, CDKB1;1 and CDT1a. In addition, we show that xal1-2 mutant plants have a premature transition to differentiation with root hairs appearing closer to the root tip, while endoreplication in these plants is partially compromised. Coincidently, the final size of cortex cells in the mutant is shorter than wild-type cells. Finally, XAL1 overexpression-lines corroborate that this transcription factor is able to promote cell proliferation at the stem-cell niche. Conclusion XAL1 seems to be an important component of the networks that modulate cell proliferation/differentiation transition and stem-cell proliferation during Arabidopsis root development; it also regulates several cell-cycle components. PMID:27474508

Defective endomitosis during megakaryopoiesis leads to thrombocytopenia in Fanca−/− mice

PubMed Central

Pawlikowska, Patrycja; Fouchet, Pierre; Vainchenker, William; Rosselli, Filippo

2014-01-01

Fanconi anemia (FA) is an inherited chromosomal instability syndrome that is characterized by progressive bone marrow failure. One of the main causes of morbidity and mortality in FA is a bleeding tendency, resulting from low platelet counts. Platelets are the final products of megakaryocyte (MK) maturation. Here, we describe a previously unappreciated role of Fanconi anemia group A protein (Fanca) during the endomitotic process of MK differentiation. Fanca deficiency leads to the accumulation of MKs with low nuclear ploidy and to decreased platelet production. We show, for the first time, that Fanca−/− mice are characterized by limited number and proliferative capacity of MK progenitors. Defective megakaryopoiesis of Fanca−/− cells is associated with the formation of nucleoplasmic bridges and increased chromosomal instability, indicating that inaccurate endoreplication and karyokinesis occur during MK polyploidization. Sustained DNA damage forces Fanca−/− MKs to enter a senescence-like state. Furthermore, inhibition of the Rho-associated kinase, a regulator of cytokinesis, improves the polyploidization of Fanca−/− MKs but greatly increases their genomic instability and diminishes their differentiation potential, supporting the notion that accumulation of DNA damage through endomitotic cycles affects MK maturation. Our study indicates that Fanca expression during endomitosis is crucial for normal megakaryopoiesis and platelet production. PMID:25261197

Defective endomitosis during megakaryopoiesis leads to thrombocytopenia in Fanca-/- mice.

PubMed

Pawlikowska, Patrycja; Fouchet, Pierre; Vainchenker, William; Rosselli, Filippo; Naim, Valeria

2014-12-04

Fanconi anemia (FA) is an inherited chromosomal instability syndrome that is characterized by progressive bone marrow failure. One of the main causes of morbidity and mortality in FA is a bleeding tendency, resulting from low platelet counts. Platelets are the final products of megakaryocyte (MK) maturation. Here, we describe a previously unappreciated role of Fanconi anemia group A protein (Fanca) during the endomitotic process of MK differentiation. Fanca deficiency leads to the accumulation of MKs with low nuclear ploidy and to decreased platelet production. We show, for the first time, that Fanca(-/-) mice are characterized by limited number and proliferative capacity of MK progenitors. Defective megakaryopoiesis of Fanca(-/-) cells is associated with the formation of nucleoplasmic bridges and increased chromosomal instability, indicating that inaccurate endoreplication and karyokinesis occur during MK polyploidization. Sustained DNA damage forces Fanca(-/-) MKs to enter a senescence-like state. Furthermore, inhibition of the Rho-associated kinase, a regulator of cytokinesis, improves the polyploidization of Fanca(-/-) MKs but greatly increases their genomic instability and diminishes their differentiation potential, supporting the notion that accumulation of DNA damage through endomitotic cycles affects MK maturation. Our study indicates that Fanca expression during endomitosis is crucial for normal megakaryopoiesis and platelet production. © 2014 by The American Society of Hematology.

The MADS-box XAANTAL1 increases proliferation at the Arabidopsis root stem-cell niche and participates in transition to differentiation by regulating cell-cycle components.

PubMed

García-Cruz, Karla V; García-Ponce, Berenice; Garay-Arroyo, Adriana; Sanchez, María De La Paz; Ugartechea-Chirino, Yamel; Desvoyes, Bénédicte; Pacheco-Escobedo, Mario A; Tapia-López, Rosalinda; Ransom-Rodríguez, Ivan; Gutierrez, Crisanto; Alvarez-Buylla, Elena R

2016-07-29

Morphogenesis depends on the concerted modulation of cell proliferation and differentiation. Such modulation is dynamically adjusted in response to various external and internal signals via complex transcriptional regulatory networks that mediate between such signals and regulation of cell-cycle and cellular responses (proliferation, growth, differentiation). In plants, which are sessile, the proliferation/differentiation balance is plastically adjusted during their life cycle and transcriptional networks are important in this process. MADS-box genes are key developmental regulators in eukaryotes, but their role in cell proliferation and differentiation modulation in plants remains poorly studied. We characterize the XAL1 loss-of-function xal1-2 allele and overexpression lines using quantitative cellular and cytometry analyses to explore its role in cell cycle, proliferation, stem-cell patterning and transition to differentiation. We used quantitative PCR and cellular markers to explore if XAL1 regulates cell-cycle components and PLETHORA1 (PLT1) gene expression, as well as confocal microscopy to analyse stem-cell niche organization. We previously showed that XAANTAL1 (XAL1/AGL12) is necessary for Arabidopsis root development as a promoter of cell proliferation in the root apical meristem. Here, we demonstrate that XAL1 positively regulates the expression of PLT1 and important components of the cell cycle: CYCD3;1, CYCA2;3, CYCB1;1, CDKB1;1 and CDT1a In addition, we show that xal1-2 mutant plants have a premature transition to differentiation with root hairs appearing closer to the root tip, while endoreplication in these plants is partially compromised. Coincidently, the final size of cortex cells in the mutant is shorter than wild-type cells. Finally, XAL1 overexpression-lines corroborate that this transcription factor is able to promote cell proliferation at the stem-cell niche. XAL1 seems to be an important component of the networks that modulate cell proliferation/differentiation transition and stem-cell proliferation during Arabidopsis root development; it also regulates several cell-cycle components. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Heterochromatin and rDNA sites distribution in the holocentric chromosomes of Cuscuta approximata Bab. (Convolvulaceae).

PubMed

Guerra, Marcelo; García, Miguel A

2004-02-01

Cuscuta is a widely distributed genus of holoparasitic plants. Holocentric chromosomes have been reported only in species of one of its subgenera (Cuscuta subg. Cuscuta). In this work, a representative of this subgenus, Cuscuta approximata, was investigated looking for its mitotic and meiotic chromosome behaviour and the heterochromatin distribution. The mitotic chromosomes showed neither primary constriction nor Rabl orientation whereas the meiotic ones exhibited the typical quadripartite structure characteristic of holocentrics, supporting the assumption of holocentric chromosomes as a synapomorphy of Cuscuta subg. Cuscuta. Chromosomes and interphase nuclei displayed many heterochromatic blocks that stained deeply with hematoxylin, 4',6-diamidino-2-phenylindole (DAPI), or after C banding. The banded karyotype showed terminal or subterminal bands in all chromosomes and central bands in some of them. The single pair of 45S rDNA sites was observed at the end of the largest chromosome pair, close to a DAPI band and a 5S rDNA site. Two other 5S rDNA site pairs were found, both closely associated with DAPI bands. The noteworthy giant nuclei of glandular cells of petals and ovary wall exhibited large chromocentres typical of polytenic nuclei. The chromosomal location of heterochromatin and rDNA sites and the structure of the endoreplicated nuclei of C. approximata seemed to be similar to those known in monocentric nuclei, suggesting that centromeric organization has little or no effect on chromatin organization.

Immunohistochemical study of Ulex europaeus agglutinin 1 (UEA-1) binding of megakaryocytes in bone marrow biopsy specimens: demonstration of heterogeneity in staining pattern reflecting the stages of differentiation.

PubMed

Liu, S M; Li, C Y

1996-01-01

During differentiation, megakaryocytes undergo nuclear endoreplication, an increase in cell size, cytoplasmic granulation, and release of platelets. The changes in highly lobulated nuclei with varying degree of polyploidy and increasing cell size are easily recognized morphologically. However, the actual cytoplasmic changes are more difficult to perceive morphologically. With the peroxidase-antiperoxidase (PAP) method using UEA-1 as the binding protein to the alpha-L-fucose of glycoprotein synthesized by megakaryocytes, we observed significant variation in cytoplasmic staining of megakaryocytes in routinely processed bone marrow biopsy sections. A total of 3344 megakaryocytes in bone marrow sections from 10 patients with nonhematologic diseases and from 10 patients with idiopathic thrombocytopenic purpura (ITP) was studied. According to the intensity and pattern of cytoplasmic staining, we divided megakaryocytes into at least six groups: (1) low granular (LG), (2) diffuse granular (DG), (3) diffuse dense granular (DDG), (4) marginal granular (MG), (5) denuded (DMK), and (6) endomitotic (EndoM). Most of the megakaryocytes were DG (mean, 42.75% +/- 19.21%) and DDG (mean, 50.25% +/- 21.23%). In correlation with nuclear morphology and cell size, it appears that substances binding to UEA-1 are located in the paranuclear region in early megakaryocytes and produce a low granular focal staining pattern (LG cells). Next, the granules spread throughout the cytoplasm (DG cells) and increase in quantity (DDG). This is followed by migration of granules to the periphery of the cytoplasm (MG cells) and is associated with the liberation of platelets and eventual formation of DMK megakaryocytes. Endomitosis, regulated by unknown factors, occurred in the MG stage. In comparing the group with nonhematologic disease (mean DG, 35.4% +/- 18.48%; DDG, 58.4% +/- 21.8%) and the group with ITP (mean DG, 50.1% +/- 17.82%; DDG, 42.1% +/- 18.12%), we found an increasing proportion of DG megakaryocytes in ITP, which suggests a left-shifted maturation of megakaryocytes. By understanding the staining pattern seen in the different stages of megakaryocytic differentiation, UEA-1 staining may be a practical method for studying megakaryocytopoiesis in routinely processed paraffin sections of bone marrow biopsy samples.

Concerted control of Escherichia coli cell division

PubMed Central

Osella, Matteo; Nugent, Eileen; Cosentino Lagomarsino, Marco

2014-01-01

The coordination of cell growth and division is a long-standing problem in biology. Focusing on Escherichia coli in steady growth, we quantify cell division control using a stochastic model, by inferring the division rate as a function of the observable parameters from large empirical datasets of dividing cells. We find that (i) cells have mechanisms to control their size, (ii) size control is effected by changes in the doubling time, rather than in the single-cell elongation rate, (iii) the division rate increases steeply with cell size for small cells, and saturates for larger cells. Importantly, (iv) the current size is not the only variable controlling cell division, but the time spent in the cell cycle appears to play a role, and (v) common tests of cell size control may fail when such concerted control is in place. Our analysis illustrates the mechanisms of cell division control in E. coli. The phenomenological framework presented is sufficiently general to be widely applicable and opens the way for rigorous tests of molecular cell-cycle models. PMID:24550446

Modular Battery Controller

NASA Technical Reports Server (NTRS)

Button, Robert M (Inventor); Gonzalez, Marcelo C (Inventor)

2017-01-01

Some embodiments of the present invention describe a battery including a plurality of master-less controllers. Each controller is operatively connected to a corresponding cell in a string of cells, and each controller is configured to bypass a fraction of current around the corresponding cell when the corresponding cell has a greater charge than one or more other cells in the string of cells.

Electrolytic Valving Isolation for Cell Co-Culture Microenvironment with Controlled Cell Pairing Ratios

PubMed Central

Chen, Yu-Chih; Ingram, Patrick; Yoon, Euisik

2016-01-01

Cancer-stromal interaction is a critical process in tumorigenesis. Conventional dish-based co-culture assays simply mix two cell types in the same dish; thus, they are deficient in controlling cell locations and precisely tracking single cell behavior from heterogeneous cell populations. Microfluidic technology can provide a good spatial temporal control of microenvironments, but the control has been typically realized by using external pumps, making long-term cultures cumbersome and bulky. In this work, we present a cell-cell interaction microfluidic platform that can accurately control co-culture microenvironment by using a novel electrolytic cell isolation scheme without using any valves or pneumatic pumps. The proposed microfluidic platform can also precisely control the number of interacting cells and pairing ratios to emulate cancer niches. More than 80% of the chambers captured the desired number of cells. The duration of cell isolation can be adjusted by electrolytic bubble generation and removal. We verified that electrolytic process has a negligible effect on cell viability and proliferation in our platform. To the best of our knowledge, this work is the first attempt to incorporate electrolytic bubble generation as a cell isolation method in microfluidics. For proof of feasibility, we performed cell-cell interaction assays between prostate cancer (PC3) cells and myoblast (C2C12) cells. The preliminary results demonstrated the potential of using electrolysis for micro-environmental control during cell culture. Also, the ratio controlled cell-cell interaction assays was successfully performed showing that the cell pairing ratios of PC3 to C2C12 affected the proliferation rate of myoblast cells due to increased secretion of growth factors from prostate cancer cells. PMID:25118341

A light-controlled cell lysis system in bacteria.

PubMed

Wang, Geyi; Lu, Xin; Zhu, Yisha; Zhang, Wei; Liu, Jiahui; Wu, Yankang; Yu, Liyang; Sun, Dongchang; Cheng, Feng

2018-05-08

Intracellular products (e.g., insulin), which are obtained through cell lysis, take up a big share of the biotech industry. It is often time-consuming, laborious, and environment-unfriendly to disrupt bacterial cells with traditional methods. In this study, we developed a molecular device for controlling cell lysis with light. We showed that intracellular expression of a single lysin protein was sufficient for efficient bacterial cell lysis. By placing the lysin-encoding gene under the control of an improved light-controlled system, we successfully controlled cell lysis by switching on/off light: OD 600 of the Escherichia coli cell culture was decreased by twofold when the light-controlled system was activated under dark condition. We anticipate that our work would not only pave the way for cell lysis through a convenient biological way in fermentation industry, but also provide a paradigm for applying the light-controlled system in other fields of biotech industry.

Cooperativity of HIV-Specific Cytolytic CD4 T Cells and CD8 T Cells in Control of HIV Viremia

PubMed Central

Johnson, Susan; Eller, Michael; Teigler, Jeffrey E.; Maloveste, Sebastien M.; Schultz, Bruce T.; Soghoian, Damien Z.; Lu, Richard; Oster, Alexander F.; Chenine, Agnès-Laurence; Alter, Galit; Dittmer, Ulf; Marovich, Mary; Robb, Merlin L.; Michael, Nelson L.; Bolton, Diane

2015-01-01

ABSTRACT CD4+ T cells play a pivotal role in the control of chronic viral infections. Recently, nontraditional CD4+ T cell functions beyond helper effects have been described, and a role for cytolytic CD4+ T cells in the control of HIV infection has been suggested. We define here the transcriptional, phenotypic, and functional profiles of HIV-specific cytolytic CD4+ T cells. Fluidigm BioMark and multiparameter flow cytometric analysis of HIV-specific cytolytic CD4+ T cells revealed a distinct transcriptional signature compared to Th1 CD4+ cells but shared similar features with HIV-specific cytolytic CD8+ T cells. Furthermore, HIV-specific cytolytic CD4+ T cells showed comparable killing activity relative to HIV-specific CD8+ T cells and worked cooperatively in the elimination of virally infected cells. Interestingly, we found that cytolytic CD4+ T cells emerge early during acute HIV infection and tightly follow acute viral load trajectory. This emergence was associated to the early viral set point, suggesting an involvement in early control, in spite of CD4 T cell susceptibility to HIV infection. Our data suggest cytolytic CD4+ T cells as an independent subset distinct from Th1 cells that show combined activity with CD8+ T cells in the long-term control of HIV infection. IMPORTANCE The ability of the immune system to control chronic HIV infection is of critical interest to both vaccine design and therapeutic approaches. Much research has focused on the effect of the ability of CD8+ T cells to control the virus, while CD4+ T cells have been overlooked as effectors in HIV control due to the fact that they are preferentially infected. We show here that a subset of HIV-specific CD4+ T cells cooperate in the cytolytic control of HIV replication. Moreover, these cells represent a distinct subset of CD4+ T cells showing significant transcriptional and phenotypic differences compared to HIV-specific Th1 cells but with similarities to CD8+ T cells. These findings are important for our understanding of HIV immunopathology. PMID:25972560

Specific CD8+ T Cell Responses Correlate with Control of Simian Immunodeficiency Virus Replication in Mauritian Cynomolgus Macaques

PubMed Central

Budde, Melisa L.; Greene, Justin M.; Chin, Emily N.; Ericsen, Adam J.; Scarlotta, Matthew; Cain, Brian T.; Pham, Ngoc H.; Becker, Ericka A.; Harris, Max; Weinfurter, Jason T.; O'Connor, Shelby L.; Piatak, Michael; Lifson, Jeffrey D.; Gostick, Emma; Price, David A.; Friedrich, Thomas C.

2012-01-01

Specific major histocompatibility complex (MHC) class I alleles are associated with an increased frequency of spontaneous control of human and simian immunodeficiency viruses (HIV and SIV). The mechanism of control is thought to involve MHC class I-restricted CD8+ T cells, but it is not clear whether particular CD8+ T cell responses or a broad repertoire of epitope-specific CD8+ T cell populations (termed T cell breadth) are principally responsible for mediating immunologic control. To test the hypothesis that heterozygous macaques control SIV replication as a function of superior T cell breadth, we infected MHC-homozygous and MHC-heterozygous cynomolgus macaques with the pathogenic virus SIVmac239. As measured by a gamma interferon enzyme-linked immunosorbent spot assay (IFN-γ ELISPOT) using blood, T cell breadth did not differ significantly between homozygotes and heterozygotes. Surprisingly, macaques that controlled SIV replication, regardless of their MHC zygosity, shared durable T cell responses against similar regions of Nef. While the limited genetic variability in these animals prevents us from making generalizations about the importance of Nef-specific T cell responses in controlling HIV, these results suggest that the T cell-mediated control of virus replication that we observed is more likely the consequence of targeting specificity rather than T cell breadth. PMID:22573864

The cell cycle.

PubMed

Singh, N; Lim, R B; Sawyer, M A

2000-07-01

The cell cycle and the cell cycle control system are the engines that drive life. They allow for the processes of cell renewal and the growth of organisms, under controlled conditions. The control system is essential for the monitoring of normal cell growth and replication of genetic material and to ensure that normal, functional daughter cells are produced at completion of each cell cycle. Although certain clinical applications exist which take advantage of the events of the cell cycle, our understanding of its mechanisms and how to manipulate them is infantile. The next decades will continue to see the effort of many researchers focused upon unlocking the mysteries of the cell cycle and the cell cycle control system.

Culture and Characterization of Circulating Endothelial Progenitor Cells in Patients with Renal Cell Carcinoma.

PubMed

Gu, Wenyu; Sun, Wei; Guo, Changcheng; Yan, Yang; Liu, Min; Yao, Xudong; Yang, Bin; Zheng, Junhua

2015-07-01

Although emerging evidence demonstrates increased circulating endothelial progenitor cells in patients with solid tumors, to our knowledge it is still unknown whether such cells can be cultured from patients with highly angiogenic renal cell carcinoma. We cultured and characterized circulating endothelial progenitor cells from patients with renal cell carcinoma. The circulating endothelial progenitor cell level (percent of CD45(-)CD34(+) VEGF-R2(+) cells in total peripheral blood mononuclear cells) was quantified in 47 patients with renal cell carcinoma and 40 healthy controls. Peripheral blood mononuclear cells were then isolated from 33 patients with renal cell carcinoma and 30 healthy controls to culture and characterize circulating endothelial progenitor cells. The circulating endothelial progenitor cell level was significantly higher in patients with renal cell carcinoma than in healthy controls (0.276% vs 0.086%, p <0.001). A colony of circulating endothelial progenitor cells first emerged significantly earlier in patient than in control preparations (6.72 vs 14.67 days, p <0.001). The culture success rate (87.8% vs 40.0% of participants) and the number of colonies (10.06 vs 1.83) were significantly greater for patients than for controls (each p <0.001). The circulating endothelial progenitor cell level correlated positively with the number of patient colonies (r = 0.762, p <0.001). Cells cultured from patients and controls showed a similar growth pattern, immunophenotype, ability to uptake Ac-LDL and bind lectin, and form capillary tubes in vitro. However, significantly more VEGF-R2(+) circulating endothelial progenitor cells were found in preparations from patients with renal cell carcinoma than from healthy controls (21.1% vs 13.4%, p <0.001). Earlier emergence of circulating endothelial progenitor cell colonies, a higher cell culture success rate and more colonies were found for patients with renal cell carcinoma than for healthy controls. Results indicate the important significance of VEGF-R2(+) circulating endothelial progenitors in patients with renal cell carcinoma. Copyright © 2015 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

Antibody-Independent Control of γ-Herpesvirus Latency via B Cell Induction of Anti-Viral T Cell Responses

PubMed Central

McClellan, Kelly B; Gangappa, Shivaprakash; Speck, Samuel H; Virgin, Herbert W.

2006-01-01

B cells can use antibody-dependent mechanisms to control latent viral infections. It is unknown whether this represents the sole function of B cells during chronic viral infection. We report here that hen egg lysozyme (HEL)-specific B cells can contribute to the control of murine γ-herpesvirus 68 (γHV68) latency without producing anti-viral antibody. HEL-specific B cells normalized defects in T cell numbers and proliferation observed in B cell−/− mice during the early phase of γHV68 latency. HEL-specific B cells also reversed defects in CD8 and CD4 T cell cytokine production observed in B cell−/− mice, generating CD8 and CD4 T cells necessary for control of latency. Furthermore, HEL-specific B cells were able to present virally encoded antigen to CD8 T cells. Therefore, B cells have antibody independent functions, including antigen presentation, that are important for control of γ-herpesvirus latency. Exploitation of this property of B cells may allow enhanced vaccine responses to chronic virus infection. PMID:16789842

Directional control of lamellipodia extension by constraining cell shape and orienting cell tractional forces

NASA Technical Reports Server (NTRS)

Parker, Kevin Kit; Brock, Amy Lepre; Brangwynne, Cliff; Mannix, Robert J.; Wang, Ning; Ostuni, Emanuele; Geisse, Nicholas A.; Adams, Josephine C.; Whitesides, George M.; Ingber, Donald E.

2002-01-01

Directed cell migration is critical for tissue morphogenesis and wound healing, but the mechanism of directional control is poorly understood. Here we show that the direction in which cells extend their leading edge can be controlled by constraining cell shape using micrometer-sized extracellular matrix (ECM) islands. When cultured on square ECM islands in the presence of motility factors, cells preferentially extended lamellipodia, filopodia, and microspikes from their corners. Square cells reoriented their stress fibers and focal adhesions so that tractional forces were concentrated in these corner regions. When cell tension was dissipated, lamellipodia extension ceased. Mechanical interactions between cells and ECM that modulate cytoskeletal tension may therefore play a key role in the control of directional cell motility.

[Distribution of lymphocyte subpopulations and plasma cells in the colonic mucosa of children with ulcerative colitis].

PubMed

Arató, A; Savilahti, E; Tainio, V M

1990-09-02

The distribution of lymphocyte subpopulations and plasma cells of the colonic and rectal mucosae were studied in eight children with ulcerative colitis and 12 healthy controls. In four patients the examinations were also carried out 3 months after the beginning of treatment. No difference in the number of intraepithelial lymphocytes was found between the patients and controls. The majority of these cells were T-cells, and among them the suppressor/cytotoxic cells were preponderant. In the lamina propria of both untreated and treated patients the numbers of T-cells, helper T-cells, and B-cells were elevated compared to controls. In the patients the number of IgG-containing cells was three times that of the controls; the number of IgE positive cells was also elevated. The numbers of IgA- and IgM-containing cells were not different from that of the controls. The results suggest that in ulcerative colitis the place of primary immunological processes inside the large bowel mucosa is the lamina propria.

Chronic inorganic arsenic exposure in vitro induces a cancer cell phenotype in human peripheral lung epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Person, Rachel J.; Olive Ngalame, Ntube N.; Makia, Ngome L.

Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomousmore » growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer. - Highlights: • Chronic arsenic exposure transforms a human peripheral lung epithelia cell line. • Cells acquire characteristics in common with human lung adenocarcinoma cells. • These transformed cells provide a valuable model for arsenic-induced lung cancer.« less

The architecture and conservation pattern of whole-cell control circuitry.

PubMed

McAdams, Harley H; Shapiro, Lucy

2011-05-27

The control circuitry that directs and paces Caulobacter cell cycle progression involves the entire cell operating as an integrated system. This control circuitry monitors the environment and the internal state of the cell, including the cell topology, as it orchestrates orderly activation of cell cycle subsystems and Caulobacter's asymmetric cell division. The proteins of the Caulobacter cell cycle control system and its internal organization are co-conserved across many alphaproteobacteria species, but there are great differences in the regulatory apparatus' functionality and peripheral connectivity to other cellular subsystems from species to species. This pattern is similar to that observed for the "kernels" of the regulatory networks that regulate development of metazoan body plans. The Caulobacter cell cycle control system has been exquisitely optimized as a total system for robust operation in the face of internal stochastic noise and environmental uncertainty. When sufficient details accumulate, as for Caulobacter cell cycle regulation, the system design has been found to be eminently rational and indeed consistent with good design practices for human-designed asynchronous control systems. Copyright © 2011 Elsevier Ltd. All rights reserved.

Fabrication of corneal epithelial cell sheets maintaining colony-forming cells without feeder cells by oxygen-controlled method.

PubMed

Nakajima, Ryota; Takeda, Shizu

2014-01-01

The use of murine 3T3 feeder cells needs to be avoided when fabricating corneal epithelial cell sheets for use in treating ocular surface diseases. However, the expression level of the epithelial stem/progenitor cell marker, p63, is down-regulated in feeder-free culture systems. In this study, in order to fabricate corneal epithelial cell sheets that maintain colony-forming cells without using any feeder cells, we investigated the use of an oxygen-controlled method that was developed previously to fabricate cell sheets efficiently. Rabbit limbal epithelial cells were cultured under hypoxia (1-10% O2) and under normoxia during stratification after reaching confluence. Multilayered corneal epithelial cell sheets were fabricated using an oxygen-controlled method, and immunofluorescence analysis showed that cytokeratin 3 and p63 was expressed in appropriate localization in the cell sheets. The colony-forming efficiency of the cell sheets fabricated by the oxygen-controlled method without feeder cells was significantly higher than that of cell sheets fabricated under 20% O2 without feeder cells. These results indicate that the oxygen-controlled method has the potential to achieve a feeder-free culture system for fabricating corneal epithelial cell sheets for corneal regeneration. Copyright © 2013 Elsevier Ltd. All rights reserved.

Remote control of therapeutic T cells through a small molecule-gated chimeric receptor

PubMed Central

Wu, Chia-Yung; Roybal, Kole T.; Puchner, Elias M.; Onuffer, James; Lim, Wendell A.

2016-01-01

There is growing promise in using engineered cells as therapeutic agents. For example, synthetic Chimeric Antigen Receptors (CARs) can redirect T cells to recognize and eliminate tumor cells expressing specific antigens. Despite promising clinical results, excessive activity and poor control over such engineered T cells can cause severe toxicities. We present the design of “ON-switch” CARs that enable small molecule-control over T cell therapeutic functions, while still retaining antigen specificity. In these split receptors, antigen binding and intracellular signaling components only assemble in the presence of a heterodimerizing small molecule. This titratable pharmacologic regulation could allow physicians to precisely control the timing, location, and dosage of T cell activity, thereby mitigating toxicity. This work illustrates the potential of combining cellular engineering with orthogonal chemical tools to yield safer therapeutic cells that tightly integrate both cell autonomous recognition and user control. PMID:26405231

Remote control of therapeutic T cells through a small molecule-gated chimeric receptor.

PubMed

Wu, Chia-Yung; Roybal, Kole T; Puchner, Elias M; Onuffer, James; Lim, Wendell A

2015-10-16

There is growing interest in using engineered cells as therapeutic agents. For example, synthetic chimeric antigen receptors (CARs) can redirect T cells to recognize and eliminate tumor cells expressing specific antigens. Despite promising clinical results, these engineered T cells can exhibit excessive activity that is difficult to control and can cause severe toxicity. We designed "ON-switch" CARs that enable small-molecule control over T cell therapeutic functions while still retaining antigen specificity. In these split receptors, antigen-binding and intracellular signaling components assemble only in the presence of a heterodimerizing small molecule. This titratable pharmacologic regulation could allow physicians to precisely control the timing, location, and dosage of T cell activity, thereby mitigating toxicity. This work illustrates the potential of combining cellular engineering with orthogonal chemical tools to yield safer therapeutic cells that tightly integrate cell-autonomous recognition and user control. Copyright © 2015, American Association for the Advancement of Science.

The mediator subunit Med23 contributes to controlling T-cell activation and prevents autoimmunity.

PubMed

Sun, Yang; Zhu, Xiaoyan; Chen, Xufeng; Liu, Haifeng; Xu, Yu; Chu, Yajing; Wang, Gang; Liu, Xiaolong

2014-10-10

T-cell activation is critical for successful immune responses and is controlled at multiple levels. Although many changes of T-cell receptor-associated signalling molecules affect T-cell activation, the transcriptional mechanisms that control this process remain largely unknown. Here we find that T cell-specific deletion of the mediator subunit Med23 leads to hyperactivation of T cells and aged Med23-deficient mice exhibit an autoimmune syndrome. Med23 specifically and consistently promotes the transcription of multiple negative regulators of T-cell activation. In the absence of Med23, the T-cell activation threshold is lower, which results in enhanced antitumour T-cell function. Cumulatively, our data suggest that Med23 contributes to controlling T-cell activation at the transcriptional level and prevents the development of autoimmunity.

Using a simulation cell for exercise realism.

PubMed

Lerner, Ken

2013-01-01

A simulation cell or SimCell is an effective and flexible tool for control of emergency management exercises. It allows exercise participants to interact, via simulation, with a wide variety of nonplaying organizations and officials. Adapted from military application, the Chemical Stockpile Emergency Preparedness Program (CSEPP) applied, developed, and refined the SimCell concept for emergency management exercises. It has now been incorporated into national exercise guidance through the Homeland Security Exercise and Evaluation Program, and has been used in a wide variety of national, regional, and local exercises. This article reviews development of the SimCell concept in CSEPP, briefly surveys current practice incorporating SimCells in exercise control, and offers practical lessons-learned and tips on using a SimCell to best advantage. Lessons learned include using a SimCell as an exercise-control hub; preparing inject material for exercise controllers as part of the Master Scenario Event List; laying the groundwork for success through exercise player and controller training; developing protocol for SimCell communications; and capturing feedback from SimCell controllers for inclusion in the exercise evaluation reporting process. The SimCell concept is flexible and can be applied to a variety of exercise types and through a variety of methods.

Architecture and inherent robustness of a bacterial cell-cycle control system.

PubMed

Shen, Xiling; Collier, Justine; Dill, David; Shapiro, Lucy; Horowitz, Mark; McAdams, Harley H

2008-08-12

A closed-loop control system drives progression of the coupled stalked and swarmer cell cycles of the bacterium Caulobacter crescentus in a near-mechanical step-like fashion. The cell-cycle control has a cyclical genetic circuit composed of four regulatory proteins with tight coupling to processive chromosome replication and cell division subsystems. We report a hybrid simulation of the coupled cell-cycle control system, including asymmetric cell division and responses to external starvation signals, that replicates mRNA and protein concentration patterns and is consistent with observed mutant phenotypes. An asynchronous sequential digital circuit model equivalent to the validated simulation model was created. Formal model-checking analysis of the digital circuit showed that the cell-cycle control is robust to intrinsic stochastic variations in reaction rates and nutrient supply, and that it reliably stops and restarts to accommodate nutrient starvation. Model checking also showed that mechanisms involving methylation-state changes in regulatory promoter regions during DNA replication increase the robustness of the cell-cycle control. The hybrid cell-cycle simulation implementation is inherently extensible and provides a promising approach for development of whole-cell behavioral models that can replicate the observed functionality of the cell and its responses to changing environmental conditions.

Modular Battery Charge Controller

NASA Technical Reports Server (NTRS)

Button, Robert; Gonzalez, Marcelo

2009-01-01

A new approach to masterless, distributed, digital-charge control for batteries requiring charge control has been developed and implemented. This approach is required in battery chemistries that need cell-level charge control for safety and is characterized by the use of one controller per cell, resulting in redundant sensors for critical components, such as voltage, temperature, and current. The charge controllers in a given battery interact in a masterless fashion for the purpose of cell balancing, charge control, and state-of-charge estimation. This makes the battery system invariably fault-tolerant. The solution to the single-fault failure, due to the use of a single charge controller (CC), was solved by implementing one CC per cell and linking them via an isolated communication bus [e.g., controller area network (CAN)] in a masterless fashion so that the failure of one or more CCs will not impact the remaining functional CCs. Each micro-controller-based CC digitizes the cell voltage (V(sub cell)), two cell temperatures, and the voltage across the switch (V); the latter variable is used in conjunction with V(sub cell) to estimate the bypass current for a given bypass resistor. Furthermore, CC1 digitizes the battery current (I1) and battery voltage (V(sub batt) and CC5 digitizes a second battery current (I2). As a result, redundant readings are taken for temperature, battery current, and battery voltage through the summation of the individual cell voltages given that each CC knows the voltage of the other cells. For the purpose of cell balancing, each CC periodically and independently transmits its cell voltage and stores the received cell voltage of the other cells in an array. The position in the array depends on the identifier (ID) of the transmitting CC. After eight cell voltage receptions, the array is checked to see if one or more cells did not transmit. If one or more transmissions are missing, the missing cell(s) is (are) eliminated from cell-balancing calculations. The cell-balancing algorithm is based on the error between the cell s voltage and the other cells and is categorized into four zones of operation. The algorithm is executed every second and, if cell balancing is activated, the error variable is set to a negative low value. The largest error between the cell and the other cells is found and the zone of operation determined. If the error is zero or negative, then the cell is at the lowest voltage and no balancing action is needed. If the error is less than a predetermined negative value, a Cell Bad Flag is set. If the error is positive, then cell balancing is needed, but a hysteretic zone is added to prevent the bypass circuit from triggering repeatedly near zero error. This approach keeps the cells within a predetermined voltage range.

Oxygen-controlled automated neural differentiation of mouse embryonic stem cells.

PubMed

Mondragon-Teran, Paul; Tostoes, Rui; Mason, Chris; Lye, Gary J; Veraitch, Farlan S

2013-03-01

Automation and oxygen tension control are two tools that provide significant improvements to the reproducibility and efficiency of stem cell production processes. the aim of this study was to establish a novel automation platform capable of controlling oxygen tension during both the cell-culture and liquid-handling steps of neural differentiation processes. We built a bespoke automation platform, which enclosed a liquid-handling platform in a sterile, oxygen-controlled environment. An airtight connection was used to transfer cell culture plates to and from an automated oxygen-controlled incubator. Our results demonstrate that our system yielded comparable cell numbers, viabilities, metabolism profiles and differentiation efficiencies when compared with traditional manual processes. Interestingly, eliminating exposure to ambient conditions during the liquid-handling stage resulted in significant improvements in the yield of MAP2-positive neural cells, indicating that this level of control can improve differentiation processes. This article describes, for the first time, an automation platform capable of maintaining oxygen tension control during both the cell-culture and liquid-handling stages of a 2D embryonic stem cell differentiation process.

A correlate of HIV-1 control consisting of both innate and adaptive immune parameters best predicts viral load by multivariable analysis in HIV-1 infected viremic controllers and chronically-infected non-controllers.

PubMed

Tomescu, Costin; Liu, Qin; Ross, Brian N; Yin, Xiangfan; Lynn, Kenneth; Mounzer, Karam C; Kostman, Jay R; Montaner, Luis J

2014-01-01

HIV-1 infected viremic controllers maintain durable viral suppression below 2000 copies viral RNA/ml without anti-retroviral therapy (ART), and the immunological factor(s) associated with host control in presence of low but detectable viral replication are of considerable interest. Here, we utilized a multivariable analysis to identify which innate and adaptive immune parameters best correlated with viral control utilizing a cohort of viremic controllers (median 704 viral RNA/ml) and non-controllers (median 21,932 viral RNA/ml) that were matched for similar CD4+ T cell counts in the absence of ART. We observed that HIV-1 Gag-specific CD8+ T cell responses were preferentially targeted over Pol-specific responses in viremic controllers (p = 0.0137), while Pol-specific responses were positively associated with viral load (rho = 0.7753, p = 0.0001, n = 23). Viremic controllers exhibited significantly higher NK and plasmacytoid dendritic cells (pDC) frequency as well as retained expression of the NK CD16 receptor and strong target cell-induced NK cell IFN-gamma production compared to non-controllers (p<0.05). Despite differences in innate and adaptive immune function however, both viremic controllers (p<0.05) and non-controller subjects (p<0.001) exhibited significantly increased CD8+ T cell activation and spontaneous NK cell degranulation compared to uninfected donors. Overall, we identified that a combination of innate (pDC frequency) and adaptive (Pol-specific CD8+ T cell responses) immune parameters best predicted viral load (R2 = 0.5864, p = 0.0021, n = 17) by a multivariable analysis. Together, this data indicates that preferential Gag-specific over Pol-specific CD8+ T cell responses along with a retention of functional innate subsets best predict host control over viral replication in HIV-1 infected viremic controllers compared to chronically-infected non-controllers.

Enhanced expression of PD-1 and other activation markers by CD4+ T cells of young but not old patients with metastatic melanoma.

PubMed

van den Brom, Rob R H; van der Geest, Kornelis S M; Brouwer, Elisabeth; Hospers, Geke A P; Boots, Annemieke M H

2018-06-01

The biological behavior of melanoma is unfavorable in the elderly when compared to young subjects. We hypothesized that differences in T-cell responses might underlie the distinct behavior of melanoma in young and old melanoma patients. Therefore, we investigated the circulating T-cell compartment of 34 patients with metastatic melanoma and 42 controls, which were classified as either young or old. Absolute numbers of CD4+ T cells were decreased in young and old melanoma patients when compared to the age-matched control groups. Percentages of naive and memory CD4+ T cells were not different when comparing old melanoma patients to age-matched controls. Percentages of memory CD4+ T cells tended to be increased in young melanoma patients compared to young controls. Proportions of naive CD4+ T cells were lower in young patients than in age-matched controls, and actually comparable to those in old patients and controls. This was accompanied with increased percentages of memory CD4+ T cells expressing HLA-DR, Ki-67, and PD-1 in young melanoma patients in comparison to the age-matched controls, but not in old patients. Proportions of CD45RA-FOXP3 high memory regulatory T cells were increased in young and old melanoma patients when compared to their age-matched controls, whereas those of CD45RA+FOXP3 low naive regulatory T cells were similar. We observed no clear modulation of the circulating CD8+ T-cell repertoire in melanoma patients. In conclusion, we show that CD4+ T cells of young melanoma patients show signs of activation, whereas these signs are less clear in CD4+ T cells of old patients.

Outside-in control -Does plant cell wall integrity regulate cell cycle progression?

PubMed

Gigli-Bisceglia, Nora; Hamann, Thorsten

2018-04-13

During recent years it has become accepted that plant cell walls are not inert objects surrounding all plant cells but are instead highly dynamic, plastic structures. They are involved in a large number of cell biological processes and contribute actively to plant growth, development and interaction with environment. Therefore, it is not surprising that cellular processes can control plant cell wall integrity while, simultaneously, cell wall integrity can influence cellular processes. In yeast and animal cells such a bi-directional relationship also exists between the yeast/animal extra-cellular matrices and the cell cycle. In yeast, the cell wall integrity maintenance mechanism and a dedicated plasmamembrane integrity checkpoint are mediating this relationship. Recent research has yielded insights into the mechanism controlling plant cell wall metabolism during cytokinesis. However, knowledge regarding putative regulatory pathways controlling adaptive modifications in plant cell cycle activity in response to changes in the state of the plant cell wall are not yet identified. In this review, we summarize similarities and differences in regulatory mechanisms coordinating extra cellular matrices and cell cycle activity in animal and yeast cells, discuss the available evidence supporting the existence of such a mechanism in plants and suggest that the plant cell wall integrity maintenance mechanism might also control cell cycle activity in plant cells. This article is protected by copyright. All rights reserved.

Leaf shape: genetic controls and environmental factors.

PubMed

Tsukaya, Hirokazu

2005-01-01

In recent years, many genes have been identified that are involved in the developmental processes of leaf morphogenesis. Here, I review the mechanisms of leaf shape control in a model plant, Arabidopsis thaliana, focusing on genes that fulfill special roles in leaf development. The lateral, two-dimensional expansion of leaf blades is highly dependent on the determination of the dorsoventrality of the primordia, a defining characteristic of leaves. Having a determinate fate is also a characteristic feature of leaves and is controlled by many factors. Lateral expansion is not only controlled by general regulators of cell cycling, but also by the multi-level regulation of meristematic activities, e.g., specific control of cell proliferation in the leaf-length direction, in leaf margins and in parenchymatous cells. In collaboration with the polarized control of leaf cell elongation, these redundant and specialized regulating systems for cell cycling in leaf lamina may realize the elegantly smooth, flat structure of leaves. The unified, flat shape of leaves is also dependent on the fine integration of cell proliferation and cell enlargement. Interestingly, while a decrease in the number of cells in leaf primordia can trigger a cell volume increase, an increase in the number of cells does not trigger a cell volume decrease. This phenomenon is termed compensation and suggests the existence of some systems for integration between cell cycling and cell enlargement in leaf primordia via cell-cell communication. The environmental adjustment of leaf expansion to light conditions and gravity is also summarized.

Decorating an individual living cell with a shell of controllable thickness by cytocompatible surface initiated graft polymerization.

PubMed

Wang, Guan; Zhang, Kai; Wang, Yindian; Zhao, Changwen; He, Bin; Ma, Yuhong; Yang, Wantai

2018-05-03

Surface engineering of individual living cells is a promising field for cell-based applications. However, engineering individual cells with controllable thickness by chemical methods has been rarely studied. This article describes the development of a new cytocompatible chemical strategy to decorate individual living cells. The thicknesses of the crosslinked shells could be conveniently controlled by the irradiation time, visible light intensity, or monomer concentration. Moreover, the lag phase of the yeast cell division was extended and their stability against lysis was improved, which could also be tuned by controlling the shell thickness.

Micromechanical Devices for Control of Cell-Cell Interaction, and Methods of Use Thereof

NASA Technical Reports Server (NTRS)

Bhatia, Sangeeta N. (Inventor); Hui, Elliot (Inventor)

2017-01-01

The development and function of living tissues depends largely on interactions between cells that can vary in both time and space; however, temporal control of cell-cell interaction is experimentally challenging. By employing a micromachined silicon substrate with moving parts, herein is disclosed the dynamic regulation of cell-cell interactions via direct manipulation of adherent cells with micron-scale precision. The inventive devices and methods allow mechanical control of both tissue composition and spatial organization. The inventive device and methods enable the investigation of dynamic cell-cell interaction in a multitude of applications, such as intercellular communication, spanning embryogenesis, homeostasis, and pathogenic processes.

Functionalized iron oxide nanoparticles for controlling the movement of immune cells

NASA Astrophysics Data System (ADS)

White, Ethan E.; Pai, Alex; Weng, Yiming; Suresh, Anil K.; van Haute, Desiree; Pailevanian, Torkom; Alizadeh, Darya; Hajimiri, Ali; Badie, Behnam; Berlin, Jacob M.

2015-04-01

Immunotherapy is currently being investigated for the treatment of many diseases, including cancer. The ability to control the location of immune cells during or following activation would represent a powerful new technique for this field. Targeted magnetic delivery is emerging as a technique for controlling cell movement and localization. Here we show that this technique can be extended to microglia, the primary phagocytic immune cells in the central nervous system. The magnetized microglia were generated by loading the cells with iron oxide nanoparticles functionalized with CpG oligonucleotides, serving as a proof of principle that nanoparticles can be used to both deliver an immunostimulatory cargo to cells and to control the movement of the cells. The nanoparticle-oligonucleotide conjugates are efficiently internalized, non-toxic, and immunostimulatory. We demonstrate that the in vitro migration of the adherent, loaded microglia can be controlled by an external magnetic field and that magnetically-induced migration is non-cytotoxic. In order to capture video of this magnetically-induced migration of loaded cells, a novel 3D-printed ``cell box'' was designed to facilitate our imaging application. Analysis of cell movement velocities clearly demonstrate increased cell velocities toward the magnet. These studies represent the initial step towards our final goal of using nanoparticles to both activate immune cells and to control their trafficking within the diseased brain.Immunotherapy is currently being investigated for the treatment of many diseases, including cancer. The ability to control the location of immune cells during or following activation would represent a powerful new technique for this field. Targeted magnetic delivery is emerging as a technique for controlling cell movement and localization. Here we show that this technique can be extended to microglia, the primary phagocytic immune cells in the central nervous system. The magnetized microglia were generated by loading the cells with iron oxide nanoparticles functionalized with CpG oligonucleotides, serving as a proof of principle that nanoparticles can be used to both deliver an immunostimulatory cargo to cells and to control the movement of the cells. The nanoparticle-oligonucleotide conjugates are efficiently internalized, non-toxic, and immunostimulatory. We demonstrate that the in vitro migration of the adherent, loaded microglia can be controlled by an external magnetic field and that magnetically-induced migration is non-cytotoxic. In order to capture video of this magnetically-induced migration of loaded cells, a novel 3D-printed ``cell box'' was designed to facilitate our imaging application. Analysis of cell movement velocities clearly demonstrate increased cell velocities toward the magnet. These studies represent the initial step towards our final goal of using nanoparticles to both activate immune cells and to control their trafficking within the diseased brain. Electronic supplementary information (ESI) available: Transmission electron microscopy images of the particles, additional independent experiments for the NFκB activity and exocytosis assays, TEM images for the SPION untreated cells, bright field microscopy images of the cells alone in the presence and absence of magnet, images of the magnetic movement experiments at higher doses of SPION, full uncropped images of the post-migration LIVE/DEAD assay, and a video file of cell movement. See DOI: 10.1039/c3nr04421a

Quantifying intrinsic and extrinsic control of single-cell fates in cancer and stem/progenitor cell pedigrees with competing risks analysis

PubMed Central

Cornwell, J. A.; Hallett, R. M.; der Mauer, S. Auf; Motazedian, A.; Schroeder, T.; Draper, J. S.; Harvey, R. P.; Nordon, R. E.

2016-01-01

The molecular control of cell fate and behaviour is a central theme in biology. Inherent heterogeneity within cell populations requires that control of cell fate is studied at the single-cell level. Time-lapse imaging and single-cell tracking are powerful technologies for acquiring cell lifetime data, allowing quantification of how cell-intrinsic and extrinsic factors control single-cell fates over time. However, cell lifetime data contain complex features. Competing cell fates, censoring, and the possible inter-dependence of competing fates, currently present challenges to modelling cell lifetime data. Thus far such features are largely ignored, resulting in loss of data and introducing a source of bias. Here we show that competing risks and concordance statistics, previously applied to clinical data and the study of genetic influences on life events in twins, respectively, can be used to quantify intrinsic and extrinsic control of single-cell fates. Using these statistics we demonstrate that 1) breast cancer cell fate after chemotherapy is dependent on p53 genotype; 2) granulocyte macrophage progenitors and their differentiated progeny have concordant fates; and 3) cytokines promote self-renewal of cardiac mesenchymal stem cells by symmetric divisions. Therefore, competing risks and concordance statistics provide a robust and unbiased approach for evaluating hypotheses at the single-cell level. PMID:27250534

Simplified Load-Following Control for a Fuel Cell System

NASA Technical Reports Server (NTRS)

Vasquez, Arturo

2010-01-01

A simplified load-following control scheme has been proposed for a fuel cell power system. The scheme could be used to control devices that are important parts of a fuel cell system but are sometimes characterized as parasitic because they consume some of the power generated by the fuel cells.

T-cell-dependent control of acute Giardia lamblia infections in mice.

PubMed

Singer, S M; Nash, T E

2000-01-01

We have studied immune mechanisms responsible for control of acute Giardia lamblia and Giardia muris infections in adult mice. Association of chronic G. lamblia infection with hypogammaglobulinemia and experimental infections of mice with G. muris have led to the hypothesis that antibodies are required to control these infections. We directly tested this hypothesis by infecting B-cell-deficient mice with either G. lamblia or G. muris. Both wild-type mice and B-cell-deficient mice eliminated the vast majority of parasites between 1 and 2 weeks postinfection with G. lamblia. G. muris was also eliminated in both wild-type and B-cell-deficient mice. In contrast, T-cell-deficient and scid mice failed to control G. lamblia infections, as has been shown previously for G. muris. Treatment of wild-type or B-cell-deficient mice with antibodies to CD4 also prevented elimination of G. lamblia, confirming a role for T cells in controlling infections. By infecting mice deficient in either alphabeta- or gammadelta-T-cell receptor (TCR)-expressing T cells, we show that the alphabeta-TCR-expressing T cells are required to control parasites but that the gammadelta-TCR-expressing T cells are not. Finally, infections in mice deficient in production of gamma interferon or interleukin 4 (IL-4) and mice deficient in responding to IL-4 and IL-13 revealed that neither the Th1 nor the Th2 subset is absolutely required for protection from G. lamblia. We conclude that a T-cell-dependent mechanism is essential for controlling acute Giardia infections and that this mechanism is independent of antibody and B cells.

Daughter-Specific Transcription Factors Regulate Cell Size Control in Budding Yeast

PubMed Central

Di Talia, Stefano; Wang, Hongyin; Skotheim, Jan M.; Rosebrock, Adam P.; Futcher, Bruce; Cross, Frederick R.

2009-01-01

In budding yeast, asymmetric cell division yields a larger mother and a smaller daughter cell, which transcribe different genes due to the daughter-specific transcription factors Ace2 and Ash1. Cell size control at the Start checkpoint has long been considered to be a main regulator of the length of the G1 phase of the cell cycle, resulting in longer G1 in the smaller daughter cells. Our recent data confirmed this concept using quantitative time-lapse microscopy. However, it has been proposed that daughter-specific, Ace2-dependent repression of expression of the G1 cyclin CLN3 had a dominant role in delaying daughters in G1. We wanted to reconcile these two divergent perspectives on the origin of long daughter G1 times. We quantified size control using single-cell time-lapse imaging of fluorescently labeled budding yeast, in the presence or absence of the daughter-specific transcriptional regulators Ace2 and Ash1. Ace2 and Ash1 are not required for efficient size control, but they shift the domain of efficient size control to larger cell size, thus increasing cell size requirement for Start in daughters. Microarray and chromatin immunoprecipitation experiments show that Ace2 and Ash1 are direct transcriptional regulators of the G1 cyclin gene CLN3. Quantification of cell size control in cells expressing titrated levels of Cln3 from ectopic promoters, and from cells with mutated Ace2 and Ash1 sites in the CLN3 promoter, showed that regulation of CLN3 expression by Ace2 and Ash1 can account for the differential regulation of Start in response to cell size in mothers and daughters. We show how daughter-specific transcriptional programs can interact with intrinsic cell size control to differentially regulate Start in mother and daughter cells. This work demonstrates mechanistically how asymmetric localization of cell fate determinants results in cell-type-specific regulation of the cell cycle. PMID:19841732

Evidence of functional cell-mediated immune responses to nontypeable Haemophilus influenzae in otitis-prone children

PubMed Central

Seppanen, Elke; Tan, Dino; Corscadden, Karli J.; Currie, Andrew J.; Richmond, Peter C.; Thornton, Ruth B.

2018-01-01

Otitis media (OM) remains a common paediatric disease, despite advances in vaccinology. Susceptibility to recurrent acute OM (rAOM) has been postulated to involve defective cell-mediated immune responses to common otopathogenic bacteria. We compared the composition of peripheral blood mononuclear cells (PBMC) from 20 children with a history of rAOM (otitis-prone) and 20 healthy non-otitis-prone controls, and assessed innate and cell-mediated immune responses to the major otopathogen nontypeable Haemophilus influenzae (NTHi). NTHi was a potent stimulator of inflammatory cytokine secretion from PBMC within 4 hours, with no difference in cytokine levels produced between PBMC from cases or controls. In the absence of antigen stimulation, otitis-prone children had more circulating Natural Killer (NK) cells (p<0.01), particularly NKdim (CD56lo) cells (p<0.01), but fewer CD4+ T cells (p<0.01) than healthy controls. NTHi challenge significantly increased the proportion of activated (CD107a+) NK cells in otitis-prone and non-otitis-prone children (p<0.01), suggesting that NK cells from otitis-prone children are functional and respond to NTHi. CD8+ T cells and NK cells from both cases and controls produced IFNγ in response to polyclonal stimulus (Staphylococcal enterotoxin B; SEB), with more IFNγ+ CD8+ T cells present in cases than controls (p<0.05) but similar proportions of IFNγ+ NK cells. Otitis-prone children had more circulating IFNγ-producing NK cells (p<0.05) and more IFNγ-producing CD4+ (p<0.01) or CD8+ T-cells (p<0.05) than healthy controls. In response to SEB, more CD107a-expressing CD8+ T cells were present in cases than controls (p<0.01). Despite differences in PBMC composition, PBMC from otitis-prone children mounted innate and T cell-mediated responses to NTHi challenge that were comparable to healthy children. These data provide evidence that otitis-prone children do not have impaired functional cell mediated immunity. PMID:29621281

Cell Patterning Chip for Controlling the Stem Cell Microenvironment

PubMed Central

Rosenthal, Adam; Macdonald, Alice; Voldman, Joel

2007-01-01

Cell-cell signaling is an important component of the stem cell microenvironment, affecting both differentiation and self-renewal. However, traditional cell-culture techniques do not provide precise control over cell-cell interactions, while existing cell patterning technologies are limited when used with proliferating or motile cells. To address these limitations, we created the Bio Flip Chip (BFC), a microfabricated polymer chip containing thousands of microwells, each sized to trap down to a single stem cell. We have demonstrated the functionality of the BFC by patterning a 50×50 grid of murine embryonic stem cells (mESCs), with patterning efficiencies > 75%, onto a variety of substrates – a cell-culture dish patterned with gelatin, a 3-D substrate, and even another layer of cells. We also used the BFC to pattern small groups of cells, with and without cell-cell contact, allowing incremental and independent control of contact-mediated signaling. We present quantitative evidence that cell-cell contact plays an important role in depressing mESC colony formation, and show that E-cadherin is involved in this negative regulatory pathway. Thus, by allowing exquisite control of the cellular microenvironment, we provide a technology that enables new applications in tissue engineering and regenerative medicine. PMID:17434582

Engineering cells with intracellular agent–loaded microparticles to control cell phenotype

PubMed Central

Ankrum, James A; Miranda, Oscar R; Ng, Kelvin S; Sarkar, Debanjan; Xu, Chenjie; Karp, Jeffrey M

2014-01-01

Cell therapies enable unprecedented treatment options to replace tissues, destroy tumors and facilitate regeneration. The greatest challenge facing cell therapy is the inability to control the fate and function of cells after transplantation. We have developed an approach to control cell phenotype in vitro and after transplantation by engineering cells with intracellular depots that continuously release phenotype-altering agents for days to weeks. The platform enables control of cells’ secretome, viability, proliferation and differentiation, and the platform can be used to deliver drugs or other factors (e.g., dexamethasone, rhodamine and iron oxide) to the cell’s microenvironment. The preparation, efficient internalization and intracellular stabilization of ~1-μm drug-loaded microparticles are critical for establishing sustained control of cell phenotype. Herein we provide a protocol to generate and characterize micrometer-sized agent-doped poly(lactic-co-glycolic) acid (PLGA) particles by using a single-emulsion evaporation technique (7 h), to uniformly engineer cultured cells (15 h), to confirm particle internalization and to troubleshoot commonly experienced obstacles. PMID:24407352

Clinical, virologic, and immunologic outcomes in lymphoma survivors and in cancer-free, HIV-1-infected patients: a matched cohort study.

PubMed

Spagnuolo, Vincenzo; Travi, Giovanna; Galli, Laura; Cossarini, Francesca; Guffanti, Monica; Gianotti, Nicola; Salpietro, Stefania; Lazzarin, Adriano; Castagna, Antonella

2013-08-01

The objective of this study was to compare immunologic, virologic, and clinical outcomes between living human immunodeficiency virus (HIV)-infected individuals who had a diagnosis of lymphoma versus outcomes in a control group of cancer-free, HIV-infected patients. In this matched cohort study, patients in the case group were survivors of incident lymphomas that occurred between 1997 and June 2010. Controls were living, cancer-free, HIV-infected patients who were matched to cases at a 4:1 ratio by age, sex, nadir CD4 cell count, and year of HIV diagnosis. The date of lymphoma diagnosis served as the baseline in cases and in the corresponding controls. In total, 62 patients (cases) who had lymphoma (20 with Hodgkin disease [HD] and 42 with non-Hodgkin lymphoma [NHL]) were compared with 211 controls. The overall median follow-up was 4.8 years (interquartile range, 2.0-7.9 years). The CD4 cell count at baseline was 278 cells/mm³ (interquartile range, 122-419 cells/mm³) in cases versus 421 cells/mm³ (interquartile range, 222-574 cells/mm³) in controls (P = .003). At the last available visit, the CD4 cell count was 412 cells/mm³ (range, 269-694 cells/mm³) in cases versus 518 cells/mm³ (interquartile range, 350-661 cells/mm³) in controls (P = .087). The proportion of patients who achieved virologic success increased from 30% at baseline to 74% at the last available visit in cases (P = .008) and from 51% to 81% in controls (P = .0286). Patients with HD reached higher CD4 cell counts at their last visit than patients with NHL (589 cells/mm³ [range, 400-841 cells/mm³] vs 332 cells/mm³ [interquartile range, 220-530 cells/mm³], respectively; P = .003). Virologic success was similar between patients with HD and patients with NHL at the last visit. Forty cases (65%) and 76 controls (36%) experienced at least 1 clinical event after baseline (P < .0001); cases were associated with a shorter time to occurrence of the first clinical event compared with controls (P < .0001). HIV-infected lymphoma survivors experienced more clinical events than controls, especially during the first year of follow-up, but they reached similar long-term immunologic and virologic outcomes. © 2013 American Cancer Society.

Successful slush nitrogen vitrification of human ovarian tissue.

PubMed

Talevi, Riccardo; Barbato, Vincenza; Fiorentino, Ilaria; Braun, Sabrina; De Stefano, Cristofaro; Ferraro, Raffaele; Sudhakaran, Sam; Gualtieri, Roberto

2016-06-01

To study whether slush nitrogen vitrification improves the preservation of human ovarian tissue. Control vs. treatment study. University research laboratory. Ovarian biopsies collected from nine women (aged 14-35 years) during laparoscopic surgery for benign gynecologic conditions. None. Ovarian cortical strips of 2 × 5 × 1 mm were vitrified with liquid or slush nitrogen. Fresh and vitrified cortical strips were analyzed for cryodamage and viability under light, confocal, and transmission electron microscopy. Compared with liquid nitrogen, vitrification with slush nitrogen preserves [1] follicle quality (grade 1 follicles: fresh control, 50%; liquid nitrogen, 27%; slush nitrogen, 48%); [2] granulosa cell ultrastructure (intact cells: fresh control, 92%; liquid nitrogen, 45%; slush nitrogen, 73%), stromal cell ultrastructure (intact cells: fresh control, 59.8%; liquid nitrogen, 24%; slush nitrogen, 48.7%), and DNA integrity (TUNEL-positive cells: fresh control, 0.5%; liquid nitrogen, 2.3%; slush nitrogen, 0.4%); and [3] oocyte, granulosa, and stromal cell viability (oocyte: fresh control, 90%; liquid nitrogen, 63%; slush nitrogen, 87%; granulosa cells: fresh control, 93%; liquid nitrogen, 53%; slush nitrogen, 81%; stromal cells: fresh control, 63%; liquid nitrogen, 30%; slush nitrogen, 52%). The histology, ultrastructure, and viability of follicles and stromal cells are better preserved after vitrification with slush nitrogen compared with liquid nitrogen. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Optimization and control of perfusion cultures using a viable cell probe and cell specific perfusion rates.

PubMed

Dowd, Jason E; Jubb, Anthea; Kwok, K Ezra; Piret, James M

2003-05-01

Consistent perfusion culture production requires reliable cell retention and control of feed rates. An on-line cell probe based on capacitance was used to assay viable biomass concentrations. A constant cell specific perfusion rate controlled medium feed rates with a bioreactor cell concentration of approximately 5 x 10(6) cells mL(-1). Perfusion feeding was automatically adjusted based on the cell concentration signal from the on-line biomass sensor. Cell specific perfusion rates were varied over a range of 0.05 to 0.4 nL cell(-1) day(-1). Pseudo-steady-state bioreactor indices (concentrations, cellular rates and yields) were correlated to cell specific perfusion rates investigated to maximize recombinant protein production from a Chinese hamster ovary cell line. The tissue-type plasminogen activator concentration was maximized ( approximately 40 mg L(-1)) at 0.2 nL cell(-1) day(-1). The volumetric protein productivity ( approximately 60 mg L(-1) day(-1) was maximized above 0.3 nL cell(-1) day(-1). The use of cell specific perfusion rates provided a straightforward basis for controlling, modeling and optimizing perfusion cultures.

Nanopipette Apparatus for Manipulating Cells

NASA Technical Reports Server (NTRS)

Vilozny, Boaz (Inventor); Seger, R. Adam (Inventor); Actis, Paolo (Inventor); Pourmand, Nader (Inventor)

2017-01-01

Disclosed herein are methods and systems for controlled ejection of desired material onto surfaces including in single cells using nanopipettes, as well as ejection onto and into cells. Some embodiments are directed to a method and system comprising nanopipettes combined with an xyz controller for depositing a user defined pattern on an arbitrary substrate for the purpose of controlled cell adhesion and growth. Alternate embodiments are directed to a method and system comprising nanopipettes combined with an xyz controller and electronic control of a voltage differential in a bore of the nanopipette electroosmotically injecting material into a cell in a high-throughput manner and with minimal damage to the cell. Yet other embodiments are directed to method and system comprising functionalized nanopipettes combined with scanning ion conductance microscopy for studying molecular interactions and detection of biomolecules inside a single living cell.

11. ENGINE TEST CELL BUILDING INTERIOR. CONTROL ROOM FOR CELLS ...

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

11. ENGINE TEST CELL BUILDING INTERIOR. CONTROL ROOM FOR CELLS 2 AND 4. LOOKING SOUTHEAST. - Fairchild Air Force Base, Engine Test Cell Building, Near intersection of Arnold Street & George Avenue, Spokane, Spokane County, WA

A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes without Growth Factor Stimulation

DTIC Science & Technology

2011-01-01

A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes Without Growth Factor Stimulation...Ph.D.3 This work describes the differentiation of adipose-derived mesenchymal stem cells (ASC) in a composite hy- drogel for use as a vascularized...tissue from a single population of ASC. This work underscores the importance of the extracellular matrix in controlling stem cell phenotype. It is our

Functionalized Iron Oxide Nanoparticles for Controlling the Movement of Immune Cells

PubMed Central

White, Ethan E; Pai, Alex; Weng, Yiming; Suresh, Anil K.; Van Haute, Desiree; Pailevanian, Torkom; Alizadeh, Darya; Hajimiri, Ali; Badie, Behnam; Berlin, Jacob M.

2015-01-01

Immunotherapy is currently being investigated for the treatment of many diseases, including cancer. The ability to control the location of immune cells during or following activation would represent a powerful new technique for this field. Targeted magnetic delivery is emerging as a technique for controlling cell movement and localization. Here we show that this technique can be extended to microglia, the primary phagocytic immune cells in the central nervous system. The magnetized microglia were generated by loading the cells with iron oxide nanoparticles functionalized with CpG oligonucleotides, serving as a proof of principle that nanoparticles can be used to both deliver an immunostimulatory cargo to cells and to control the movement of the cells. The nanoparticle-oligonucleotide conjugates are efficiently internalized, non-toxic, and immunostimulatory. We demonstrate that the in vitro migration of the adherent, loaded microglia can be controlled by an external magnetic field and that magnetically-induced migration is non-cytotoxic. In order to capture video of this magnetically-induced migration of loaded cells, a novel 3D-printed “cell box” was designed to facilitate our imaging application. Analysis of cell movement velocities clearly demonstrate increased cell velocities toward the magnet. These studies represent the initial step towards our final goal of using nanoparticles to both activate immune cells and to control their trafficking within the diseased brain. PMID:25848983

Functionalized iron oxide nanoparticles for controlling the movement of immune cells.

PubMed

White, Ethan E; Pai, Alex; Weng, Yiming; Suresh, Anil K; Van Haute, Desiree; Pailevanian, Torkom; Alizadeh, Darya; Hajimiri, Ali; Badie, Behnam; Berlin, Jacob M

2015-05-07

Immunotherapy is currently being investigated for the treatment of many diseases, including cancer. The ability to control the location of immune cells during or following activation would represent a powerful new technique for this field. Targeted magnetic delivery is emerging as a technique for controlling cell movement and localization. Here we show that this technique can be extended to microglia, the primary phagocytic immune cells in the central nervous system. The magnetized microglia were generated by loading the cells with iron oxide nanoparticles functionalized with CpG oligonucleotides, serving as a proof of principle that nanoparticles can be used to both deliver an immunostimulatory cargo to cells and to control the movement of the cells. The nanoparticle-oligonucleotide conjugates are efficiently internalized, non-toxic, and immunostimulatory. We demonstrate that the in vitro migration of the adherent, loaded microglia can be controlled by an external magnetic field and that magnetically-induced migration is non-cytotoxic. In order to capture video of this magnetically-induced migration of loaded cells, a novel 3D-printed "cell box" was designed to facilitate our imaging application. Analysis of cell movement velocities clearly demonstrate increased cell velocities toward the magnet. These studies represent the initial step towards our final goal of using nanoparticles to both activate immune cells and to control their trafficking within the diseased brain.

Histone H3K9 Trimethylase Eggless Controls Germline Stem Cell Maintenance and Differentiation

PubMed Central

Zhou, Jian; McDowell, William; Park, Jungeun; Haug, Jeff; Staehling, Karen; Tang, Hong; Xie, Ting

2011-01-01

Epigenetic regulation plays critical roles in the regulation of cell proliferation, fate determination, and survival. It has been shown to control self-renewal and lineage differentiation of embryonic stem cells. However, epigenetic regulation of adult stem cell function remains poorly defined. Drosophila ovarian germline stem cells (GSCs) are a productive adult stem cell system for revealing regulatory mechanisms controlling self-renewal and differentiation. In this study, we show that Eggless (Egg), a H3K9 methyltransferase in Drosophila, is required in GSCs for controlling self-renewal and in escort cells for regulating germ cell differentiation. egg mutant ovaries primarily exhibit germ cell differentiation defects in young females and gradually lose GSCs with time, indicating that Egg regulates both germ cell maintenance and differentiation. Marked mutant egg GSCs lack expression of trimethylated H3K9 (H3k9me3) and are rapidly lost from the niche, but their mutant progeny can still differentiate into 16-cell cysts, indicating that Egg is required intrinsically to control GSC self-renewal but not differentiation. Interestingly, BMP-mediated transcriptional repression of differentiation factor bam in marked egg mutant GSCs remains normal, indicating that Egg is dispensable for BMP signaling in GSCs. Normally, Bam and Bgcn interact with each other to promote GSC differentiation. Interestingly, marked double mutant egg bgcn GSCs are still lost, but their progeny are able to differentiate into 16-cell cysts though bgcn mutant GSCs normally do not differentiate, indicating that Egg intrinsically controls GSC self-renewal through repressing a Bam/Bgcn-independent pathway. Surprisingly, RNAi-mediated egg knockdown in escort cells leads to their gradual loss and a germ cell differentiation defect. The germ cell differentiation defect is at least in part attributed to an increase in BMP signaling in the germ cell differentiation niche. Therefore, this study has revealed the essential roles of histone H3K9 trimethylation in controlling stem cell maintenance and differentiation through distinct mechanisms. PMID:22216012

Remote Control of Tissue Interactions via Engineered Photo-switchable Cell Surfaces

NASA Astrophysics Data System (ADS)

Luo, Wei; Pulsipher, Abigail; Dutta, Debjit; Lamb, Brian M.; Yousaf, Muhammad N.

2014-09-01

We report a general cell surface molecular engineering strategy via liposome fusion delivery to create a dual photo-active and bio-orthogonal cell surface for remote controlled spatial and temporal manipulation of microtissue assembly and disassembly. Cell surface tailoring of chemoselective functional groups was achieved by a liposome fusion delivery method and quantified by flow cytometry and characterized by a new cell surface lipid pull down mass spectrometry strategy. Dynamic co-culture spheroid tissue assembly in solution and co-culture tissue multilayer assembly on materials was demonstrated by an intercellular photo-oxime ligation that could be remotely cleaved and disassembled on demand. Spatial and temporal control of microtissue structures containing multiple cell types was demonstrated by the generation of patterned multilayers for controlling stem cell differentiation. Remote control of cell interactions via cell surface engineering that allows for real-time manipulation of tissue dynamics may provide tools with the scope to answer fundamental questions of cell communication and initiate new biotechnologies ranging from imaging probes to drug delivery vehicles to regenerative medicine, inexpensive bioreactor technology and tissue engineering therapies.

Overproduction of S-adenosylmethionine decarboxylase in ethylglyoxal-bis(guanylhydrazone)-resistant mouse FM3A cells.

PubMed

Suzuki, T; Sadakata, Y; Kashiwagi, K; Hoshino, K; Kakinuma, Y; Shirahata, A; Igarashi, K

1993-07-15

A variant cell line, termed SAM-1, which overproduced S-adenosylmethionine decarboxylase (AdoMetDC), was isolated by treatment of mouse FM3A cells with N-methyl-N'-nitro-N-nitrosoguanidine and subsequent incubation with ethylglyoxal bis(guanylhydrazone), an inhibitor of the enzyme. The cells were resistant to ethylglyoxal bis(guanylhydrazone), and showed AdoMetDC activity approximately five-times higher than control cells. The rate of AdoMetDC synthesis and the amount of AdoMetDC existing in SAM-1 cells were about five-times those in control cells. The amount of AdoMetDC mRNA existing in SAM-1 cells was five-times more than that in control cells. The amount of 5'-([(Z)-4-amino-2-butenyl]methylamino)-5'-deoxyadenosine, an irreversible inhibitor of AdoMetDC, necessary to inhibit cell growth was also five-times more in SAM-1 cells than in control cells. However, the following were the same in both SAM-1 and control cells; the amount of genomic DNA for AdoMetDC, the size and nucleotide sequence of 5' untranslated region of AdoMetDC mRNA, the deduced amino acid sequence (334 residues) from the nucleotide sequence of AdoMetDC cDNA and the degradation rate (t1/2 = about 4 h) of AdoMetDC. In addition, AdoMetDC mRNA in control cells was slightly more stable than that in SAM-1 cells. The results indicate that the overproduction of AdoMetDC in SAM-1 cells was caused by the increase of AdoMetDC mRNA. The variant cell line is convenient for studying the regulation of AdoMetDC and the physiological function of polyamines.

Developmental control of transcriptional and proliferative potency during the evolutionary emergence of animals

PubMed Central

Arenas-Mena, Cesar; Coffman, James A.

2016-01-01

Summary It is proposed that the evolution of complex animals required repressive genetic mechanisms for controlling the transcriptional and proliferative potency of cells. Unicellular organisms are transcriptionally potent, able to express their full genetic complement as the need arises through their life cycle, whereas differentiated cells of multicellular organisms can only express a fraction of their genomic potential. Likewise, whereas cell proliferation in unicellular organisms is primarily limited by nutrient availability, cell proliferation in multicellular organisms is developmentally regulated. Repressive genetic controls limiting the potency of cells at the end of ontogeny would have stabilized the gene expression states of differentiated cells and prevented disruptive proliferation, allowing the emergence of diverse cell types and functional shapes. We propose that distal cis-regulatory elements represent the primary innovations that set the stage for the evolution of developmental gene regulatory networks and the repressive control of key multipotency and cell-cycle control genes. The testable prediction of this model is that the genomes of extant animals, unlike those of our unicellular relatives, encode gene regulatory circuits dedicated to the developmental control of transcriptional and proliferative potency. PMID:26173445

Inhibition of the expression of aquaporin‑1 by RNA interference in pulmonary epithelial cells and its effects on water transport.

PubMed

Zhang, Qiuyue; Fu, Jianhua; Xue, Xindong

2016-01-01

In the present study, the effect of aquaporin‑1 (AQP1) on fluid transportation in pulmonary epithelial cells, and the role of AQP1 in alveolar fluid clearance were investigated to provide an experimental foundation to elucidate the pathogenesis of hyperoxic lung edema. An siRNA transfection technique was used to silence AQP1 in the A549 cell line. The transfected cells were randomized into a hyperoxia exposure and an air control group, with a negative control group set for each group. Cell volume was determined using flow cytometry, and Pf values were used to determine osmotic water permeability. Cell volume was found to be reduced in the AQP1‑silenced A549 cells, compared with the negative control group 72 h following air exposure. In addition, cell volume was reduced in the AQP1‑silenced A549 cells, compared with the negative control group 48 and 72 h following hyperoxia exposure. The osmotic water permeability of the AQP1‑silenced cells was reduced in the air control and hyperoxia exposure groups, compared with the negative control group 48 and 72 h following exposure. The volume and cell membrane osmotic water permeability of the A549 cells were reduced, compared with those in the control group following AQP1‑silencing, which indicated that the downregulation of AQP1 impedes extracellular to intracellular fluid transportation. Therefore, the disturbance in alveolar fluid clearance resulting from the downregulation of AQP1 following hyperoxia exposure may be one of the key mechanisms responsible for hyperoxic lung edema.

“Engineering Substrate Micro- and Nanotopography to Control Cell Function”

PubMed Central

Bettinger, Christopher J; Langer, Robert; Borenstein, Jeffrey T

2010-01-01

Lead-In The interaction of mammalian cells with nanoscale topography has proven to be an important signaling modality in controlling cell function. Naturally occurring nanotopographic structures within the extracellular matrix present surrounding cells with mechanotransductive cues that influence local migration, cell polarization, and other functions. Synthetically nanofabricated topography can also influence cell morphology, alignment, adhesion, migration, proliferation, and cytoskeleton organization. Here we review the use of in vitro synthetic cell-nanotopography interactions to control cell behavior and influence complex cellular processes including stem cell differentiation and tissue organization. Future challenges and opportunities in cell-nanotopography engineering will also be discussed including the elucidation of mechanisms and applications in tissue engineering. PMID:19492373

Protecting and rescuing the effectors: roles of differentiation and survival in the control of memory T cell development

PubMed Central

Kurtulus, Sema; Tripathi, Pulak; Hildeman, David A.

2013-01-01

Vaccines, arguably the single most important intervention in improving human health, have exploited the phenomenon of immunological memory. The elicitation of memory T cells is often an essential part of successful long-lived protective immunity. Our understanding of T cell memory has been greatly aided by the development of TCR Tg mice and MHC tetrameric staining reagents that have allowed the precise tracking of antigen-specific T cell responses. Indeed, following acute infection or immunization, naïve T cells undergo a massive expansion culminating in the generation of a robust effector T cell population. This peak effector response is relatively short-lived and, while most effector T cells die by apoptosis, some remain and develop into memory cells. Although the molecular mechanisms underlying this cell fate decision remain incompletely defined, substantial progress has been made, particularly with regards to CD8+ T cells. For example, the effector CD8+ T cells generated during a response are heterogeneous, consisting of cells with more or less potential to develop into full-fledged memory cells. Development of CD8+ T cell memory is regulated by the transcriptional programs that control the differentiation and survival of effector T cells. While the type of antigenic stimulation and level of inflammation control effector CD8+ T cell differentiation, availability of cytokines and their ability to control expression and function of Bcl-2 family members governs their survival. These distinct differentiation and survival programs may allow for finer therapeutic intervention to control both the quality and quantity of CD8+ T cell memory. Effector to memory transition of CD4+ T cells is less well characterized than CD8+ T cells, emerging details will be discussed. This review will focus on the recent progress made in our understanding of the mechanisms underlying the development of T cell memory with an emphasis on factors controlling survival of effector T cells. PMID:23346085

Control of proliferation and cancer growth by the Hippo signaling pathway

PubMed Central

Ehmer, Ursula; Sage, Julien

2015-01-01

The control of cell division is essential for normal development and the maintenance of cellular homeostasis. Abnormal cell proliferation is associated with multiple pathological states, including cancer. While the Hippo/YAP signaling pathway was initially thought to control organ size and growth, increasing evidence indicates that this pathway also plays a major role in the control of proliferation independent of organ size control. In particular, accumulating evidence indicates that the Hippo/YAP signaling pathway functionally interacts with multiple other cellular pathways and serves as a central node in the regulation of cell division, especially in cancer cells. Here recent observations are highlighted that connect Hippo/YAP signaling to transcription, the basic cell cycle machinery, and the control of cell division. Furthermore, the oncogenic and tumor suppressive attributes of YAP/TAZ are reviewed which emphasizes the relevance of the Hippo pathway in cancer. PMID:26432795

Regulation of cell arrangement using a novel composite micropattern.

PubMed

Liu, Xiaoyi; Liu, Yaoping; Zhao, Feng; Hun, Tingting; Li, Shan; Wang, Yuguang; Sun, Weijie; Wang, Wei; Sun, Yan; Fan, Yubo

2017-11-01

Micropatterning technique has been used to control single cell geometry in many researches, however, this is no report that it is used to control multicelluar geometry, which not only control single cell geometry but also organize those cells by a certain pattern. In this work, a composite protein micropattern is developed to control both cell shape and cell location simultaneously. The composite micropattern consists of a central circle 15 μm in diameter for single-cell capture, surrounded by small, square arrays (3 μm × 3 μm) for cell spreading. This is surrounded by a border 2 μm wide for restricting cell edges. The composite pattern results in two-cell and three-cell capture efficiencies of 32.1% ± 1.94% and 24.2% ± 2.89%, respectively, representing an 8.52% and 9.58% increase, respectively, over rates of original patterns. Fluorescent imaging of cytoskeleton alignment demonstrates that actin is gradually aligned parallel to the direction of the entire pattern arrangement, rather than to that of a single pattern. This indicates that cell arrangement is also an important factor in determining cell physiology. This composite micropattern could be a potential method to precisely control multi-cells for cell junctions, cell interactions, cell signal transduction, and eventually for tissue rebuilding study. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3093-3101, 2017. © 2017 Wiley Periodicals, Inc.

Progress in Aluminum Electrolysis Control and Future Direction for Smart Aluminum Electrolysis Plant

NASA Astrophysics Data System (ADS)

Zhang, Hongliang; Li, Tianshuang; Li, Jie; Yang, Shuai; Zou, Zhong

2017-02-01

The industrial aluminum reduction cell is an electrochemistry reactor that operates under high temperatures and highly corrosive conditions. However, these conditions have restricted the measurement of key control parameters, making the control of aluminum reduction cells a difficult problem in the industry. Because aluminum electrolysis control systems have a significant economic influence, substantial research has been conducted on control algorithms, control systems and information systems for aluminum reduction cells. This article first summarizes the development of control systems and then focuses on the progress made since 2000, including alumina concentration control, temperature control and electrolyte molecular ratio control, fault diagnosis, cell condition prediction and control system expansion. Based on these studies, the concept of a smart aluminum electrolysis plant is proposed. The frame construction, key problems and current progress are introduced. Finally, several future directions are discussed.

Myosin-II controls cellular branching morphogenesis and migration in 3D by minimizing cell surface curvature

PubMed Central

Elliott, Hunter; Fischer, Robert A.; Myers, Kenneth A.; Desai, Ravi A.; Gao, Lin; Chen, Christopher S.; Adelstein, Robert; Waterman, Clare M.; Danuser, Gaudenz

2014-01-01

In many cases cell function is intimately linked to cell shape control. We utilized endothelial cell branching morphogenesis as a model to understand the role of myosin-II in shape control of invasive cells migrating in 3D collagen gels. We applied principles of differential geometry and mathematical morphology to 3D image sets to parameterize cell branch structure and local cell surface curvature. We find that Rho/ROCK-stimulated myosin-II contractility minimizes cell-scale branching by recognizing and minimizing local cell surface curvature. Utilizing micro-fabrication to constrain cell shape identifies a positive feedback mechanism in which low curvature stabilizes myosin-II cortical association, where it acts to maintain minimal curvature. The feedback between myosin-II regulation by and control of curvature drives cycles of localized cortical myosin-II assembly and disassembly. These cycles in turn mediate alternating phases of directionally biased branch initiation and retraction to guide 3D cell migration. PMID:25621949

Regulation of Water in Plant Cells

ERIC Educational Resources Information Center

Kowles, Richard V.

2010-01-01

Cell water relationships are important topics to be included in cell biology courses. Differences exist in the control of water relationships in plant cells relative to control in animal cells. One important reason for these differences is that turgor pressure is a consideration in plant cells. Diffusion and osmosis are the underlying factors…

A Comparative Study of the ReCell® Device and Autologous Spit-thickness Meshed Skin Graft in the Treatment of Acute Burn Injuries.

PubMed

Holmes, J H; Molnar, J A; Carter, J E; Hwang, J; Cairns, B A; King, B T; Smith, D J; Cruse, C W; Foster, K N; Peck, M D; Sood, R; Feldman, M J; Jordan, M H; Mozingo, D W; Greenhalgh, D G; Palmieri, T L; Griswold, J A; Dissanaike, S; Hickerson, W L

2018-05-24

Early excision and autografting are standard care for deeper burns. However, donor sites are a source of significant morbidity. To address this, the ReCell® Autologous Cell Harvesting Device (ReCell) was designed for use at the point-of-care to prepare a non-cultured, autologous skin cell suspension (ASCS) capable of epidermal regeneration utilizing minimal donor skin. A prospective study was conducted to evaluate the clinical performance of ReCell versus meshed split-thickness skin grafts (STSG, Control) for the treatment of deep partial-thickness (DPT) burns. Effectiveness measures were assessed to 1 year for both ASCS and Control treatment sites and donor sites, including the incidence of healing, scarring, and pain. At 4 weeks, 98% of the ASCS-treated sites were healed compared to 100% of the Controls. Pain and assessments of scarring at the treatment sites were reported to be similar between groups. Significant differences were observed between ReCell and Control donor sites. The mean ReCell donor area was approximately 40 times smaller than that of the Control (194.1±158.5 cm2; p<0.0001), and after 1 week, significantly more ReCell donor sites were healed than Controls (p=0.04). Over the first 16 weeks, patients reported significantly less pain at the ReCell donor sites compared with Controls (p≤0.05 at each time point). Long-term, patients reported higher satisfaction with ReCell donor site outcomes compared with the Controls. This study provides evidence that the treatment of DPT burns with ASCS results in comparable healing, with significantly reduced donor site size and pain and improved appearance relative to STSG.

Analysis of Retinal Thinning Using Spectral-domain Optical Coherence Tomography Imaging of Sickle Cell Retinopathy Eyes Compared to Age- and Race-Matched Control Eyes.

PubMed

Lim, Jennifer I; Cao, Dingcai

2018-03-17

To determine whether the retina is thinner in sickle cell patients than in race- and age-matched controls, and, if it is thinner, whether there is any association with systemic diseases. Sickle cell and control (age- and race-matched) patients were prospectively enrolled from a university retina clinic into this observational study. Participants underwent visual acuity testing, slit-lamp biomicroscopy, dilated ophthalmoscopy, and spectral-domain optical coherence tomography imaging. Sickle cell retinal lesions, degree of vascular tortuosity, caliber of arteriovenous anastomosis, and stage of retinopathy were noted. Early Treatment Diabetic Retinopathy Study (ETDRS) subfield measurements were compared between sickle cell and control subjects and also among sickle cell hemoglobin subtypes. Associations between ETDRS subfield measurements and hemoglobin subtype, retinopathy stage, and systemic diseases were assessed. A total of 513 sickle cell eyes (260 patients) and 75 control eyes (39 patients) had median visual acuities of 20/20. ETDRS central (P = .002), inner (nasal P = .009, superior P = .021, temporal P < .001, inferior P = .017), and temporal outer (P = .012) subfield measurements were thinner in sickle cell eyes compared to control eyes. Hemoglobin SS eyes had significantly thinner inner ETDRS subfield measurements compared to SC and SThal eyes. Retinal thinning in all subfields was associated with age (P = .017) for sickle cell and control eyes. No association was found between retinal thinning and hydroxyurea use or arteriovenous anastomosis caliber. The macula is thinner in sickle cell eyes compared to control eyes; retinal thickness decreases with increasing age and sickle cell retinopathy stage and is most severe in hemoglobin SS subtypes. Copyright © 2018 Elsevier Inc. All rights reserved.

Dynamic Electrochemical Control of Cell Capture-and-Release Based on Redox-Controlled Host-Guest Interactions.

PubMed

Gao, Tao; Li, Liudi; Wang, Bei; Zhi, Jun; Xiang, Yang; Li, Genxi

2016-10-18

Artificial control of cell adhesion on smart surface is an on-demand technique in areas ranging from tissue engineering, stem cell differentiation, to the design of cell-based diagnostic system. In this paper, we report an electrochemical system for dynamic control of cell catch-and-release, which is based on the redox-controlled host-guest interaction. Experimental results reveal that the interaction between guest molecule (ferrocene, Fc) and host molecule (β-cyclodextrin, β-CD) is highly sensitive to electrochemical stimulus. By applying a reduction voltage, the uncharged Fc can bind to β-CD that is immobilized at the electrode surface. Otherwise, it is disassociated from the surface as a result of electrochemical oxidation, thus releasing the captured cells. The catch-and-release process on this voltage-responsive surface is noninvasive with the cell viability over 86%. Moreover, because Fc can act as an electrochemical probe for signal readout, the integration of this property has further extended the ability of this system to cell detection. Electrochemical signal has been greatly enhanced for cell detection by introducing branched polymer scaffold that are carrying large quantities of Fc moieties. Therefore, a minimum of 10 cells can be analyzed. It is anticipated that such redox-controlled system can be an important tool in biological and biomedical research, especially for electrochemical stimulated tissue engineering and cell-based clinical diagnosis.

High spatial and temporal resolution cell manipulation techniques in microchannels.

PubMed

Novo, Pedro; Dell'Aica, Margherita; Janasek, Dirk; Zahedi, René P

2016-03-21

The advent of microfluidics has enabled thorough control of cell manipulation experiments in so called lab on chips. Lab on chips foster the integration of actuation and detection systems, and require minute sample and reagent amounts. Typically employed microfluidic structures have similar dimensions as cells, enabling precise spatial and temporal control of individual cells and their local environments. Several strategies for high spatio-temporal control of cells in microfluidics have been reported in recent years, namely methods relying on careful design of the microfluidic structures (e.g. pinched flow), by integration of actuators (e.g. electrodes or magnets for dielectro-, acousto- and magneto-phoresis), or integrations thereof. This review presents the recent developments of cell experiments in microfluidics divided into two parts: an introduction to spatial control of cells in microchannels followed by special emphasis in the high temporal control of cell-stimulus reaction and quenching. In the end, the present state of the art is discussed in line with future perspectives and challenges for translating these devices into routine applications.

Friction-Controlled Traction Force in Cell Adhesion

PubMed Central

Pompe, Tilo; Kaufmann, Martin; Kasimir, Maria; Johne, Stephanie; Glorius, Stefan; Renner, Lars; Bobeth, Manfred; Pompe, Wolfgang; Werner, Carsten

2011-01-01

The force balance between the extracellular microenvironment and the intracellular cytoskeleton controls the cell fate. We report a new (to our knowledge) mechanism of receptor force control in cell adhesion originating from friction between cell adhesion ligands and the supporting substrate. Adherent human endothelial cells have been studied experimentally on polymer substrates noncovalently coated with fluorescent-labeled fibronectin (FN). The cellular traction force correlated with the mobility of FN during cell-driven FN fibrillogenesis. The experimental findings have been explained within a mechanistic two-dimensional model of the load transfer at focal adhesion sites. Myosin motor activity in conjunction with sliding of FN ligands noncovalently coupled to the surface of the polymer substrates is shown to result in a controlled traction force of adherent cells. We conclude that the friction of adhesion ligands on the supporting substrate is important for mechanotransduction and cell development of adherent cells in vitro and in vivo. PMID:22004739

B-cell subset alterations and correlated factors in HIV-1 infection.

PubMed

Pensieroso, Simone; Galli, Laura; Nozza, Silvia; Ruffin, Nicolas; Castagna, Antonella; Tambussi, Giuseppe; Hejdeman, Bo; Misciagna, Donatella; Riva, Agostino; Malnati, Mauro; Chiodi, Francesca; Scarlatti, Gabriella

2013-05-15

During HIV-1 infection, the development, phenotype, and functionality of B cells are impaired. Transitional B cells and aberrant B-cell populations arise in blood, whereas a declined percentage of resting memory B cells is detected. Our study aimed at pinpointing the demographic, immunological, and viral factors driving these pathological findings, and the role of antiretroviral therapy in reverting these alterations. B-cell phenotype and correlating factors were evaluated. Variations in B-cell subsets were evaluated by flow cytometry in HIV-1-infected individuals naive to therapy, elite controllers, and patients treated with antiretroviral drugs (virological control or failure). Multivariable analysis was performed to identify variables independently associated with the B-cell alterations. Significant differences were observed among patients' groups in relation to all B-cell subsets. Resting memory B cells were preserved in patients naive to therapy and elite controllers, but reduced in treated patients. Individuals naive to therapy and experiencing multidrug failure, as well as elite controllers, had significantly higher levels of activated memory B cells compared to healthy controls. In the multivariate analysis, plasma viral load and nadir CD4 T cells independently correlated with major B-cell alterations. Coinfection with hepatitis C but not hepatitis B virus also showed an impact on specific B-cell subsets. Successful protracted antiretroviral treatment led to normalization of all B-cell subsets with exception of resting memory B cells. Our results indicate that viremia and nadir CD4 T cells are important prognostic markers of B-cell perturbations and provide evidence that resting memory B-cell depletion during chronic infection is not reverted upon successful antiretroviral therapy.

Control assembly for controlling a fuel cell system during shutdown and restart

DOEpatents

Venkataraman, Ramki; Berntsen, George; Carlson, Glenn L.; Farooque, Mohammad; Beachy, Dan; Peterhans, Stefan; Bischoff, Manfred

2010-06-15

A fuel cell system and method in which the fuel cell system receives and an input oxidant gas and an input fuel gas, and in which a fuel processing assembly is provided and is adapted to at least humidify the input fuel gas which is to be supplied to the anode of the fuel cell of the system whose cathode receives the oxidant input gas via an anode oxidizing assembly which is adapted to couple the output of the anode of the fuel cell to the inlet of the cathode of the fuel cell during normal operation, shutdown and restart of the fuel cell system, and in which a control assembly is further provided and is adapted to respond to shutdown of the fuel cell system during which input fuel gas and input oxidant gas cease to be received by the fuel cell system, the control assembly being further adapted to, when the fuel cell system is shut down: control the fuel cell system so as to enable a purging gas to be able to flow through the fuel processing assembly to remove humidified fuel gas from the processing assembly and to enable a purging gas to be able to flow through the anode of the fuel cell.

The Adder Phenomenon Emerges from Independent Control of Pre- and Post-Start Phases of the Budding Yeast Cell Cycle.

PubMed

Chandler-Brown, Devon; Schmoller, Kurt M; Winetraub, Yonatan; Skotheim, Jan M

2017-09-25

Although it has long been clear that cells actively regulate their size, the molecular mechanisms underlying this regulation have remained poorly understood. In budding yeast, cell size primarily modulates the duration of the cell-division cycle by controlling the G1/S transition known as Start. We have recently shown that the rate of progression through Start increases with cell size, because cell growth dilutes the cell-cycle inhibitor Whi5 in G1. Recent phenomenological studies in yeast and bacteria have shown that these cells add an approximately constant volume during each complete cell cycle, independent of their size at birth. These results seem to be in conflict, as the phenomenological studies suggest that cells measure the amount they grow, rather than their size, and that size control acts over the whole cell cycle, rather than specifically in G1. Here, we propose an integrated model that unifies the adder phenomenology with the molecular mechanism of G1/S cell-size control. We use single-cell microscopy to parameterize a full cell-cycle model based on independent control of pre- and post-Start cell-cycle periods. We find that our model predicts the size-independent amount of cell growth during the full cell cycle. This suggests that the adder phenomenon is an emergent property of the independent regulation of pre- and post-Start cell-cycle periods rather than the consequence of an underlying molecular mechanism measuring a fixed amount of growth. Copyright © 2017 Elsevier Ltd. All rights reserved.

A High Frequency of HIV-Specific Circulating Follicular Helper T Cells Is Associated with Preserved Memory B Cell Responses in HIV Controllers.

PubMed

Claireaux, M; Galperin, M; Benati, D; Nouël, A; Mukhopadhyay, M; Klingler, J; de Truchis, P; Zucman, D; Hendou, S; Boufassa, F; Moog, C; Lambotte, O; Chakrabarti, L A

2018-05-08

Follicular helper T cells (Tfh) play an essential role in the affinity maturation of the antibody response by providing help to B cells. To determine whether this CD4 + T cell subset may contribute to the spontaneous control of HIV infection, we analyzed the phenotype and function of circulating Tfh (cTfh) in patients from the ANRS CO21 CODEX cohort who naturally controlled HIV-1 replication to undetectable levels and compared them to treated patients with similarly low viral loads. HIV-specific cTfh (Tet + ), detected by Gag-major histocompatibility complex class II (MHC-II) tetramer labeling in the CD45RA - CXCR5 + CD4 + T cell population, proved more frequent in the controller group ( P = 0.002). The frequency of PD-1 expression in Tet + cTfh was increased in both groups (median, >75%) compared to total cTfh (<30%), but the intensity of PD-1 expression per cell remained higher in the treated patient group ( P = 0.02), pointing to the persistence of abnormal immune activation in treated patients. The function of cTfh, analyzed by the capacity to promote IgG secretion in cocultures with autologous memory B cells, did not show major differences between groups in terms of total IgG production but proved significantly more efficient in the controller group when measuring HIV-specific IgG production. The frequency of Tet + cTfh correlated with HIV-specific IgG production ( R = 0.71 for Gag-specific and R = 0.79 for Env-specific IgG, respectively). Taken together, our findings indicate that key cTfh-B cell interactions are preserved in controlled HIV infection, resulting in potent memory B cell responses that may play an underappreciated role in HIV control. IMPORTANCE The rare patients who spontaneously control HIV replication in the absence of therapy provide a unique model to identify determinants of an effective anti-HIV immune response. HIV controllers show signs of particularly efficient antiviral T cell responses, while their humoral response was until recently considered to play only a minor role in viral control. However, emerging evidence suggests that HIV controllers maintain a significant but "silent" antiviral memory B cell population that can be reactivated upon antigenic stimulation. We report that cTfh help likely contributes to the persistence of controller memory B cell responses, as the frequency of HIV-specific cTfh correlated with the induction of HIV-specific antibodies in functional assays. These findings suggest that T follicular help may contribute to HIV control and highlight the need for inducing such help in HIV vaccine strategies that aim at eliciting persistent B cell responses. Copyright © 2018 Claireaux et al.

Review of microfluidic cell culture devices for the control of gaseous microenvironments in vitro

NASA Astrophysics Data System (ADS)

Wu, H.-M.; Lee, T.-A.; Ko, P.-L.; Chiang, H.-J.; Peng, C.-C.; Tung, Y.-C.

2018-04-01

Gaseous microenvironments play important roles in various biological activities in vivo. However, it is challenging to precisely control gaseous microenvironments in vitro for cell culture due to the high diffusivity nature of gases. In recent years, microfluidics has paved the way for the development of new types of cell culture devices capable of manipulating cellular microenvironments, and provides a powerful tool for in vitro cell studies. This paper reviews recent developments of microfluidic cell culture devices for the control of gaseous microenvironments, and discusses the advantages and limitations of current devices. We conclude with suggestions for the future development of microfluidic cell culture devices for the control of gaseous microenvironments.

Nanoscale definition of substrate materials to direct human adult stem cells towards tissue specific populations.

PubMed

Curran, Judith M; Chen, Rui; Stokes, Robert; Irvine, Eleanor; Graham, Duncan; Gubbins, Earl; Delaney, Deany; Amro, Nabil; Sanedrin, Raymond; Jamil, Haris; Hunt, John A

2010-03-01

The development of homogenously nano-patterned chemically modified surfaces that can be used to initiate a cellular response, particularly stem cell differentiation, in a highly controlled manner without the need for exogenous biological factors has never been reported, due to that fact that precisely defined and reproducible systems have not been available that can be used to study cell/material interactions and unlock the potential of a material driven cell response. Until now material driven stem cell (furthermore any cell) responses have been variable due to the limitations in definition and reproducibility of the underlying substrate and the lack of true homogeneity of modifications that can dictate a cellular response at a sub-micron level that can effectively control initial cell interactions of all cells that contact the surface. Here we report the successful design and use of homogenously molecularly nanopatterned surfaces to control initial stem cell adhesion and hence function. The highly specified nano-patterned arrays were compared directly to silane modified bulk coated substrates that have previously been proven to initiate mesenchymal stem cell (MSC) differentiation in a heterogenous manner, the aim of this study was to prove the efficiency of these previously observed cell responses could be enhanced by the incorporation of nano-patterns. Nano-patterned surfaces were prepared by Dip Pen Nanolithography (DPN) to produce arrays of 70 nm sized dots separated by defined spacings of 140, 280 and 1000 nm with terminal functionalities of carboxyl, amino, methyl and hydroxyl and used to control cell growth. These nanopatterned surfaces exhibited unprecedented control of initial cell interactions and will change the capabilities for stem cell definition in vitro and then cell based medical therapies. In addition to highlighting the ability of the materials to control stem cell functionality on an unprecedented scale this research also introduces the successful scale-up of DPN and the novel chemistries and systems to facilitate the production of homogeneously patterned substrates (5 mm2) that are applicable for use in in vitro cell conditions over prolonged periods for complete control of material driven cell responses.

Study on the Mechanism of Cell Cycle Checkpoint Kinase 2 (CHEK2) Gene Dysfunction in Chemotherapeutic Drug Resistance of Triple Negative Breast Cancer Cells.

PubMed

Luo, Li; Gao, Wei; Wang, Jinghui; Wang, Dingxue; Peng, Xiaobo; Jia, Zhaoyang; Jiang, Ye; Li, Gongzhuo; Tang, Dongxin; Wang, Yajie

2018-05-15

BACKGROUND This study aimed to investigate the mechanism of CHEK2 gene dysfunction in drug resistance of triple negative breast cancer (TNBC) cells. MATERIAL AND METHODS To perform our study, a stable CHEK2 wild type (CHEK2 WT) or CHEK2 Y390C mutation (CHEK2 Y390C) expressed MDA-MB-231 cell line was established. MTT assay, cell apoptosis assay and cell cycle assay were carried out to analyze the cell viability, apoptosis, and cell cycle respectively. Western blotting and qRT-PCR were applied for related protein and gene expression detection. RESULTS We found that the IC50 value of DDP (Cisplatin) to CHEK2 Y390C expressed MDA-MB-231 cells was significantly higher than that of the CHEK2 WT expressed cells and the control cells. After treatment with DDP for 48 h, cells expressing CHEK2 WT showed lower cell viability than that of the CHEK2 Y390C expressed cells and the control cells; compared with the CHEK2 Y390C expressed cells and the control cells, cells expressing CHEK2 WT showed significant G1/S arrest. Meanwhile, we found that compared with the CHEK2 Y390C expressed cells and the control cells, cell apoptosis was significantly increased in CHEK2 WT expressed cells. Moreover, our results suggested that cells expressing CHEK2 WT showed higher level of p-CDC25A, p-p53, p21, Bax, PUMA, and Noxa than that of the CHEK2 Y390C expressed cells and the control cells. CONCLUSIONS Our findings indicated that CHEK2 Y390C mutation induced the drug resistance of TNBC cells to chemotherapeutic drugs through administrating cell apoptosis and cell cycle arrest via regulating p53 activation and CHEK2-p53 apoptosis pathway.

Effective Control of Chronic γ-Herpesvirus Infection by Unconventional MHC Class Ia–Independent CD8 T Cells

PubMed Central

Tibbetts, Scott A; McClellan, Kelly B

2006-01-01

Control of virus infection is mediated in part by major histocompatibility complex (MHC) Class Ia presentation of viral peptides to conventional CD8 T cells. Although important, the absolute requirement for MHC Class Ia–dependent CD8 T cells for control of chronic virus infection has not been formally demonstrated. We show here that mice lacking MHC Class Ia molecules (Kb−/−xDb−/− mice) effectively control chronic γ-herpesvirus 68 (γHV68) infection via a robust expansion of β2-microglobulin (β2-m)-dependent, but CD1d-independent, unconventional CD8 T cells. These unconventional CD8 T cells expressed: (1) CD8αβ and CD3, (2) cell surface molecules associated with conventional effector/memory CD8 T cells, (3) TCRαβ with a significant Vβ4, Vβ3, and Vβ10 bias, and (4) the key effector cytokine interferon-γ (IFNγ). Unconventional CD8 T cells utilized a diverse TCR repertoire, and CDR3 analysis suggests that some of that repertoire may be utilized even in the presence of conventional CD8 T cells. This is the first demonstration to our knowledge that β2-m–dependent, but Class Ia–independent, unconventional CD8 T cells can efficiently control chronic virus infection, implicating a role for β2-n–dependent non-classical MHC molecules in control of chronic viral infection. We speculate that similar unconventional CD8 T cells may be able to control of other chronic viral infections, especially when viruses evade immunity by inhibiting generation of Class Ia–restricted T cells. PMID:16733540

Cell-cycle control in the face of damage--a matter of life or death.

PubMed

Clarke, Paul R; Allan, Lindsey A

2009-03-01

Cells respond to DNA damage or defects in the mitotic spindle by activating checkpoints that arrest the cell cycle. Alternatively, damaged cells can undergo cell death by the process of apoptosis. The correct balance between these pathways is important for the maintenance of genomic integrity while preventing unnecessary cell death. Although the molecular mechanisms of the cell cycle and apoptosis have been elucidated, the links between them have not been clear. Recent work, however, indicates that common components directly link the regulation of apoptosis with cell-cycle checkpoints operating during interphase, whereas in mitosis, the control of apoptosis is directly coupled to the cell-cycle machinery. These findings shed new light on how the balance between cell-cycle progression and cell death is controlled.

Protective effects of SGLT2 inhibitor luseogliflozin on pancreatic β-cells in obese type 2 diabetic db/db mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Okauchi, Seizo, E-mail: okauchi@med.kawasaki-m.ac.jp; Shimoda, Masashi; Obata, Atsushi

It is well known that Sodium-Glucose Co-transporter 2 (SGLT2) inhibitors, new hypoglycemic agents, improve glycemic control by increasing urine glucose excretion, but it remained unclear how they exert protective effects on pancreatic β-cells. In this study, we examined the effects of SGLT2 inhibitor luseogliflozin on β-cell function and mass using obese type 2 diabetic db/db mice. Ten-week-old male diabetic db/db mice were treated with luseogliflozin 0.0025% or 0.01% in chow (Luse 0.0025% or Luse 0.01%) or vehicle (control) for 4 weeks. Urinary glucose excretion was increased in Luse groups (0.0025% and 0.01%) compared to control mice 3 days after themore » intervention. Fasting blood glucose levels were significantly lower in mice treated with Luse compared to control mice. Fasting serum insulin concentrations were significantly higher in mice treated with Luse compared to control mice. Triglyceride levels tended to be lower in Luse groups compared to control mice. In immunohistochemical study using pancreas tissues, β-cell mass was larger in Luse groups compared to control group which was due to the increase of β-cell proliferation and decrease of β-cell apoptosis. Furthermore, in gene analysis using isolated islets, insulin 1, insulin 2, MafA, PDX-1 and GLUT2 gene expression levels were significantly higher in Luse groups compared to control group. In contrast, expression levels of fibrosis-related gene such as TGFβ, fibronectin, collagen I and collagen III were significantly lower in Luse groups. In conclusion, SGLT2 inhibitor luseogliflozin ameliorates glycemic control and thus exerts protective effects on pancreatic β-cell mass and function. - Highlights: • SGLT2 inhibitor luseogliflozin ameliorates glycemic control in db/db mice. • Luseogliflozin increases β-cell proliferation and decreases β-cell apoptosis. • Luseogliflozin preserves various β-cell-specific gene expression. • Luseogliflozin decreases various fibrosis-related factors in db/db mice.« less

Advanced Cell-Level Control for Extending Electric Vehicle Battery Pack Lifetime

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rehman, M. Muneeb Ur; Zhang, Fan; Evzelman, Michael

A cell-level control approach for electric vehicle battery packs is presented that enhances traditional battery balancing goals to not only provide cell balancing but also achieve significant pack lifetime extension. These goals are achieved by applying a new life-prognostic based control algorithm that biases individual cells differently based on their state of charge, capacity and internal resistance. The proposed life control approach reduces growth in capacity mismatch typically seen in large battery packs over life while optimizing usable energy of the pack. The result is a longer lifetime of the overall pack and a more homogeneous distribution of cell capacitiesmore » at the end of the first life for vehicle applications. Active cell balancing circuits and associated algorithms are used to accomplish the cell-level life extension objectives. This paper presents details of the cell-level control approach, selection and design of the active balancing system, and low-complexity state-of-charge, capacity, and series-resistance estimation algorithms. A laboratory prototype is used to demonstrate the proposed control approach. The prototype consists of twenty-one 25 Ah Panasonic lithium-Ion NMC battery cells from a commercial electric vehicle and an integrated BMS/DC-DC system that provides 750 W to the vehicle low voltage auxiliary loads.« less

Advanced Cell-Level Control for Extending Electric Vehicle Battery Pack Lifetime

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rehman, M. Muneeb Ur; Zhang, Fan; Evzelman, Michael

2017-02-16

A cell-level control approach for electric vehicle battery packs is presented that enhances traditional battery balancing goals to not only provide cell balancing but also achieve significant pack lifetime extension. These goals are achieved by applying a new life-prognostic based control algorithm that biases individual cells differently based on their state of charge, capacity and internal resistance. The proposed life control approach reduces growth in capacity mismatch typically seen in large battery packs over life while optimizing usable energy of the pack. The result is a longer lifetime of the overall pack and a more homogeneous distribution of cell capacitiesmore » at the end of the first life for vehicle applications. Active cell balancing circuits and associated algorithms are used to accomplish the cell-level life extension objectives. This paper presents details of the cell-level control approach, selection and design of the active balancing system, and low-complexity state-of-charge, capacity, and series-resistance estimation algorithms. A laboratory prototype is used to demonstrate the proposed control approach. The prototype consists of twenty-one 25 Ah Panasonic lithium-Ion NMC battery cells from a commercial electric vehicle and an integrated BMS/DC-DC system that provides 750 W to the vehicle low voltage auxiliary loads.« less

Demonstration of Passive Fuel Cell Thermal Management Technology

NASA Technical Reports Server (NTRS)

Burke, Kenneth A.; Jakupca, Ian; Colozza, Anthony; Wynne, Robert; Miller, Michael; Meyer, Al; Smith, William

2012-01-01

The NASA Glenn Research Center is developing advanced passive thermal management technology to reduce the mass and improve the reliability of space fuel cell systems for the NASA Exploration program. The passive thermal management system relies on heat conduction within highly thermally conductive cooling plates to move the heat from the central portion of the cell stack out to the edges of the fuel cell stack. Using the passive approach eliminates the need for a coolant pump and other cooling loop components within the fuel cell system which reduces mass and improves overall system reliability. Previous development demonstrated the performance of suitable highly thermally conductive cooling plates and integrated heat exchanger technology to collect the heat from the cooling plates (Ref. 1). The next step in the development of this passive thermal approach was the demonstration of the control of the heat removal process and the demonstration of the passive thermal control technology in actual fuel cell stacks. Tests were run with a simulated fuel cell stack passive thermal management system outfitted with passive cooling plates, an integrated heat exchanger and two types of cooling flow control valves. The tests were run to demonstrate the controllability of the passive thermal control approach. Finally, successful demonstrations of passive thermal control technology were conducted with fuel cell stacks from two fuel cell stack vendors.

Design of a compact microfludic device for controllable cell distribution.

PubMed

Li, Jing-Liang; Day, Daniel; Gu, Min

2010-11-21

A compact microfluidic device with 96 microchambers allocated within four circular units was designed and examined for cell distribution. In each unit, cells were distributed to the surrounding chambers radially from the center. The circular arrangement of the chambers makes the design simple and compact. A controllable and quantitative cell distribution is achievable in this device. This design is significant to the microfluidic applications where controllable distribution of cells in multipule microchambers is demanded.

Fuel sensor-less control of a liquid feed fuel cell under dynamic loading conditions for portable power sources (II)

NASA Astrophysics Data System (ADS)

Chang, C. L.; Chen, C. Y.; Sung, C. C.; Liou, D. H.; Chang, C. Y.; Cha, H. C.

This work presents a new fuel sensor-less control scheme for liquid feed fuel cells that is able to control the supply to a fuel cell system for operation under dynamic loading conditions. The control scheme uses cell-operating characteristics, such as potential, current, and power, to regulate the fuel concentration of a liquid feed fuel cell without the need for a fuel concentration sensor. A current integral technique has been developed to calculate the quantity of fuel required at each monitoring cycle, which can be combined with the concentration regulating process to control the fuel supply for stable operation. As verified by systematic experiments, this scheme can effectively control the fuel supply of a liquid feed fuel cell with reduced response time, even under conditions where the membrane electrolyte assembly (MEA) deteriorates gradually. This advance will aid the commercialization of liquid feed fuel cells and make them more adaptable for use in portable and automotive power units such as laptops, e-bikes, and handicap cars.

Distinct Inflammatory Profiles of Myelin-Reactive T cells from Patients with Multiple Sclerosis

PubMed Central

Cao, Yonghao; Goods, Brittany A.; Raddassi, Khadir; Nepom, Gerald T.; Kwok, William W.; Love, J. Christopher; Hafler, David A.

2015-01-01

Myelin-reactive T cells have been identified in patients with multiple sclerosis (MS) and healthy subjects with comparable frequencies, but the functional programs of self-reactive T cells that promote disease remain unknown. A total of 13,324 T cell libraries generated from blood of 23 patients and 22 healthy controls were interrogated for reactivity to myelin antigens. Libraries derived from CCR6+ myelin-reactive T cells from patients with MS exhibited significantly enhanced production of IFN-γ, IL-17, and GM-CSF compared to healthy controls. Single-cell clones isolated by MHC/peptide tetramers from CCR6+ T cell libraries also secreted more pro-inflammatory cytokines while clones isolated from controls secreted more IL-10. The transcriptomes of myelin-specific CCR6+ T cells from patients with MS were distinct from those derived from healthy controls, and of note, were enriched in Th17-induced experimental autoimmune encephalitis (EAE) gene signatures and gene signatures derived from Th17 cells isolated other human autoimmune diseases. These data, although not casual, imply that functional differences between antigen specific T cells from MS and healthy controls is fundamental to disease development and support the notion that IL-10 production from myelin-reactive T cells may act to limit disease progression, or even pathogenesis. PMID:25972006

Growth control of the eukaryote cell: a systems biology study in yeast.

PubMed

Castrillo, Juan I; Zeef, Leo A; Hoyle, David C; Zhang, Nianshu; Hayes, Andrew; Gardner, David Cj; Cornell, Michael J; Petty, June; Hakes, Luke; Wardleworth, Leanne; Rash, Bharat; Brown, Marie; Dunn, Warwick B; Broadhurst, David; O'Donoghue, Kerry; Hester, Svenja S; Dunkley, Tom Pj; Hart, Sarah R; Swainston, Neil; Li, Peter; Gaskell, Simon J; Paton, Norman W; Lilley, Kathryn S; Kell, Douglas B; Oliver, Stephen G

2007-01-01

Cell growth underlies many key cellular and developmental processes, yet a limited number of studies have been carried out on cell-growth regulation. Comprehensive studies at the transcriptional, proteomic and metabolic levels under defined controlled conditions are currently lacking. Metabolic control analysis is being exploited in a systems biology study of the eukaryotic cell. Using chemostat culture, we have measured the impact of changes in flux (growth rate) on the transcriptome, proteome, endometabolome and exometabolome of the yeast Saccharomyces cerevisiae. Each functional genomic level shows clear growth-rate-associated trends and discriminates between carbon-sufficient and carbon-limited conditions. Genes consistently and significantly upregulated with increasing growth rate are frequently essential and encode evolutionarily conserved proteins of known function that participate in many protein-protein interactions. In contrast, more unknown, and fewer essential, genes are downregulated with increasing growth rate; their protein products rarely interact with one another. A large proportion of yeast genes under positive growth-rate control share orthologs with other eukaryotes, including humans. Significantly, transcription of genes encoding components of the TOR complex (a major controller of eukaryotic cell growth) is not subject to growth-rate regulation. Moreover, integrative studies reveal the extent and importance of post-transcriptional control, patterns of control of metabolic fluxes at the level of enzyme synthesis, and the relevance of specific enzymatic reactions in the control of metabolic fluxes during cell growth. This work constitutes a first comprehensive systems biology study on growth-rate control in the eukaryotic cell. The results have direct implications for advanced studies on cell growth, in vivo regulation of metabolic fluxes for comprehensive metabolic engineering, and for the design of genome-scale systems biology models of the eukaryotic cell.

Growth control of the eukaryote cell: a systems biology study in yeast

PubMed Central

Castrillo, Juan I; Zeef, Leo A; Hoyle, David C; Zhang, Nianshu; Hayes, Andrew; Gardner, David CJ; Cornell, Michael J; Petty, June; Hakes, Luke; Wardleworth, Leanne; Rash, Bharat; Brown, Marie; Dunn, Warwick B; Broadhurst, David; O'Donoghue, Kerry; Hester, Svenja S; Dunkley, Tom PJ; Hart, Sarah R; Swainston, Neil; Li, Peter; Gaskell, Simon J; Paton, Norman W; Lilley, Kathryn S; Kell, Douglas B; Oliver, Stephen G

2007-01-01

Background Cell growth underlies many key cellular and developmental processes, yet a limited number of studies have been carried out on cell-growth regulation. Comprehensive studies at the transcriptional, proteomic and metabolic levels under defined controlled conditions are currently lacking. Results Metabolic control analysis is being exploited in a systems biology study of the eukaryotic cell. Using chemostat culture, we have measured the impact of changes in flux (growth rate) on the transcriptome, proteome, endometabolome and exometabolome of the yeast Saccharomyces cerevisiae. Each functional genomic level shows clear growth-rate-associated trends and discriminates between carbon-sufficient and carbon-limited conditions. Genes consistently and significantly upregulated with increasing growth rate are frequently essential and encode evolutionarily conserved proteins of known function that participate in many protein-protein interactions. In contrast, more unknown, and fewer essential, genes are downregulated with increasing growth rate; their protein products rarely interact with one another. A large proportion of yeast genes under positive growth-rate control share orthologs with other eukaryotes, including humans. Significantly, transcription of genes encoding components of the TOR complex (a major controller of eukaryotic cell growth) is not subject to growth-rate regulation. Moreover, integrative studies reveal the extent and importance of post-transcriptional control, patterns of control of metabolic fluxes at the level of enzyme synthesis, and the relevance of specific enzymatic reactions in the control of metabolic fluxes during cell growth. Conclusion This work constitutes a first comprehensive systems biology study on growth-rate control in the eukaryotic cell. The results have direct implications for advanced studies on cell growth, in vivo regulation of metabolic fluxes for comprehensive metabolic engineering, and for the design of genome-scale systems biology models of the eukaryotic cell. PMID:17439666

Gammadelta receptor bearing T cells in scleroderma: enhanced interaction with vascular endothelial cells in vitro.

PubMed

Kahaleh, M B; Fan, P S; Otsuka, T

1999-05-01

In view of the documented perivascular mononuclear cell infiltration in the involved organs in scleroderma (SSc) and the reported accumulation of gammadelta-T cells in SSc skin and lung, we evaluated gammadelta-T cell interaction with endothelial cells (EC) in vitro. gammadelta- and alphabeta-T cells were isolated from BPMN of SSc patients with early diffuse disease and of matched control subjects by an immunomagnetic method after stimulation with mycobacterium lysate and interleukin-2 for 2 weeks. Lymphocyte adhesion, proliferation, and cytotoxicity to EC were investigated. SSc gammadelta-T cells adhered to cultured EC and proliferated at higher rates than control cells. Furthermore, significant EC cytotoxicity by SSc gammadelta was seen. The cytotoxicity was blocked by addition of anti-gammadelta-TCR antibody and by anti-granzyme A antibody but not by anti-MHC class I and II antibodies. Expression of granzyme A mRNA was seen in five/five SSc gammadelta-T cells and in one/five control cells. alphabeta-T cells from both SSc and control subjects were significantly less interactive with EC than gammadelta-T cells. The data demonstrate EC recognition by SSc gammadelta-T cells and propose gammadelta-T cells as a possible effector cell type in the immune pathogenesis of SSc. Copyright 1999 Academic Press.

Drop-on-Demand Single Cell Isolation and Total RNA Analysis

PubMed Central

Moon, Sangjun; Kim, Yun-Gon; Dong, Lingsheng; Lombardi, Michael; Haeggstrom, Edward; Jensen, Roderick V.; Hsiao, Li-Li; Demirci, Utkan

2011-01-01

Technologies that rapidly isolate viable single cells from heterogeneous solutions have significantly contributed to the field of medical genomics. Challenges remain both to enable efficient extraction, isolation and patterning of single cells from heterogeneous solutions as well as to keep them alive during the process due to a limited degree of control over single cell manipulation. Here, we present a microdroplet based method to isolate and pattern single cells from heterogeneous cell suspensions (10% target cell mixture), preserve viability of the extracted cells (97.0±0.8%), and obtain genomic information from isolated cells compared to the non-patterned controls. The cell encapsulation process is both experimentally and theoretically analyzed. Using the isolated cells, we identified 11 stem cell markers among 1000 genes and compare to the controls. This automated platform enabling high-throughput cell manipulation for subsequent genomic analysis employs fewer handling steps compared to existing methods. PMID:21412416

Automated single cell microbioreactor for monitoring intracellular dynamics and cell growth in free solution†

PubMed Central

Johnson-Chavarria, Eric M.; Agrawal, Utsav; Tanyeri, Melikhan; Kuhlman, Thomas E.

2014-01-01

We report an automated microfluidic-based platform for single cell analysis that allows for cell culture in free solution with the ability to control the cell growth environment. Using this approach, cells are confined by the sole action of gentle fluid flow, thereby enabling non-perturbative analysis of cell growth away from solid boundaries. In addition, the single cell microbioreactor allows for precise and time-dependent control over cell culture media, with the combined ability to observe the dynamics of non-adherent cells over long time scales. As a proof-of-principle demonstration, we used the platform to observe dynamic cell growth, gene expression, and intracellular diffusion of repressor proteins while precisely tuning the cell growth environment. Overall, this microfluidic approach enables the direct observation of cellular dynamics with exquisite control over environmental conditions, which will be useful for quantifying the behaviour of single cells in well-defined media. PMID:24836754

Microprocessor controlled advanced battery management systems

NASA Technical Reports Server (NTRS)

Payne, W. T.

1978-01-01

The advanced battery management system described uses the capabilities of an on-board microprocessor to: (1) monitor the state of the battery on a cell by cell basis; (2) compute the state of charge of each cell; (3) protect each cell from reversal; (4) prevent overcharge on each individual cell; and (5) control dual rate reconditioning to zero volts per cell.

Calpain-Mediated positional information directs cell wall orientation to sustain plant stem cell activity, growth and development

USDA-ARS?s Scientific Manuscript database

Eukaryotic development and stem cell control depend on the integration of cell positional sensing with cell cycle control and cell wall positioning, yet few factors that directly link these events are known. The DEFECTIVE KERNEL1 (DEK1) gene encoding the unique plant calpain protein is fundamental f...

Increased IFN-gamma production by NK and CD3+/CD56+ cells in sexually HIV-1-exposed but uninfected individuals.

PubMed

Montoya, Carlos Julio; Velilla, Paula Andrea; Chougnet, Claire; Landay, Alan L; Rugeles, Maria Teresa

2006-08-01

The mechanisms involved in controlling the establishment of HIV-1 infection are not fully understood. In particular, the role of innate immunity in natural resistance exhibited by individuals who are continuously exposed to HIV-1 but remain seronegative (ESN) has not been thoroughly evaluated. We determined the frequency and function of peripheral blood innate immune cells (plasmacytoid and myeloid dendritic cells, monocytes, NK cells, CD3+/CD56+ cells and invariant NKT cells) in ESN, chronically HIV-1-infected and low-risk HIV-1 seronegative individuals. ESN demonstrated a similar frequency of innate immune cells in comparison to controls and a higher frequency of dendritic cells, NK and invariant NKT cells compared to HIV-1-infected subjects. Incubation of mononuclear cells with stimulatory CpG ODN induced CD86 and CD69 up-regulation to a similar degree on innate cells from the three study groups. CpG ODN-stimulated secretion of cytokines was also similar between ESN and controls, while secretion of IFN-alpha was significantly decreased in HIV-1+ individuals. Importantly, expression of IFN-gamma by PMA/Ionomycin-activated CD56(bright) NK cells and CD3+/CD56+ cells was significantly higher in ESN when compared with controls. The anti-viral effects of IFN-gamma are well established, and so our results suggest that IFN-gamma production by innate immune cells might be one of the multiple factors involved in controlling the establishment of sexually transmitted HIV-1 infection.

Assembly of multiple cell gradients directed by three-dimensional microfluidic channels.

PubMed

Li, Yiwei; Feng, Xiaojun; Wang, Yachao; Du, Wei; Chen, Peng; Liu, Chao; Liu, Bi-Feng

2015-08-07

Active control over the cell gradient is essential for understanding biological systems and the reconstitution of the functionality of many types of tissues, particularly for organ-on-a-chip. Here, we propose a three-dimensional (3D) microfluidic strategy for generating controllable cell gradients. In this approach, a homogeneous cell suspension is loaded into a 3D stair-shaped PDMS microchannel to generate a cell gradient within 10 min by sedimentation. We demonstrate that cell gradients of various profiles (exponential and piecewise linear) can be achieved by precisely controlling the height of each layer during the fabrication. With sequential seeding, we further demonstrate the generation of two overlapping cell gradients on the same glass substrate with pre-defined designs. The cell gradient-based QD cytotoxicity assay also demonstrated that cell behaviors and resistances were regulated by the changes in cell density. These results reveal that the proposed 3D microfluidic strategy provides a simple and versatile means for establishing controllable gradients in cell density, opening up a new avenue for reconstructing functional tissues.

A PML/Slit Axis Controls Physiological Cell Migration and Cancer Invasion in the CNS.

PubMed

Amodeo, Valeria; A, Deli; Betts, Joanne; Bartesaghi, Stefano; Zhang, Ying; Richard-Londt, Angela; Ellis, Matthew; Roshani, Rozita; Vouri, Mikaella; Galavotti, Sara; Oberndorfer, Sarah; Leite, Ana Paula; Mackay, Alan; Lampada, Aikaterini; Stratford, Eva Wessel; Li, Ningning; Dinsdale, David; Grimwade, David; Jones, Chris; Nicotera, Pierluigi; Michod, David; Brandner, Sebastian; Salomoni, Paolo

2017-07-11

Cell migration through the brain parenchyma underpins neurogenesis and glioblastoma (GBM) development. Since GBM cells and neuroblasts use the same migratory routes, mechanisms underlying migration during neurogenesis and brain cancer pathogenesis may be similar. Here, we identify a common pathway controlling cell migration in normal and neoplastic cells in the CNS. The nuclear scaffold protein promyelocytic leukemia (PML), a regulator of forebrain development, promotes neural progenitor/stem cell (NPC) and neuroblast migration in the adult mouse brain. The PML pro-migratory role is active also in transformed mouse NPCs and in human primary GBM cells. In both normal and neoplastic settings, PML controls cell migration via Polycomb repressive complex 2 (PRC2)-mediated repression of Slits, key regulators of axon guidance. Finally, a PML/SLIT1 axis regulates sensitivity to the PML-targeting drug arsenic trioxide in primary GBM cells. Taken together, these findings uncover a drug-targetable molecular axis controlling cell migration in both normal and neoplastic cells. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

System for controlling the operating temperature of a fuel cell

DOEpatents

Fabis, Thomas R.; Makiel, Joseph M.; Veyo, Stephen E.

2006-06-06

A method and system are provided for improved control of the operating temperature of a fuel cell (32) utilizing an improved temperature control system (30) that varies the flow rate of inlet air entering the fuel cell (32) in response to changes in the operating temperature of the fuel cell (32). Consistent with the invention an improved temperature control system (30) is provided that includes a controller (37) that receives an indication of the temperature of the inlet air from a temperature sensor (39) and varies the heat output by at least one heat source (34, 36) to maintain the temperature of the inlet air at a set-point T.sub.inset. The controller (37) also receives an indication of the operating temperature of the fuel cell (32) and varies the flow output by an adjustable air mover (33), within a predetermined range around a set-point F.sub.set, in order to maintain the operating temperature of the fuel cell (32) at a set-point T.sub.opset.

Introduction of N-cadherin-binding motif to alginate hydrogels for controlled stem cell differentiation.

PubMed

Lee, Jae Won; An, Hyoseok; Lee, Kuen Yong

2017-07-01

Control of stem cell fate and phenotype using biomimetic synthetic extracellular matrices (ECMs) is an important tissue engineering approach. Many studies have focused on improving cell-matrix interactions. However, proper control of cell-cell interactions using synthetic ECMs could be critical for tissue engineering, especially with undifferentiated stem cells. In this study, alginate hydrogels were modified with a peptide derived from the low-density lipoprotein receptor-related protein 5 (LRP5), which is known to bind to N-cadherin, as a cell-cell interaction motif. In vitro changes in the morphology and differentiation of mouse bone marrow stromal cells (D1 stem cells) cultured in LRP5-alginate hydrogels were investigated. LRP5-alginate gels successfully induced stem cell aggregation and enhanced chondrogenic differentiation of D1 stem cells, compared to RGD-alginate gels, at low cell density. This approach to tailoring synthetic biomimetic ECMs using cell-cell interaction motifs may be critical in tissue engineering approaches using stem cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Lithium-Ion Cell Charge-Control Unit Developed

NASA Technical Reports Server (NTRS)

Reid, Concha M.; Manzo, Michelle A.; Buton, Robert M.; Gemeiner, Russel

2005-01-01

A lithium-ion (Li-ion) cell charge-control unit was developed as part of a Li-ion cell verification program. This unit manages the complex charging scheme that is required when Li-ion cells are charged in series. It enables researchers to test cells together as a pack, while allowing each cell to charge individually. This allows the inherent cell-to-cell variations to be addressed on a series string of cells and reduces test costs substantially in comparison to individual cell testing.

Voltage controlled nano-injection system for single-cell surgery

PubMed Central

Seger, R. Adam; Actis, Paolo; Penfold, Catherine; Maalouf, Michelle; Vilozny, Boaz; Pourmand, Nader

2015-01-01

Manipulation and analysis of single cells is the next frontier in understanding processes that control the function and fate of cells. Herein we describe a single-cell injection platform based on nanopipettes. The system uses scanning microscopy techniques to detect cell surfaces, and voltage pulses to deliver molecules into individual cells. As a proof of concept, we injected adherent mammalian cells with fluorescent dyes. PMID:22899383

Voltage controlled nano-injection system for single-cell surgery.

PubMed

Adam Seger, R; Actis, Paolo; Penfold, Catherine; Maalouf, Michelle; Vilozny, Boaz; Pourmand, Nader

2012-09-28

Manipulation and analysis of single cells is the next frontier in understanding processes that control the function and fate of cells. Herein we describe a single-cell injection platform based on nanopipettes. The system uses scanning microscopy techniques to detect cell surfaces, and voltage pulses to deliver molecules into individual cells. As a proof of concept, we injected adherent mammalian cells with fluorescent dyes.

Fuel sensor-less control of a liquid feed fuel cell system under steady load for portable applications

NASA Astrophysics Data System (ADS)

Chang, C. L.; Chen, C. Y.; Sung, C. C.; Liou, D. H.

This study presents a novel fuel sensor-less control scheme for a liquid feed fuel cell system that does not rely on a fuel concentration sensor. The proposed approach simplifies the design and reduces the cost and complexity of a liquid feed fuel cell system, and is especially suited to portable power sources, of which the volume and weight are important. During the reaction of a fuel cell, the cell's operating characteristics, such as potential, current and power are measured to control the supply of fuel and regulate its concentration to optimize performance. Experiments were conducted to verify that the fuel sensor-less control algorithm is effective in the liquid feed fuel cell system.

Nonlinear Recurrent Neural Network Predictive Control for Energy Distribution of a Fuel Cell Powered Robot

PubMed Central

Chen, Qihong; Long, Rong; Quan, Shuhai

2014-01-01

This paper presents a neural network predictive control strategy to optimize power distribution for a fuel cell/ultracapacitor hybrid power system of a robot. We model the nonlinear power system by employing time variant auto-regressive moving average with exogenous (ARMAX), and using recurrent neural network to represent the complicated coefficients of the ARMAX model. Because the dynamic of the system is viewed as operating- state- dependent time varying local linear behavior in this frame, a linear constrained model predictive control algorithm is developed to optimize the power splitting between the fuel cell and ultracapacitor. The proposed algorithm significantly simplifies implementation of the controller and can handle multiple constraints, such as limiting substantial fluctuation of fuel cell current. Experiment and simulation results demonstrate that the control strategy can optimally split power between the fuel cell and ultracapacitor, limit the change rate of the fuel cell current, and so as to extend the lifetime of the fuel cell. PMID:24707206

Controlling Cell Function with Geometry

NASA Astrophysics Data System (ADS)

Mrksich, Milan

2012-02-01

This presentation will describe the use of patterned substrates to control cell shape with examples that illustrate the ways in which cell shape can regulate cell function. Most cells are adherent and must attach to and spread on a surface in order to survive, proliferate and function. In tissue, this surface is the extracellular matrix (ECM), an insoluble scaffold formed by the assembly of several large proteins---including fibronectin, the laminins and collagens and others---but in the laboratory, the surface is prepared by adsorbing protein to glass slides. To pattern cells, gold-coated slides are patterned with microcontact printing to create geometric features that promote cell attachment and that are surrounded by inert regions. Cells attach to these substrates and spread to adopt the shape defined by the underlying pattern and remain stable in culture for several days. Examples will be described that used a series of shapes to reveal the relationship between the shape of the cell and the structure of its cytoskeleton. These geometric cues were used to control cell polarity and the tension, or contractility, present in the cytoskeleton. These rules were further used to control the shapes of mesenchymal stem cells and in turn to control the differentiation of these cells into specialized cell types. For example, stem cells that were patterned into a ``star'' shape preferentially differentiated into bone cells whereas those that were patterned into a ``flower'' shape preferred a fat cell fate. These influences of shape on differentiation depend on the mechanical properties of the cytoskeleton. These examples, and others, reveal that shape is an important cue that informs cell function and that can be combined with the more common soluble cues to direct and study cell function.

Photovoltaic Powering And Control System For Electrochromic Windows

DOEpatents

Schulz, Stephen C.; Michalski, Lech A.; Volltrauer, Hermann N.; Van Dine, John E.

2000-04-25

A sealed insulated glass unit is provided with an electrochromic device for modulating light passing through the unit. The electrochromic device is controlled from outside the unit by a remote control electrically unconnected to the device. Circuitry within the unit may be magnetically controlled from outside. The electrochromic device is powered by a photovoltaic cells. The photovoltaic cells may be positioned so that at least a part of the light incident on the cell passes through the electrochromic device, providing a form of feedback control. A variable resistance placed in parallel with the electrochromic element is used to control the response of the electrochromic element to changes in output of the photovoltaic cell.

12. ENGINE TEST CELL BUILDING INTERIOR. DETAIL OF CONTROL CONSOLE ...

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

12. ENGINE TEST CELL BUILDING INTERIOR. DETAIL OF CONTROL CONSOLE FOR ENGINE TEST CELL 4. LOOKING NORTH. - Fairchild Air Force Base, Engine Test Cell Building, Near intersection of Arnold Street & George Avenue, Spokane, Spokane County, WA

Exploring the Role of Ubiquitination in Progesterone Receptor Transcriptional Activation and Turnover in Breast Cancer Cells

DTIC Science & Technology

2006-06-01

factors. T47DY cells were cotransfected with a PR construct, a PRE- luciferase plasmid and a renilla plasmid, for transfection control. The cells...PR-B or S294A PR-B, PRE-luciferase reporter constructs and a Renilla control plasmid. Cells were treated for 24hrs with or without R5020 (10nM...plasmid and a plasmid constitutively expressing renilla luciferase for transfection control. Cell were starved for one day and treated with or without

Matrix Elasticity of Void-Forming Hydrogels Controls Transplanted Stem Cell-Mediated Bone Formation

PubMed Central

Huebsch, Nathaniel; Lippens, Evi; Lee, Kangwon; Mehta, Manav; Koshy, Sandeep T; Darnell, Max C; Desai, Rajiv; Madl, Christopher M.; Xu, Maria; Zhao, Xuanhe; Chaudhuri, Ovijit; Verbeke, Catia; Kim, Woo Seob; Alim, Karen; Mammoto, Akiko; Ingber, Donald E.; Duda, Georg N; Mooney, David J.

2015-01-01

The effectiveness of stem-cell therapies has been hampered by cell death and limited control over fate1. These problems can be partially circumvented by using macroporous biomaterials that improve the survival of transplanted stem cells and provide molecular cues to direct cell phenotype2–4. Stem cell behavior can also be controlled in vitro by manipulating the elasticity of both porous and non-porous materials5–7, yet translation to therapeutic processes in vivo remains elusive. Here, by developing injectable, void-forming hydrogels that decouple pore formation from elasticity, we show that mesenchymal stem cell (MSC) osteogenesis in vitro, and cell deployment in vitro and in vivo, can be controlled by modifying, respectively, the hydrogel's elastic modulus or its chemistry. When the hydrogels were used to transplant MSCs, the hydrogel's elasticity regulated bone regeneration, with optimal bone formation at 60 kPa. Our findings show that biophysical cues can be harnessed to direct therapeutic stem-cell behaviors in situ. PMID:26366848

Matrix elasticity of void-forming hydrogels controls transplanted-stem-cell-mediated bone formation

NASA Astrophysics Data System (ADS)

Huebsch, Nathaniel; Lippens, Evi; Lee, Kangwon; Mehta, Manav; Koshy, Sandeep T.; Darnell, Max C.; Desai, Rajiv M.; Madl, Christopher M.; Xu, Maria; Zhao, Xuanhe; Chaudhuri, Ovijit; Verbeke, Catia; Kim, Woo Seob; Alim, Karen; Mammoto, Akiko; Ingber, Donald E.; Duda, Georg N.; Mooney, David J.

2015-12-01

The effectiveness of stem cell therapies has been hampered by cell death and limited control over fate. These problems can be partially circumvented by using macroporous biomaterials that improve the survival of transplanted stem cells and provide molecular cues to direct cell phenotype. Stem cell behaviour can also be controlled in vitro by manipulating the elasticity of both porous and non-porous materials, yet translation to therapeutic processes in vivo remains elusive. Here, by developing injectable, void-forming hydrogels that decouple pore formation from elasticity, we show that mesenchymal stem cell (MSC) osteogenesis in vitro, and cell deployment in vitro and in vivo, can be controlled by modifying, respectively, the hydrogel’s elastic modulus or its chemistry. When the hydrogels were used to transplant MSCs, the hydrogel’s elasticity regulated bone regeneration, with optimal bone formation at 60 kPa. Our findings show that biophysical cues can be harnessed to direct therapeutic stem cell behaviours in situ.

Dendritic Cell Immune Responses in HIV-1 Controllers.

PubMed

Martin-Gayo, Enrique; Yu, Xu G

2017-02-01

Robust HIV-1-specific CD8 T cell responses are currently regarded as the main correlate of immune defense in rare individuals who achieve natural, drug-free control of HIV-1; however, the mechanisms that support evolution of such powerful immune responses are not well understood. Dendritic cells (DCs) are specialized innate immune cells critical for immune recognition, immune regulation, and immune induction, but their possible contribution to HIV-1 immune defense in controllers remains ill-defined. Recent studies suggest that myeloid DCs from controllers have improved abilities to recognize HIV-1 through cytoplasmic immune sensors, resulting in more potent, cell-intrinsic type I interferon secretion in response to viral infection. This innate immune response may facilitate DC-mediated induction of highly potent antiviral HIV-1-specific T cells. Moreover, protective HLA class I isotypes restricting HIV-1-specific CD8 T cells may influence DC function through specific interactions with innate myelomonocytic MHC class I receptors from the leukocyte immunoglobulin-like receptor family. Bi-directional interactions between dendritic cells and HIV-1-specific T cells may contribute to natural HIV-1 immune control, highlighting the importance of a fine-tuned interplay between innate and adaptive immune activities for effective antiviral immune defense.

Cell mechanics: a dialogue.

PubMed

Tao, Jiaxiang; Li, Yizeng; Vig, Dhruv K; Sun, Sean X

2017-03-01

Under the microscope, eukaryotic animal cells can adopt a variety of different shapes and sizes. These cells also move and deform, and the physical mechanisms driving these movements and shape changes are important in fundamental cell biology, tissue mechanics, as well as disease biology. This article reviews some of the basic mechanical concepts in cells, emphasizing continuum mechanics description of cytoskeletal networks and hydrodynamic flows across the cell membrane. We discuss how cells can generate movement and shape changes by controlling mass fluxes at the cell boundary. These mass fluxes can come from polymerization/depolymerization of actin cytoskeleton, as well as osmotic and hydraulic pressure-driven flow of water across the cell membrane. By combining hydraulic pressure control with force balance conditions at the cell surface, we discuss a quantitative mechanism of cell shape and volume control. The broad consequences of this model on cell mechanosensation and tissue mechanics are outlined.

Cell mechanics: a dialogue

PubMed Central

Tao, Jiaxiang; Li, Yizeng; Vig, Dhruv K; Sun, Sean X

2017-01-01

Under the microscope, eukaryotic animal cells can adopt a variety of different shapes and sizes. These cells also move and deform, and the physical mechanisms driving these movements and shape changes are important in fundamental cell biology, tissue mechanics, as well as disease biology. This article reviews some of the basic mechanical concepts in cells, emphasizing continuum mechanics description of cytoskeletal networks and hydrodynamic flows across the cell membrane. We discuss how cells can generate movement and shape changes by controlling mass fluxes at the cell boundary. These mass fluxes can come from polymerization/depolymerization of actin cytoskeleton, as well as osmotic and hydraulic pressure-driven flow of water across the cell membrane. By combining hydraulic pressure control with force balance conditions at the cell surface, we discuss a quantitative mechanism of cell shape and volume control. The broad consequences of this model on cell mechanosensation and tissue mechanics are outlined. PMID:28129208

Cell mechanics: a dialogue

NASA Astrophysics Data System (ADS)

Tao, Jiaxiang; Li, Yizeng; Vig, Dhruv K.; Sun, Sean X.

2017-03-01

Under the microscope, eukaryotic animal cells can adopt a variety of different shapes and sizes. These cells also move and deform, and the physical mechanisms driving these movements and shape changes are important in fundamental cell biology, tissue mechanics, as well as disease biology. This article reviews some of the basic mechanical concepts in cells, emphasizing continuum mechanics description of cytoskeletal networks and hydrodynamic flows across the cell membrane. We discuss how cells can generate movement and shape changes by controlling mass fluxes at the cell boundary. These mass fluxes can come from polymerization/depolymerization of actin cytoskeleton, as well as osmotic and hydraulic pressure-driven flow of water across the cell membrane. By combining hydraulic pressure control with force balance conditions at the cell surface, we discuss a quantitative mechanism of cell shape and volume control. The broad consequences of this model on cell mechanosensation and tissue mechanics are outlined.

Asymmetric cell division during T cell development controls downstream fate

PubMed Central

Pham, Kim; Shimoni, Raz; Charnley, Mirren; Ludford-Menting, Mandy J.; Hawkins, Edwin D.; Ramsbottom, Kelly; Oliaro, Jane; Izon, David; Ting, Stephen B.; Reynolds, Joseph; Lythe, Grant; Molina-Paris, Carmen; Melichar, Heather; Robey, Ellen; Humbert, Patrick O.; Gu, Min

2015-01-01

During mammalian T cell development, the requirement for expansion of many individual T cell clones, rather than merely expansion of the entire T cell population, suggests a possible role for asymmetric cell division (ACD). We show that ACD of developing T cells controls cell fate through differential inheritance of cell fate determinants Numb and α-Adaptin. ACD occurs specifically during the β-selection stage of T cell development, and subsequent divisions are predominantly symmetric. ACD is controlled by interaction with stromal cells and chemokine receptor signaling and uses a conserved network of polarity regulators. The disruption of polarity by deletion of the polarity regulator, Scribble, or the altered inheritance of fate determinants impacts subsequent fate decisions to influence the numbers of DN4 cells arising after the β-selection checkpoint. These findings indicate that ACD enables the thymic microenvironment to orchestrate fate decisions related to differentiation and self-renewal. PMID:26370500

Fuel cell with internal flow control

DOEpatents

Haltiner, Jr., Karl J.; Venkiteswaran, Arun [Karnataka, IN

2012-06-12

A fuel cell stack is provided with a plurality of fuel cell cassettes where each fuel cell cassette has a fuel cell with an anode and cathode. The fuel cell stack includes an anode supply chimney for supplying fuel to the anode of each fuel cell cassette, an anode return chimney for removing anode exhaust from the anode of each fuel cell cassette, a cathode supply chimney for supplying oxidant to the cathode of each fuel cell cassette, and a cathode return chimney for removing cathode exhaust from the cathode of each fuel cell cassette. A first fuel cell cassette includes a flow control member disposed between the anode supply chimney and the anode return chimney or between the cathode supply chimney and the cathode return chimney such that the flow control member provides a flow restriction different from at least one other fuel cell cassettes.

Cytomorphometric and Morphological Analysis in Women with Trichomonas vaginalis Infection: Micronucleus Frequency in Exfoliated Cervical Epithelial Cells.

PubMed

Safi Oz, Zehra; Doğan Gun, Banu; Gun, Mustafa Ozkan; Ozdamar, Sukru Oguz

2015-01-01

The aim of this study was to explore the cytomorphometric and morphological effects of Trichomonas vaginalis in exfoliated epithelial cells. Ninety-six Pap-stained cervical smears were divided into a study group and two control groups as follows: T. vaginalis cases, a first control group with inflammation, and a second control group without inflammation. Micronucleated, binucleated, karyorrhectic, karyolytic, and karyopyknotic cells and cells with perinuclear halos per 1,000 epithelial cells were counted. Nuclear and cellular areas were evaluated in 70 clearly defined cells in each smear using image analysis. The frequencies of morphological parameters in the T. vaginalis cases were higher than the values of the two control groups, and the difference among groups was found to be significant (p < 0.05). The nuclear and cytoplasmic areas of epithelial cells were diminished in patients with trichomoniasis. The mean nucleus/cytoplasm ratio in T. vaginalis patients was higher than the value in the control groups, and the difference between the study group and control group 1 was significant. However, there was no statistically significant increase between the study group and control group 2. T. vaginalis exhibited significant changes in the cellular size and nuclear structure of the cells. The rising frequency of micronuclei, nuclear abnormalities, and changing nucleus/cytoplasm ratio may reflect genotoxic damage in trichomoniasis. © 2015 S. Karger AG, Basel.

Microfluidic cell trap array for controlled positioning of single cells on adhesive micropatterns.

PubMed

Lin, Laiyi; Chu, Yeh-Shiu; Thiery, Jean Paul; Lim, Chwee Teck; Rodriguez, Isabel

2013-02-21

Adhesive micropattern arrays permit the continuous monitoring and systematic study of the behavior of spatially confined cells of well-defined shape and size in ordered configurations. This technique has contributed to defining mechanisms that control cell polarity and cell functions, including proliferation, apoptosis, differentiation and migration in two-dimensional cell culture systems. These micropattern studies often involve isolating a single cell on one adhesive protein micropattern using random seeding methods. Random seeding has been successful for isolated and, to a lesser degree, paired patterns, where two patterns are placed in close proximity. Using this method, we found that the probability of obtaining one cell per pattern decreases significantly as the number of micropatterns in a cluster increases, from 16% for paired micropatterns to 0.3% for clusters of 6 micropatterns. This work presents a simple yet effective platform based on a microfludic sieve-like trap array to exert precise control over the positioning of single cells on micropatterns. We observed a 4-fold improvement over random seeding in the efficiency of placing a pair of single cells on paired micropattern and a 40-fold improvement for 6-pattern clusters. The controlled nature of this platform can also allow the juxtaposition of two different cell populations through a simple modification in the trap arrangement. With excellent control of the identity, number and position of neighbouring cells, this cell-positioning platform provides a unique opportunity for the extension of two-dimensional micropattern studies beyond paired micropatterns to organizations containing many cells or different cell types.

Persistence of decidual NK cells and KIR genotypes in healthy pregnant and preeclamptic women: a case-control study in the third trimester of gestation

PubMed Central

2011-01-01

Background Natural Killer (NK) cells are the most abundant lymphocytes in the decidua during early gestation. The interactions of NK cells with the extravillous cytotrophoblast have been associated with a normal spiral artery remodeling process, an essential event for a successful pregnancy. Recent data indicate that alterations in the amount of decidual NK (dNK) cells contribute to the development of preeclampsia (PE). Moreover, genetic studies suggest that Killer Immunoglobulin-like Receptors (KIR) expressed in dNK cells influence the susceptibility to PE. Although dNK cells have been well characterized during early pregnancy, they have been scarcely studied in the third trimester of gestation. The aim of this work was to characterize dNK cells at the last trimester of gestation and to analyze the KIR genotype of healthy and PE women. Methods Decidual samples were obtained during Caesarean section from control (n = 10) and PE (n = 9) women. Flow cytometric analysis of CD3, CD56, CD16 and CD9 was used to characterize and quantify dNK cells in both groups. Cell surface markers from decidual leukocytes were compared with PBMC from healthy donors. KIR genotyping was performed in genomic DNA (control, n = 86; PE, n = 90) using PCR-SSP. Results The results indicate that dNK cells persist throughout pregnancy. They represented 20% of total leukocytes in control and PE groups, and they expressed the same cell surface markers (CD3-, CD56+, CD16- and CD9+) as dNK in the first trimester of gestation. There were no significant differences in the percentage of dNK cells between control and PE groups. The analysis of KIR gene frequencies and genotypes was not statistically different between control and PE groups. The ratio of activating to inhibitory genes indicated that the overall inhibitory balance (0.2-0.5) was more frequent in the PE group (control, 31.3% vs PE, 45.5%), and the activating balance (0.6-1.1) was more frequent in the control group (control, 68.6% vs PE, 54.4%). However this difference was not significant. Conclusion We demonstrated the persistence of dNK cells in PE and control women at the third trimester of pregnancy; these dNK cells had a similar phenotype to those found during early pregnancy. The predominance of a KIR inhibitory balance in the PE group could be associated to the physiopathology of PE. PMID:21247496

Topological control of life and death in non-proliferative epithelia.

PubMed

Martinand-Mari, Camille; Maury, Benoit; Rousset, François; Sahuquet, Alain; Mennessier, Gérard; Rochal, Sergei; Lorman, Vladimir; Mangeat, Paul; Baghdiguian, Stephen

2009-01-01

Programmed cell death is one of the most fascinating demonstrations of the plasticity of biological systems. It is classically described to act upstream of and govern major developmental patterning processes (e.g. inter-digitations in vertebrates, ommatidia in Drosophila). We show here the first evidence that massive apoptosis can also be controlled and coordinated by a pre-established pattern of a specific 'master cell' population. This new concept is supported by the development and validation of an original model of cell patterning. Ciona intestinalis eggs are surrounded by a three-layered follicular organization composed of 60 elongated floating extensions made of as many outer and inner cells, and indirectly spread through an extracellular matrix over 1200 test cells. Experimental and selective ablation of outer and inner cells results in the abrogation of apoptosis in respective remaining neighbouring test cells. In addition incubation of outer/inner follicular cell-depleted eggs with a soluble extract of apoptotic outer/inner cells partially restores apoptosis to apoptotic-defective test cells. The 60 inner follicular cells were thus identified as 'apoptotic master' cells which collectively are induction sites for programmed cell death of the underlying test cells. The position of apoptotic master cells is controlled by topological constraints exhibiting a tetrahedral symmetry, and each cell spreads over and can control the destiny of 20 smaller test cells, which leads to optimized apoptosis signalling.

Charge-Control Unit for Testing Lithium-Ion Cells

NASA Technical Reports Server (NTRS)

Reid, Concha M.; Mazo, Michelle A.; Button, Robert M.

2008-01-01

A charge-control unit was developed as part of a program to validate Li-ion cells packaged together in batteries for aerospace use. The lithium-ion cell charge-control unit will be useful to anyone who performs testing of battery cells for aerospace and non-aerospace uses and to anyone who manufacturers battery test equipment. This technology reduces the quantity of costly power supplies and independent channels that are needed for test programs in which multiple cells are tested. Battery test equipment manufacturers can integrate the technology into their battery test equipment as a method to manage charging of multiple cells in series. The unit manages a complex scheme that is required for charging Li-ion cells electrically connected in series. The unit makes it possible to evaluate cells together as a pack using a single primary test channel, while also making it possible to charge each cell individually. Hence, inherent cell-to-cell variations in a series string of cells can be addressed, and yet the cost of testing is reduced substantially below the cost of testing each cell as a separate entity. The unit consists of electronic circuits and thermal-management devices housed in a common package. It also includes isolated annunciators to signal when the cells are being actively bypassed. These annunciators can be used by external charge managers or can be connected in series to signal that all cells have reached maximum charge. The charge-control circuitry for each cell amounts to regulator circuitry and is powered by that cell, eliminating the need for an external power source or controller. A 110-VAC source of electricity is required to power the thermal-management portion of the unit. A small direct-current source can be used to supply power for an annunciator signal, if desired.

The effect of desferrioxamine on transferrin receptors, the cell cycle and growth rates of human leukaemic cells.

PubMed Central

Bomford, A; Isaac, J; Roberts, S; Edwards, A; Young, S; Williams, R

1986-01-01

The effect of the iron chelator, desferrioxamine, on transferrin binding, growth rates and the cell cycle was investigated in the human leukaemic cell line, K562. At all concentrations of the chelator (2-50 microM) binding of 125I-transferrin was increased by 24 h and reached a maximum at 72-96 h. Maximum binding (6-8-fold increased) occurred in cells treated with 20 microM-desferrioxamine, in contrast with control cells which, at 96 h, showed a 50% decrease over initial binding. Scatchard analysis at 4 degrees C showed that this increased binding was due to an increase in the number of receptors, as the Kd was similar in induced (1.8 nM) and control (1.5 nM) cells. After 96 h cells, cultured with 20 and 50 microM-desferrioxamine accumulated 59Fe from bovine transferrin at over twice the rate found with control cells, reflecting the increase in transferrin receptors. Although iron uptake was unimpaired by the chelator there was a dose-dependent inhibition of cell growth, with control cells completing three divisions in 96 h and those in 10 microM-desferrioxamine only two divisions. At the highest concentration (50 microM), cell division was abrogated although cell viability was maintained (85%). In contrast, DNA synthesis was not markedly affected, except at 50 microM-desferrioxamine when incorporation of [3H]thymidine was 52% of that in control cells. Flow cytometry revealed that there was a progressive accumulation of the cells in the active phases of their cycle (S, G2 + M). Desferrioxamine may increase transferrin receptors in two ways: by chelating a regulatory pool of iron within the cell, and by arresting cells in S phase when receptors are maximally expressed. PMID:3790074

[Osteogenic potential of bone marrow mesenchymal stem cells from ovariectomied osteoporotic rat].

PubMed

Li, Dong-ju; Ge, Dong-xia; Wu, Wen-chao; Wu, Jiang; Li, Liang

2005-05-01

To investigate the difference of osteogenic potential of bone marrow mesenchymal stem cells (MSCs) between healthy rats and osteoporotic rats. We established the animal model of osteoporosis by performing ovariectom on the 3-month-old female Sprague-Dawley rats. Bone marrow mesenchymal stem cells(MSCs) were isolated from the rats of control group and of ovariectomized (ovx) group by means of the density-gradient centrifugation method, and the 3rd-4th passage MSCs were used in all the experiments. The experiments comprised 4 groups: (1) Marrow mesenchymal stem cells control group (MSCs control group); (2) Marrow mesenchymal stem cells ovx group (MSCs ovx group); (3) Osteogenesis induction control group (OSI control group); (4) Osteogenesis induction ovx group (OSI ovx group). Cell cycle and proliferation index (PI) of MSCs were detected by flow cytometry. The expression of alkaline phosphatase (ALP) was detected by dynamics method with substrate of phosphoric acid para-Nitro benzene. The levels of osteocalcin were detected with the isotope labelling method. (1) PI of MSCs was lower in MSCs ovx group than in MSCs control group. (2) The expression of alkaline phosphatase (ALP) was much higher in OSI control group than in the MSCs control group; the expression of alkaline phosphatase (ALP) was much higher in the OSI control group than in OSI ovx group after 7-day and 14-day osteogenic induction. (3) The level of osteocalcin was much higher in the OSI control group than in the MSCs control group after 14-day, 21-day, 28-day osteogenic induction. The level of osteocalcin was much higher in the OSI control group than in the OSI ovx group. Both the proliferative potential and the osteogenic potential of bone marrow mesenchymal stem cells (MSCs) from the ovariectomized osteoporotic rat are decreased.

S-Fms signalobody enhances myeloid cell growth and migration.

PubMed

Kawahara, Masahiro; Hitomi, Azusa; Nagamune, Teruyuki

2014-07-01

Since receptor tyrosine kinases (RTKs) control various cell fates in many types of cells, mimicry of RTK functions is promising for artificial control of cell fates. We have previously developed single-chain Fv (scFv)/receptor chimeras named signalobodies that can mimic receptor signaling in response to a specific antigen. While the RTK-based signalobodies enabled us to control cell growth and migration, further extension of applicability in another cell type would underlie the impact of the RTK-based signalobodies. In this study, we applied the scFv-c-Fms (S-Fms) signalobody in a murine myeloid progenitor cell line, FDC-P1. S-Fms transduced a fluorescein-conjugated BSA (BSA-FL)-dependent growth signal and activated downstream signaling molecules including MEK, ERK, Akt, and STAT3, which are major constituents of Ras/MAPK, PI3K/Akt, and JAK/STAT signaling pathways. In addition, S-Fms transduced a migration signal as demonstrated by the transwell-based migration assay. Direct real-time observation of the cells further confirmed that FDC/S-Fms cells underwent directional cell migration toward a positive gradient of BSA-FL. These results demonstrated the utility of the S-Fms signalobody for controlling growth and migration of myeloid cells. Further extension of our approach includes economical large-scale production of practically relevant blood cells as well as artificial control of cell migration for tissue regeneration and immune response. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Microvalve controlled multi-functional microfluidic chip for divisional cell co-culture.

PubMed

Li, Rui; Zhang, Xingjian; Lv, Xuefei; Geng, Lina; Li, Yongrui; Qin, Kuiwei; Deng, Yulin

2017-12-15

Pneumatic micro-valve controlled microfluidic chip provides precise fluidic control for cell manipulation. In this paper, a multi-functional microfluidic chip was designed for three separate experiments: 1. Different cell lines were dispensed and cultured; 2. Three transfected SH-SY5Y cells were introduced and treated with methyl-phenyl-pyridinium (MPP + ) as drug delivery mode; 3. Specific protection and interaction were observed among cell co-culture after nerve damage. The outcomes revealed the potential and practicability of our entire multi-functional pneumatic chip system on different cell biology applications. Copyright © 2017. Published by Elsevier Inc.

Cell Microenvironment Engineering and Monitoring for Tissue Engineering and Regenerative Medicine: The Recent Advances

PubMed Central

Barthes, Julien; Özçelik, Hayriye; Hindié, Mathilde; Ndreu-Halili, Albana; Hasan, Anwarul

2014-01-01

In tissue engineering and regenerative medicine, the conditions in the immediate vicinity of the cells have a direct effect on cells' behaviour and subsequently on clinical outcomes. Physical, chemical, and biological control of cell microenvironment are of crucial importance for the ability to direct and control cell behaviour in 3-dimensional tissue engineering scaffolds spatially and temporally. In this review, we will focus on the different aspects of cell microenvironment such as surface micro-, nanotopography, extracellular matrix composition and distribution, controlled release of soluble factors, and mechanical stress/strain conditions and how these aspects and their interactions can be used to achieve a higher degree of control over cellular activities. The effect of these parameters on the cellular behaviour within tissue engineering context is discussed and how these parameters are used to develop engineered tissues is elaborated. Also, recent techniques developed for the monitoring of the cell microenvironment in vitro and in vivo are reviewed, together with recent tissue engineering applications where the control of cell microenvironment has been exploited. Cell microenvironment engineering and monitoring are crucial parts of tissue engineering efforts and systems which utilize different components of the cell microenvironment simultaneously can provide more functional engineered tissues in the near future. PMID:25143954

Cell microenvironment engineering and monitoring for tissue engineering and regenerative medicine: the recent advances.

PubMed

Barthes, Julien; Özçelik, Hayriye; Hindié, Mathilde; Ndreu-Halili, Albana; Hasan, Anwarul; Vrana, Nihal Engin

2014-01-01

In tissue engineering and regenerative medicine, the conditions in the immediate vicinity of the cells have a direct effect on cells' behaviour and subsequently on clinical outcomes. Physical, chemical, and biological control of cell microenvironment are of crucial importance for the ability to direct and control cell behaviour in 3-dimensional tissue engineering scaffolds spatially and temporally. In this review, we will focus on the different aspects of cell microenvironment such as surface micro-, nanotopography, extracellular matrix composition and distribution, controlled release of soluble factors, and mechanical stress/strain conditions and how these aspects and their interactions can be used to achieve a higher degree of control over cellular activities. The effect of these parameters on the cellular behaviour within tissue engineering context is discussed and how these parameters are used to develop engineered tissues is elaborated. Also, recent techniques developed for the monitoring of the cell microenvironment in vitro and in vivo are reviewed, together with recent tissue engineering applications where the control of cell microenvironment has been exploited. Cell microenvironment engineering and monitoring are crucial parts of tissue engineering efforts and systems which utilize different components of the cell microenvironment simultaneously can provide more functional engineered tissues in the near future.

Use of nanoscale mechanical stimulation for control and manipulation of cell behaviour.

PubMed

Childs, Peter G; Boyle, Christina A; Pemberton, Gabriel D; Nikukar, Habib; Curtis, Adam S G; Henriquez, Fiona L; Dalby, Matthew J; Reid, Stuart

2016-04-01

The ability to control cell behaviour, cell fate and simulate reliable tissue models in vitro remains a significant challenge yet is crucial for various applications of high throughput screening e.g. drug discovery. Mechanotransduction (the ability of cells to convert mechanical forces in their environment to biochemical signalling) represents an alternative mechanism to attain this control with such studies developing techniques to reproducibly control the mechanical environment in techniques which have potential to be scaled. In this review, the use of techniques such as finite element modelling and precision interferometric measurement are examined to provide context for a novel technique based on nanoscale vibration, also known as "nanokicking". Studies have shown this stimulus to alter cellular responses in both endothelial and mesenchymal stem cells (MSCs), particularly in increased proliferation rate and induced osteogenesis respectively. Endothelial cell lines were exposed to nanoscale vibration amplitudes across a frequency range of 1-100 Hz, and MSCs primarily at 1 kHz. This technique provides significant potential benefits over existing technologies, as cellular responses can be initiated without the use of expensive engineering techniques and/or chemical induction factors. Due to the reproducible and scalable nature of the apparatus it is conceivable that nanokicking could be used for controlling cell behaviour within a wide array of high throughput procedures in the research environment, within drug discovery, and for clinical/therapeutic applications. The results discussed within this article summarise the potential benefits of using nanoscale vibration protocols for controlling cell behaviour. There is a significant need for reliable tissue models within the clinical and pharma industries, and the control of cell behaviour and stem cell differentiation would be highly beneficial. The full potential of this method of controlling cell behaviour has not yet been realised. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Analysis of the immune system of multiple myeloma patients achieving long-term disease control by multidimensional flow cytometry

PubMed Central

Pessoa de Magalhães, Roberto J.; Vidriales, María-Belén; Paiva, Bruno; Fernandez-Gimenez, Carlos; García-Sanz, Ramón; Mateos, Maria-Victoria; Gutierrez, Norma C.; Lecrevisse, Quentin; Blanco, Juan F; Hernández, Jose; de las Heras, Natalia; Martinez-Lopez, Joaquin; Roig, Monica; Costa, Elaine Sobral; Ocio, Enrique M.; Perez-Andres, Martin; Maiolino, Angelo; Nucci, Marcio; De La Rubia, Javier; Lahuerta, Juan-Jose; San-Miguel, Jesús F.; Orfao, Alberto

2013-01-01

Multiple myeloma remains largely incurable. However, a few patients experience more than 10 years of relapse-free survival and can be considered as operationally cured. Interestingly, long-term disease control in multiple myeloma is not restricted to patients with a complete response, since some patients revert to having a profile of monoclonal gammopathy of undetermined significance. We compared the distribution of multiple compartments of lymphocytes and dendritic cells in the bone marrow and peripheral blood of multiple myeloma patients with long-term disease control (n=28), patients with newly diagnosed monoclonal gammopathy of undetermined significance (n=23), patients with symptomatic multiple myeloma (n=23), and age-matched healthy adults (n=10). Similarly to the patients with monoclonal gammopathy of undetermined significance and symptomatic multiple myeloma, patients with long-term disease control showed an expansion of cytotoxic CD8+ T cells and natural killer cells. However, the numbers of bone marrow T-regulatory cells were lower in patients with long-term disease control than in those with symptomatic multiple myeloma. It is noteworthy that B cells were depleted in patients with monoclonal gammopathy of undetermined significance and in those with symptomatic multiple myeloma, but recovered in both the bone marrow and peripheral blood of patients with long-term disease control, due to an increase in normal bone marrow B-cell precursors and plasma cells, as well as pre-germinal center peripheral blood B cells. The number of bone marrow dendritic cells and tissue macrophages differed significantly between patients with long-term disease control and those with symptomatic multiple myeloma, with a trend to cell count recovering in the former group of patients towards levels similar to those found in healthy adults. In summary, our results indicate that multiple myeloma patients with long-term disease control have a constellation of unique immune changes favoring both immune cytotoxicity and recovery of B-cell production and homing, suggesting improved immune surveillance. PMID:22773604

Alpha 1 adrenergic receptor mediated polyphosphoinositide breakdown in DDT1-MF2 cells. Lack of evidence of desensitization after prolonged exposure to epinephrine.

PubMed

Rosenbaum, J S; Azhar, S; Hoffman, B B

1987-12-15

The DDT1-MF2 cell line is a transformed smooth muscle cell line which is known to possess both alpha 1 and beta 2 adrenergic receptors. We have utilized these cells to compare the effects of epinephrine pretreatment on the functional capabilities of these two different adrenergic receptors. Pretreatment of the cells grown in suspension with 10(-7) M epinephrine for 6 hr resulted in desensitization of beta receptor stimulated cyclic AMP accumulation. The maximal response to isoproterenol was decreased to 46 +/- 6% of the value in controls (P less than 0.05); there was also a decrease in the sensitivity of the cells to isoproterenol (log EC50 = -6.65 +/- 0.22 vs -7.26 +/- 0.11 in controls, P less than 0.05). Also, there was a decrease in the number of beta receptors from 257 +/- 29 to 163 +/- 22 fmol/mg protein. In contrast, pretreatment with 10(-6) M epinephrine for 6 hr failed to induce a loss of sensitivity in the ability of the alpha 1 receptor agonist phenylephrine to stimulate inositol triphosphate accumulation (log EC50 = -5.59 +/- 0.18 vs -5.42 +/- 0.44 in control cells). A 2-fold increase in basal inositol monophosphate accumulation was observed after epinephrine pretreatment (P less than 0.05); however, there was no change in maximal phenylephrine-stimulated inositol monophosphate accumulation in these cells. There was a small decrease in the alpha 1 receptor number after epinephrine pretreatment (Bmax = 457 +/- 89 fmol/mg protein vs 540 +/- 94 in control cells, P less than 0.05). In contrast to epinephrine pretreatment, pretreatment of cells in suspension with 10(-7) M 12-O-tetradecanoylphorbol-13-acetate (TPA) for 15 min resulted in a nearly complete blunting in the ability of both norepinephrine and phenylephrine to stimulate inositol phosphate accumulation: after norepinephrine stimulation, 774 +/- 34 dpm in TPA-pretreated cells vs 2590 +/- 10 in control cells; inositol monophosphate accumulation after phenylephrine stimulation 576 +/- 25 dpm in TPA-pretreated cells vs 1660 +/- 27 in control cells. Basal levels of inositol monophosphate remained unchanged at 544 +/- 28 dpm vs 505 +/- 31 in TPA-pretreated cells compared to control cells. These data indicate that protein kinase C may exert a negative feedback control on the alpha 1 receptor in these cells and that direct activation of protein kinase C by phorbol esters may have a different effect on the alpha 1 adrenergic receptor system in DDT1-MF2 cells than does prolonged exposure to epinephrine.

T Cell Receptor Signaling in the Control of Regulatory T Cell Differentiation and Function

PubMed Central

Li, Ming O.; Rudensky, Alexander Y.

2016-01-01

Regulatory T cells (TReg cells), a specialized T cell lineage, have a pivotal function in the control of self-tolerance and inflammatory responses. Recent studies have revealed a discrete mode of TCR signaling that regulates Treg cell differentiation, maintenance and function and that impacts on gene expression, metabolism, cell adhesion and migration of these cells. Here, we discuss the emerging understanding of TCR-guided differentiation of Treg cells in the context of their function in health and disease. PMID:27026074

Fuel cell water transport

DOEpatents

Vanderborgh, Nicholas E.; Hedstrom, James C.

1990-01-01

The moisture content and temperature of hydrogen and oxygen gases is regulated throughout traverse of the gases in a fuel cell incorporating a solid polymer membrane. At least one of the gases traverses a first flow field adjacent the solid polymer membrane, where chemical reactions occur to generate an electrical current. A second flow field is located sequential with the first flow field and incorporates a membrane for effective water transport. A control fluid is then circulated adjacent the second membrane on the face opposite the fuel cell gas wherein moisture is either transported from the control fluid to humidify a fuel gas, e.g., hydrogen, or to the control fluid to prevent excess water buildup in the oxidizer gas, e.g., oxygen. Evaporation of water into the control gas and the control gas temperature act to control the fuel cell gas temperatures throughout the traverse of the fuel cell by the gases.

Reaction temperature sensing (RTS)-based control for Li-ion battery safety

PubMed Central

Zhang, Guangsheng; Cao, Lei; Ge, Shanhai; Wang, Chao-Yang; Shaffer, Christian E.; Rahn, Christopher D.

2015-01-01

We report reaction temperature sensing (RTS)-based control to fundamentally enhance Li-ion battery safety. RTS placed at the electrochemical interface inside a Li-ion cell is shown to detect temperature rise much faster and more accurately than external measurement of cell surface temperature. We demonstrate, for the first time, that RTS-based control shuts down a dangerous short-circuit event 3 times earlier than surface temperature- based control and prevents cell overheating by 50 °C and the resultant cell damage. PMID:26658957

Optimal control of fuel overpressure in a polymer electrolyte membrane fuel cell with hydrogen transfer leak during load change

NASA Astrophysics Data System (ADS)

Ebadighajari, Alireza; DeVaal, Jake; Golnaraghi, Farid

2017-02-01

Formation of membrane pinholes is a common defect in fuel cells, inflicting more cost and making less durable cells. This work focuses on mitigating this issue, and offers a continuous online treatment instead of attempting to dynamically model the hydrogen transfer leak rate. This is achieved by controlling the differential pressure between the anode and cathode compartments at the inlet side of the fuel cell stack, known as the fuel overpressure. The model predictive control approach is used to attain the objectives in a Ballard 9-cell Mk1100 polymer electrolyte membrane fuel cell (PEMFC) with inclusion of hydrogen transfer leak. Furthermore, the pneumatic modeling technique is used to model the entire anode side of a fuel cell station. The hydrogen transfer leak is embedded in the model in a novel way, and is considered as a disturbance during the controller design. Experimental results for different sizes of hydrogen transfer leaks are provided to show the benefits of fuel overpressure control system in alleviating the effects of membrane pinholes, which in turn increases membrane longevity, and reduces hydrogen emissions in the eventual presence of transfer leaks. Moreover, the model predictive controller provides an optimal control input while satisfying the problem constraints.

On controllability and system constraints of the linear models of proton exchange membrane and solid oxide fuel cells

NASA Astrophysics Data System (ADS)

Radisavljevic, Verica

2011-10-01

In this paper we first show that the linear models of proton exchange membrane (polymer electrolyte membrane, PEM) and solid oxide (SO) fuel cells, commonly used in power and energy literature, are not controllable. The source of uncontrollability is the equation for pressure of the water vapor that is only affected by the fuel cell current, which in fact is a disturbance in this system and cannot be controlled by the given model inputs: inlet molar flow rates of hydrogen and oxygen. Being uncontrollable these models are not good candidates for studying control of dynamic processes in PEM and SO fuel cells. However, due to their simplicity, they can be used in hybrid configurations with other energy producing devices such as photovoltaic (solar) cells, wind turbine, micro gas turbine, battery (ultra capacitor) to demonstrate some other phenomena, but not for control purposes unless the hybrid models formed in such hybrid configurations are controllable. Testing controllability of such hybrid models is mandatory. Secondly, we introduce some algebraic constraints that follow from the model dynamics and the Nernst open-loop fuel cell voltage formula. These constraints must be satisfied in simulation of considered fuel cell modes, for example, via MATLAB/Simulink or any other computer software package.

Design and development of microbioreactors for long-term cell culture in controlled oxygen microenvironments.

PubMed

Abaci, Hasan E; Devendra, Raghavendra; Smith, Quinton; Gerecht, Sharon; Drazer, German

2012-02-01

The ability to control the oxygen level to which cells are exposed in tissue culture experiments is crucial for many applications. Here, we design, develop and test a microbioreactor (MBR) for long-term cell culture studies with the capability to accurately control and continuously monitor the dissolved oxygen (DO) level in the cell microenvironment. In addition, the DO level can be controlled independently from other cues, such as the viscous shear-stress acting on the cells. We first analyze the transport of oxygen in the proposed device and determine the materials and dimensions that are compatible with uniform oxygen tension and low shear-stress at the cell level. The device is also designed to culture a statistically significant number of cells. We use fully transparent materials and the overall design of the device is compatible with live-cell imaging. The proposed system includes real-time read-out of actual DO levels, is simple to fabricate at low cost, and can be easily expanded to control the concentration of other microenvironmental solutes. We performed control experiments in the absence of cells to demonstrate that the MBR can be used to accurately modulate DO levels ranging from atmospheric level to 1%, both under no flow and perfusion conditions. We also demonstrate cancer cell attachment and viability within the MBR. The proposed MBR offers the unprecedented capability to perform on-line measurement and analysis of DO levels in the microenvironment of adherent cultures and to correlate them with various cellular responses.

Modification of a neuronal network direction using stepwise photo-thermal etching of an agarose architecture.

PubMed

Suzuki, Ikurou; Sugio, Yoshihiro; Moriguchi, Hiroyuki; Jimbo, Yasuhiko; Yasuda, Kenji

2004-07-01

Control over spatial distribution of individual neurons and the pattern of neural network provides an important tool for studying information processing pathways during neural network formation. Moreover, the knowledge of the direction of synaptic connections between cells in each neural network can provide detailed information on the relationship between the forward and feedback signaling. We have developed a method for topographical control of the direction of synaptic connections within a living neuronal network using a new type of individual-cell-based on-chip cell-cultivation system with an agarose microchamber array (AMCA). The advantages of this system include the possibility to control positions and number of cultured cells as well as flexible control of the direction of elongation of axons through stepwise melting of narrow grooves. Such micrometer-order microchannels are obtained by photo-thermal etching of agarose where a portion of the gel is melted with a 1064-nm infrared laser beam. Using this system, we created neural network from individual Rat hippocampal cells. We were able to control elongation of individual axons during cultivation (from cells contained within the AMCA) by non-destructive stepwise photo-thermal etching. We have demonstrated the potential of our on-chip AMCA cell cultivation system for the controlled development of individual cell-based neural networks.

Cell cycle dependent changes in the plasma membrane organization of mammalian cells.

PubMed

Denz, Manuela; Chiantia, Salvatore; Herrmann, Andreas; Mueller, Peter; Korte, Thomas; Schwarzer, Roland

2017-03-01

Lipid membranes are major structural elements of all eukaryotic and prokaryotic organisms. Although many aspects of their biology have been studied extensively, their dynamics and lateral heterogeneity are still not fully understood. Recently, we observed a cell-to-cell variability in the plasma membrane organization of CHO-K1 cells (Schwarzer et al., 2014). We surmised that cell cycle dependent changes of the individual cells from our unsynchronized cell population account for this phenomenon. In the present study, this hypothesis was tested. To this aim, CHO-K1 cells were arrested in different cell cycle phases by chemical treatments, and the order of their plasma membranes was determined by various fluorescent lipid analogues using fluorescence lifetime imaging microscopy. Our experiments exhibit significant differences in the membrane order of cells arrested in the G2/M or S phase compared to control cells. Our single-cell analysis also enabled the specific selection of mitotic cells, which displayed a significant increase of the membrane order compared to the control. In addition, the lipid raft marker GPImYFP was used to study the lateral organization of cell cycle arrested cells as well as mitotic cells and freely cycling samples. Again, significant differences were found between control and arrested cells and even more pronounced between control and mitotic cells. Our data demonstrate a direct correlation between cell cycle progression and plasma membrane organization, underlining that cell-to-cell heterogeneities of membrane properties have to be taken into account in cellular studies especially at the single-cell level. Copyright © 2016 Elsevier B.V. All rights reserved.

Breaking the rules? X-ray examination of hematopoietic stem cell grafts at international airports.

PubMed

Petzer, Andreas L; Speth, Hans-Georg; Hoflehner, Elisabeth; Clausen, Johannes; Nachbaur, David; Gastl, Günther; Gunsilius, Eberhard

2002-06-15

Hematopoietic stem cell grafts from unrelated donors are commonly transported by aircraft. They must not be subjected to x-rays during security checks, which may cause inconvenient discussions between the courier and the airport security staff. We exposed hematopoietic stem cells from mobilized peripheral blood to a widely used x-ray hand-luggage control system. Cell viability as well as growth in vitro of mature progenitor cells (colony-forming cells), primitive progenitor cells (long-term culture-initiating cells), and lymphocytes were not altered even after 10 passages through the hand-luggage control system. Thus, repeated exposure to the low radiation dose of hand-luggage control systems (1.5 +/- 0.6 microSv per exposure) seems to be harmless for hematopoietic stem cells, which should simplify the international transport of stem cell grafts.

[Effect of low-energy 633 nm red light stimulation on proliferation and reactive oxygen species level of human epidermal cell line HaCaT].

PubMed

Chen, Z Y; Li, D L; Duan, X D; Peng, D Z

2016-09-20

To investigate the changes of proliferative activity and reactive oxygen species level of human epidermal cell line HaCaT after being irradiated with low-energy 633 nm red light. Irradiation distance was determined through preliminary experiment. HaCaT cells were conventionally sub-cultured with RPMI 1640 culture medium containing 10% fetal calf serum, 100 U/mL penicillin, and 100 μg/mL streptomycin. Cells of the third passage were used in the following experiments. (1) Cells were divided into blank control group and 0.082, 0.164, 0.245, 0.491, 1.472, 2.453, 4.910, and 9.810 J/cm(2) irradiation groups according to the random number table, with 3 wells in each group. Cells in blank control group were not irradiated, while cells in the latter 8 irradiation groups were irradiated with 633 nm red light for 10, 20, 30, 60, 180, 300, 600, and 1 200 s in turn. Cells were reirradiated once every 8 hours. After being irradiated for 48 hours (6 times) in irradiation groups, the proliferative activity of cells in 9 groups was determined with cell counting kit 8 and microplate reader (denoted as absorbance value). (2) Another batch of cells were grouped and irradiated as in experiment (1). After being irradiated for once in irradiation groups, cells in 9 groups were conventionally cultured for 60 min with detection reagent of reactive oxygen species. At post culture minute (PCM) 0 (immediately), 30, 60, and 120, reactive oxygen species level of cells was determined with microplate reader (denoted as absorbance value). (3) Another batch of cells were divided into blank control group, 0.082, 0.491, 2.453, and 9.810 J/cm(2) irradiation groups, and positive control group. Cells in blank control group and positive control group were not irradiated (positive control reagent of reactive oxygen species was added to cells in positive control group), and cells in irradiation groups were irradiated as in experiment (1) for once. The expression of reactive oxygen species in cells of each group was observed by confocal laser scanning microscope. Data were processed with one-way analysis of variance, analysis of variance for repeated measurement, and t test. (1) Irradiation distance was 10 cm. Proliferative activity of cells in blank control group and 0.082, 0.164, 0.245, 0.491, 1.472, 2.453, 4.910, and 9.810 J/cm(2) irradiation groups was 1.000, 1.116±0.031, 1.146±0.016, 1.162±0.041, 1.179±0.016, 1.207±0.016, 1.247±0.040, 1.097±0.059, and 0.951±0.118, respectively. Compared with that in blank control group, proliferative activity of cells in 0.082-2.453 J/cm(2) irradiation groups was significantly higher (with t values from -22.803 to -6.779, P values below 0.05). Proliferative activity of cells in 4.910 and 9.810 J/cm(2) irradiation groups was similar to that in blank control group (with t values respectively -2.854 and 0.711, P values above 0.05). (2) Compared with that in blank control group, reactive oxygen species level of cells was significantly enhanced at PCM 0 and 30 in 0.164-2.453 J/cm(2) irradiation groups (with t values from -12.453 to -4.684, P<0.05 or P<0.01), while that showed no significant change in 0.082, 4.910, and 9.810 J/cm(2) irradiation groups (with t values from -3.925 to -0.672, P values above 0.05). Compared with that in blank control group, reactive oxygen species level of cells was significantly enhanced at PCM 60 in 0.082-2.453 J/cm(2) irradiation groups (with t values from -11.387 to -4.717, P<0.05 or P<0.01). Compared with that in blank control group, reactive oxygen species level of cells was significantly enhanced at PCM 120 in 0.491-2.453 J/cm(2) irradiation groups (with t values from -10.657 to -6.644, P<0.05 or P<0.01). (3) Compared with that in blank control group, the expression of reactive oxygen species of cells was increased in 0.082, 0.491, and 2.453 J/cm(2) irradiation groups and positive control group. The expression of reactive oxygen species of cells in 9.810 J/cm(2) irradiation group was attenuated when compared with the expressions in the other irradiation groups. Reactive oxygen species expressed in mitochondria of cells in each group. Low-energy 633 nm red light can enhance the proliferation of human epidermal cell line HaCaT, and the effect is closely related to the increase of reactive oxygen species produced by mitochondria after being stimulated by red light irradiation.

Gag-Positive Reservoir Cells Are Susceptible to HIV-Specific Cytotoxic T Lymphocyte Mediated Clearance In Vitro and Can Be Detected In Vivo

PubMed Central

Graf, Erin H.; Pace, Matthew J.; Peterson, Bennett A.; Lynch, Lindsay J.; Chukwulebe, Steve B.; Mexas, Angela M.; Shaheen, Farida; Martin, Jeffrey N.; Deeks, Steven G.; Connors, Mark; Migueles, Stephen A.; O’Doherty, Una

2013-01-01

Resting CD4+ T cells infected with HIV persist in the presence of suppressive anti-viral therapy (ART) and are barriers to a cure. One potential curative approach, therapeutic vaccination, is fueled by recognition of the ability of a subset of elite controllers (EC) to control virus without therapy due to robust anti-HIV immune responses. Controllers have low levels of integrated HIV DNA and low levels of replication competent virus, suggesting a small reservoir. As our recent data indicates some reservoir cells can produce HIV proteins (termed GPR cells for Gag-positive reservoir cells), we hypothesized that a fraction of HIV-expressing resting CD4+ T cells could be efficiently targeted and cleared in individuals who control HIV via anti-HIV cytotoxic T lymphocytes (CTL). To test this we examined if superinfected resting CD4+ T cells from EC express HIV Gag without producing infectious virus and the susceptibility of these cells to CTL. We found that resting CD4+ T cells expressed HIV Gag and were cleared by autologous CD8+ T cells from EC. Importantly, we found the extent of CTL clearance in our in vitro assay correlates with in vivo reservoir size and that a population of Gag expressing resting CD4+ T cells exists in vivo in patients well controlled on therapy. PMID:23951263

Cell fate reprogramming by control of intracellular network dynamics

NASA Astrophysics Data System (ADS)

Zanudo, Jorge G. T.; Albert, Reka

Identifying control strategies for biological networks is paramount for practical applications that involve reprogramming a cell's fate, such as disease therapeutics and stem cell reprogramming. Although the topic of controlling the dynamics of a system has a long history in control theory, most of this work is not directly applicable to intracellular networks. Here we present a network control method that integrates the structural and functional information available for intracellular networks to predict control targets. Formulated in a logical dynamic scheme, our control method takes advantage of certain function-dependent network components and their relation to steady states in order to identify control targets, which are guaranteed to drive any initial state to the target state with 100% effectiveness and need to be applied only transiently for the system to reach and stay in the desired state. We illustrate our method's potential to find intervention targets for cancer treatment and cell differentiation by applying it to a leukemia signaling network and to the network controlling the differentiation of T cells. We find that the predicted control targets are effective in a broad dynamic framework. Moreover, several of the predicted interventions are supported by experiments. This work was supported by NSF Grant PHY 1205840.

Cell death induced by Morarah and Khaltita in hepatoma cancer cells (Huh-7).

PubMed

Baig, Saeeda; Alamgir, Mohiuddin

2009-10-01

To compare the combined and isolated growth inhibitory effects of Morarah and Khaltita (herbs) on hepatoma cell lines (Huh-7), through induction of apoptosis or necrosis. Comparative controlled in-vitro study. The Molecular Biology Laboratory, The Aga Khan University, Karachi, from June to December 2006. The growth of hepatoma cell lines (Huh-7) was checked by adding Khaltita and Morarah to the cells before culture in a 24 well plate. Six wells were selected and labeled for each of the four variables (controls, Khaltita, Morarah and mixture). After 2 days, cells were studied under an inverted phase contrast microscope and fields were recorded. Approximately four fields per slide of higher intensity were selected randomly to determine the dead cell density, and the procedure was repeated 10 or more times. Frequency and percentages were calculated for dead or alive cells in controls, Morarah, Khaltita and their mixture. Chi-square was used to compare the qualitative variables. P-values < 0.05 were considered significant. Morarah and Khaltita were found to induce statistically significant (p < 0.001) cell death in hepatoma cell lines (Huh-7). At a magnification of 40x, the controls showed 1% dead cells compared to 91% in Morarah, 83% in Khaltita and 73% in combined mixture of Khaltita and Morarah. At magnification of 20x, the controls showed 4% dead cells compared to 44% in Morarah, 47% in Khaltita and 49% in the combined mixture of Khaltita and Morarah. Morarah and Khaltita induced cell death in cultured hepatoma cells (Huh-7).

Depletion of pro-inflammatory CD161(+) double negative (CD3(+)CD4(-)CD8(-)) T cells in AIDS patients is ameliorated by expansion of the γδ T cell population.

PubMed

Singleterry, Will L; Henderson, Harold; Cruse, Julius M

2012-02-01

In this present investigation, flow cytometry was utilized to evaluate 13 healthy controls and 31 HIV-1 infected patients who had advanced to the AIDS stage of infection (CD4 count below 200 cells/mm(3)), for the expression of CD161 on CD3(+) double negative (DN) (CD3(+)CD4(-)CD8(-)) T cells, CD4(+) T cells, CD8(+) T cells and γδ T cells. The observed depletion of CD161(+) T cells from peripheral circulation was due primarily to the loss of CD4(+)CD161(+) T cells; as these cells represented 8.67±0.74% of the total healthy control peripheral T cell population, while the CD4(+)CD161(+) T cells of the AIDS group represented only 3.35±0.41% (p=<0.0001) of the total peripheral T cell population. We have also shown here that the DN T cell population was more than doubled in the AIDS group, with the DN T cell population expanding from 3.29±0.45% of the healthy control peripheral T cell population to 8.64±1.16% (p=0.0001) of the AIDS group peripheral T cell population. By evaluating the expression of CD161 on the surface of the DN T cells we showed that within the healthy control group, 47.4±4.99% of the DN T cells were positive for the expression of CD161, while only 26.4±3.54% (p=0.002) of the AIDS group's DN T cells expressed CD161. Despite CD161 expression being halved on the DN T cells of the AIDS group, when we compared the total peripheral T cell percentage of CD161(+) DN T cells between the healthy control group and the AIDS group, there was no statistical difference. Even though only 26.4% DN T cells within the AIDS group were positive for CD161(+), the overall DN T cell population had expanded to such an extent that there was no statistical difference between the groups with regard to CD161(+) DN T cells as a percentage of the total peripheral T cell population. Furthermore, we showed that within the DN T cell population, there was an approximate 2:1 ratio of γδ to αβ T cells, and this ratio was maintained in both the healthy control group and the AIDS group. While evaluating γδ T cells we also discovered that CD8(+) γδ T cells were expanded from 0.62±.09% of the healthy control peripheral T cell population to 5.01±.88% (p=<0.0001) of the peripheral T cell population of the AIDS group; and that this population of CD8(+) γδ T cells underwent the same reduction in percentage of cells expressing CD161(+), further demonstrated that the phenomenon of CD161(+) percentage reduction and compensatory increase in total cell population was affecting the entire circulating γδ T cell population. Copyright © 2011 Elsevier Inc. All rights reserved.

Anti-apoptotic BCL-2 family proteins in acute neural injury

PubMed Central

Anilkumar, Ujval; Prehn, Jochen H. M.

2014-01-01

Cells under stress activate cell survival and cell death signaling pathways. Cell death signaling frequently converges on mitochondria, a process that is controlled by the activities of pro- and anti-apoptotic B-cell lymphoma 2 (BCL-2) proteins. In this review, we summarize current knowledge on the control of neuronal survival, development and injury by anti-apoptotic BCL-2 family proteins. We discuss overlapping and differential effects of the individual family members BCL-2, BCL-extra long (BCL-XL), myeloid cell leukemia 1 (MCL-1), and BCL2-like 2 (BCL-W) in the control of survival during development and pathophysiological processes such as trophic factor withdrawal, ischemic injury, excitotoxicity, oxidative stress and energy stress. Finally we discuss recent evidence that several anti-apoptotic BCL-2 proteins influence mitochondrial bioenergetics and control neuronal Ca2+ homeostasis independent of their classical role in cell death signaling. PMID:25324720

Anti-apoptotic BCL-2 family proteins in acute neural injury.

PubMed

Anilkumar, Ujval; Prehn, Jochen H M

2014-01-01

Cells under stress activate cell survival and cell death signaling pathways. Cell death signaling frequently converges on mitochondria, a process that is controlled by the activities of pro- and anti-apoptotic B-cell lymphoma 2 (BCL-2) proteins. In this review, we summarize current knowledge on the control of neuronal survival, development and injury by anti-apoptotic BCL-2 family proteins. We discuss overlapping and differential effects of the individual family members BCL-2, BCL-extra long (BCL-XL), myeloid cell leukemia 1 (MCL-1), and BCL2-like 2 (BCL-W) in the control of survival during development and pathophysiological processes such as trophic factor withdrawal, ischemic injury, excitotoxicity, oxidative stress and energy stress. Finally we discuss recent evidence that several anti-apoptotic BCL-2 proteins influence mitochondrial bioenergetics and control neuronal Ca(2+) homeostasis independent of their classical role in cell death signaling.

Chronophin coordinates cell leading edge dynamics by controlling active cofilin levels

PubMed Central

Delorme-Walker, Violaine; Seo, Ji-Yeon; Gohla, Antje; Fowler, Bruce; Bohl, Ben; DerMardirossian, Céline

2015-01-01

Cofilin, a critical player of actin dynamics, is spatially and temporally regulated to control the direction and force of membrane extension required for cell locomotion. In carcinoma cells, although the signaling pathways regulating cofilin activity to control cell direction have been established, the molecular machinery required to generate the force of the protrusion remains unclear. We show that the cofilin phosphatase chronophin (CIN) spatiotemporally regulates cofilin activity at the cell edge to generate persistent membrane extension. We show that CIN translocates to the leading edge in a PI3-kinase–, Rac1-, and cofilin-dependent manner after EGF stimulation to activate cofilin, promotes actin free barbed end formation, accelerates actin turnover, and enhances membrane protrusion. In addition, we establish that CIN is crucial for the balance of protrusion/retraction events during cell migration. Thus, CIN coordinates the leading edge dynamics by controlling active cofilin levels to promote MTLn3 cell protrusion. PMID:26324884

Novel device for continuous spatial control and temporal delivery of nitric oxide for in vitro cell culture☆

PubMed Central

Romanowicz, Genevieve E.; He, Weilue; Nielsen, Matthew; Frost, Megan C.

2013-01-01

Nitric oxide (NO) is an ubiquitous signaling molecule of intense interest in many physiological processes. Nitric oxide is a highly reactive free radical gas that is difficult to deliver with precise control over the level and timing that cells actually experience. We describe and characterize a device that allows tunable fluxes and patterns of NO to be generated across the surface upon which cells are cultured. The system is based on a quartz microscope slide that allows for controlled light levels to be applied to a previously described photosensitive NO-releasing polydimethylsiloxane (PDMS). Cells are cultured in separate wells that are either NO-releasing or a chemically similar PDMS that does not release NO. Both wells are then top coated with DowCorning RTV-3140 PDMS and a polydopamine/gelatin layer to allow cells to grow in the culture wells. When the waveguide is illuminated, the surface of the quartz slide propagates light such that the photosensitive polymer is evenly irradiated and generates NO across the surface of the cell culture well and no light penetrates into the volume of the wells where cells are growing. Mouse smooth muscle cells (MOVAS) were grown in the system in a proof of principle experiment, whereby 60% of the cells were present in the NO-releasing well compared to control wells after 17 h. The compelling advantage of illuminating the NO-releasing polymers with the waveguide system is that light can be used to tunably control NO release while avoiding exposing cells to optical radiation. This device provides means to quantitatively control the surface flux, timing and duration of NO cells experience and allows for systematic study of cellular response to NO generated at the cell/surface interface in a wide variety of studies. PMID:24024168

Chemical Control over T-Cell Activation in Vivo Using Deprotection of trans-Cyclooctene-Modified Epitopes.

PubMed

van der Gracht, Anouk M F; de Geus, Mark A R; Camps, Marcel G M; Ruckwardt, Tracy J; Sarris, Alexi J C; Bremmers, Jessica; Maurits, Elmer; Pawlak, Joanna B; Posthoorn, Michelle M; Bonger, Kimberly M; Filippov, Dmitri V; Overkleeft, Herman S; Robillard, Marc S; Ossendorp, Ferry; van Kasteren, Sander I

2018-06-15

Activation of a cytotoxic T-cell is a complex multistep process, and tools to study the molecular events and their dynamics that result in T-cell activation in situ and in vivo are scarce. Here, we report the design and use of conditional epitopes for time-controlled T-cell activation in vivo. We show that trans-cyclooctene-protected SIINFEKL (with the lysine amine masked) is unable to elicit the T-cell response characteristic for the free SIINFEKL epitope. Epitope uncaging by means of an inverse-electron demand Diels-Alder (IEDDA) event restored T-cell activation and provided temporal control of T-cell proliferation in vivo.

Impute DC link (IDCL) cell based power converters and control thereof

DOEpatents

Divan, Deepakraj M.; Prasai, Anish; Hernendez, Jorge; Moghe, Rohit; Iyer, Amrit; Kandula, Rajendra Prasad

2016-04-26

Power flow controllers based on Imputed DC Link (IDCL) cells are provided. The IDCL cell is a self-contained power electronic building block (PEBB). The IDCL cell may be stacked in series and parallel to achieve power flow control at higher voltage and current levels. Each IDCL cell may comprise a gate drive, a voltage sharing module, and a thermal management component in order to facilitate easy integration of the cell into a variety of applications. By providing direct AC conversion, the IDCL cell based AC/AC converters reduce device count, eliminate the use of electrolytic capacitors that have life and reliability issues, and improve system efficiency compared with similarly rated back-to-back inverter system.

Pro-Inflammatory cytokines increases hepatocellular carcinoma cells thermotolerance: Evidence of how local inflammation may negatively impact radiofrequency ablation local control rates

PubMed Central

Douglas, Wade G.; Wang, Yangping; Gibbs, John F.; Tracy, Erin; Kuvshinoff, Boris; Huntoon, Kristin; Baumann, Heinz

2008-01-01

Background Hepatocellular carcinomas (HCC) associated with inflammation that undergo radiofrequency ablation (RFA) appear to have poorer local control rates. Little is known of how mediators of inflammation influence HCC cellular thermotolerance which in part is mediated by heat shock protein 70 (HSP 70). This study determines how inflammatory mediators effect cellular thermotolerance and provides insight into how associated inflammation may impact HCC RFA local control rates. Methods HepG2 cell lines were cultured in control medium (CM) or CM containing conditioned medium of endotoxin-activated macrophage (CMM). Serial dilutions of CMM established microenvironments approximating low, medium and high CMM. All groups underwent a heat shock challenge (HSC) at 45° C for 10 minutes. Western blot, northern blot, densometric analysis, along with Thymidine and clonagenic assays determined how inflammation influenced multiple biologic endpoints. Results Cells cultured in low CMM, expressed significantly more HSP 70 RNA and protein compared to control cells after HSC. The cells also had a higher proliferative and survival rate after HSC compared to control cells. Medium CMM cultured cells had no significant difference in HSP 70 RNA and protein production or proliferation and survival rates after HSC, compared to CM cultured cells. AT high CMM the inhibitory effects of inflammatory mediators prevailed, all the measured endpoints were significantly less compared to CM cultured cells. Conclusions This study demonstrates that inflammation can alter the responsiveness of HCC cells to a HSC in a dose dependent manner. This study supports the clinical observation that HCC associated with chronic inflammation have worse RFA local control rates. PMID:18262552

[Biologic effects of different concentrations of putrescine on human umbilical vein endothelial cells].

PubMed

Chen, Jianxia; Rong, Xinzhou; Fan, Guicheng; Li, Songze; Zhang, Tao; Li, Qinghui

2015-12-01

To explore the effects of different concentrations of putrescine on proliferation, migration, and apoptosis of human umbilical vein endothelial cells (HUVECs). HUVECs were routinely cultured in vitro. The 3rd to the 5th passage of HUVECs were used in the following experiments. (1) Cells were divided into 500, 1 000, and 5 000 µg/mL putrescine groups according to the random number table (the same grouping method was used for following grouping), with 3 wells in each group, which were respectively cultured with complete culture solution containing putrescine in the corresponding concentration for 24 h. Morphology of cells was observed by inverted optical microscope. (2) Cells were divided into 0.5, 1.0, 5.0, 10.0, 50.0, 100.0, 500.0, 1 000.0 µg/mL putrescine groups, and control group, with 4 wells in each group. Cells in the putrescine groups were respectively cultured with complete culture solution containing putrescine in the corresponding concentration for 24 h, and cells in control group were cultured with complete culture solution with no additional putrescine for 24 h. Cell proliferation activity (denoted as absorption value) was measured by colorimetry. (3) Cells were divided (with one well in each group) and cultured as in experiment (2), and the migration ability was detected by transwell migration assay. (4) Cells were divided (with one flask in each group) and cultured as in experiment (2), and the cell apoptosis rate was determined by flow cytometer. Data were processed with one-way analysis of variance, Kruskal-Wallis test, and Dunnett test. (1) After 24-h culture, cell attachment was good in 500 µg/mL putrescine group, and no obvious change in the shape was observed; cell attachment was less in 1 000 µg/mL putrescine group and the cells were small and rounded; cells in 5 000 µg/mL putrescine group were in fragmentation without attachment. (2) The absorption values of cells in 0.5, 1.0, 5.0, 10.0, 50.0, 100.0, 500.0, 1 000.0 µg/mL putrescine groups, and control group were respectively 0.588 ± 0.055, 0.857 ± 0.031, 0.707 ± 0.031, 0.662 ± 0.023, 0.450 ± 0.019, 0.415 ± 0.014, 0.359 ± 0.020, 0.204 ± 0.030, and 0.447 ± 0.021, with statistically significant differences among them (χ(2) = 6.86, P = 0.009). The cell proliferation activity in 0.5, 1.0, 5.0, and 10.0 µg/mL putrescine groups was higher than that in control group (P < 0.05 or P < 0.01). The cell proliferation activity in 500.0 and 1 000.0 µg/mL putrescine groups was lower than that in control group (with P values below 0.01). The cell proliferation activity in 50.0 and 100.0 µg/mL putrescine groups was close to that in control group (with P values above 0.05). (3) There were statistically significant differences in the numbers of migrated cells between the putrescine groups and control group (F = 138.662, P < 0.001). The number of migrated cells was more in 1.0, 5.0, and 10.0 µg/mL putrescine groups than in control group (with P value below 0.01). The number of migrated cells was less in 500.0 and 1 000.0 µg/mL putrescine groups than in control group (with P value below 0.01). The number of migrated cells in 0.5, 50.0, and 100.0 µg/mL putrescine groups was close to that in control group (with P values above 0.05). (4) There were statistically significant differences in the apoptosis rate between the putrescine groups and control group (χ(2)=3.971, P=0.046). The cell apoptosis rate was lower in 0.5, 1.0, 5.0, and 10.0 µg/mL putrescine groups than in control group (with P values below 0.05). The cell apoptosis rate was higher in 500.0 and 1 000.0 µg/mL putrescine groups than in control group (with P values below 0.01). The cell apoptosis rates in 50.0 and 100.0 µg/mL putrescine groups were close to the cell apoptosis rate in control group (with P values above 0.05). Low concentration of putrescine can remarkably enhance the ability of proliferation and migration of HUVECs, while a high concentration of putrescine can obviously inhibit HUVECs proliferation and migration, and it induces apoptosis.

Supplementation of conventional freezing medium with a combination of catalase and trehalose results in better protection of surface molecules and functionality of hematopoietic cells.

PubMed

Sasnoor, Lalita M; Kale, Vaijayanti P; Limaye, Lalita S

2003-10-01

Our previous studies had shown that a combination of the bio-antioxidant catalase and the membrane stabilizer trehalose in the conventional freezing mixture affords better cryoprotection to hematopoietic cells as judged by clonogenic assays. In the present investigation, we extended these studies using several parameters like responsiveness to growth factors, expression of growth factor receptors, adhesion assays, adhesion molecule expression, and long-term culture-forming ability. Cells were frozen with (test cells) or without additives (control cells) in the conventional medium containing 10% dimethylsulfoxide (DMSO). Experiments were done on mononuclear cells (MNC) from cord blood/fetal liver hematopoietic cells (CB/FL) and CD34(+) cells isolated from frozen MNC. Our results showed that the responsiveness of test cells to the two early-acting cytokines, viz. interleukin-3 (IL-3) and stem cell factor (SCF) in CFU assays was better than control cells as seen by higher colony formation at limiting concentrations of these cytokines. We, therefore, analyzed the expression of these two growth factor receptors by flow cytometry. We found that in cryopreserved test MNC, as well as CD34(+) cells isolated from them, the expression of both cytokine receptors was two- to three-fold higher than control MNC and CD34(+) cells isolated from them. Adhesion assays carried out with CB/FL-derived CD34(+) cells and KG1a cells showed significantly higher adherence of test cells to M210B4 than respective control cells. Cryopreserved test MNC as well as CD34(+) cells isolated from them showed increased expression of adhesion molecules like CD43, CD44, CD49d, and CD49e. On isolated CD34(+) cells and KG1a cells, there was a two- to three-fold increase in a double-positive population expressing CD34/L-selectin in test cells as compared to control cells. Long-term cultures (LTC) were set up with frozen MNC as well as with CD34(+) cells. Clonogenic cells from LTC were enumerated at the end of the fifth week. There was a significantly increased formation of CFU from test cells than from control cells, indicating better preservation of early progenitors in test cells. Our results suggest that use of a combination of catalase and trehalose as a supplement in the conventional freezing medium results in better protection of growth factor receptors, adhesion molecules, and functionality of hematopoietic cells, yielding a better graft quality.

Boosting airway T-regulatory cells by gastrointestinal stimulation as a strategy for asthma control.

PubMed

Strickland, D H; Judd, S; Thomas, J A; Larcombe, A N; Sly, P D; Holt, P G

2011-01-01

The hallmark of atopic asthma is transient airways hyperresponsiveness (AHR) preceded by aeroallergen-induced Th-cell activation. This is preceded by upregulation of CD86 on resident airway dendritic cells (DCs) that normally lack competence in T-cell triggering. Moreover, AHR duration is controlled via T-regulatory (Treg) cells, which can attenuate CD86 upregulation on DC. We show that airway mucosal Treg/DC interaction represents an accessible therapeutic target for asthma control. Notably, baseline airway Treg activity in sensitized rats can be boosted by microbe-derived stimulation of the gut, resulting in enhanced capacity to control CD86 expression on airway DC triggered by aeroallergen and accelerated resolution of AHR.

Microfluidic-Based Generation of Size-Controlled, Biofunctionalized Synthetic Polymer Microgels for Cell Encapsulation

PubMed Central

Headen, Devon M.; Aubry, Guillaume; Lu, Hang

2014-01-01

Cell and islet microencapsulation in synthetic hydrogels provide an immunoprotective and cell-supportive microenvironment. A microfluidic strategy for the genaration of biofunctionalized, synthetic microgel particles with precise control over particle size and molecular permeability for cell and protein delivery is presented. These engineered capsules support high cell viability and function of encapsulated human stem cells and islets. PMID:24615922

Activation of Rho GTPase Cdc42 promotes adhesion and invasion in colorectal cancer cells.

PubMed

Gao, Lei; Bai, Lan; Nan, Qing zhen

2013-07-25

The purpose of this study was to investigate the role of activated Rho GTPase cell division control protein 42 homolog (Cdc42) in colorectal cancer cell adhesion, migration, and invasion. The constitutively active form of Cdc42 (GFP-Cdc42L61) or control vector was overexpressed in the colorectal cancer cell line SW480. The localization of active Cdc42 was monitored by immunofluorescence staining, and the effects of active Cdc42 on cell migration and invasion were examined using an attachment assay, a wound healing assay, and a Matrigel migration assay in vitro. Immunofluorescence staining revealed that constitutively active Cdc42 predominately localized to the plasma membrane. Compared to SW480 cells transfected with the control vector, overexpression of constitutively active Cdc42 in SW480 cells promoted filopodia formation and cell stretch and dramatically enhanced cell adhesion to the coated plates. The wound healing assay revealed a significant increase of migration capability in SW480 cells expressing active Cdc42 compared to the control cells. Additionally, the Matrigel invasion assay demonstrated that active Cdc42 significantly promoted SW480 cell migration through the chamber. Our results suggest that active Rho GTPase Cdc42 can greatly enhance colorectal cancer cell SW480 to spread, migrate, and invade, which may contribute to colorectal cancer metastasis.

Selective deletion of Connexin 40 in renin-producing cells impairs renal baroreceptor function and is associated with arterial hypertension

PubMed Central

Wagner, Charlotte; Jobs, Alexander; Schweda, Frank; Kurtz, Lisa; Kurt, Birguel; Sequeira Lopez, Maria L.; Gomez, R. Ariel; van Veen, Toon A.B.; de Wit, Cor; Kurtz, Armin

2011-01-01

Renin-producing juxtaglomerular cells are connected to each other and to endothelial cells of afferent arterioles by gap junctions containing Connexin 40 (Cx40), abundantly expressed by these two cell types. Here, we generated mice with cell-specific deletion of Cx40 in endothelial and in renin-producing cells, as its global deletion caused local dissociation of renin-producing cells from endothelial cells, renin hypersecretion, and hypertension. In mice lacking endothelial Cx40, the blood pressure, renin-producing cell distribution, and the control of renin secretion were similar to wild-type mice. In contrast, mice deficient for Cx40 in renin-producing cells were hypertensive and these cells were ectopically localized. Although plasma renin activity and kidney renin mRNA levels of these mice were not different from controls, the negative regulation of renin secretion by pressure was inverted to a positive feedback in kidneys lacking Cx40 in renin-producing cells. Thus, our findings show that endothelial Cx40 is not essential for the control of renin expression and/or release. Cx40 in renin-producing cells is required for their correct positioning in the juxtaglomerular area and the control of renin secretion by pressure. PMID:20686449

Par-aPKC-dependent and -independent mechanisms cooperatively control cell polarity, Hippo signaling, and cell positioning in 16-cell stage mouse embryos.

PubMed

Hirate, Yoshikazu; Hirahara, Shino; Inoue, Ken-Ichi; Kiyonari, Hiroshi; Niwa, Hiroshi; Sasaki, Hiroshi

2015-10-01

In preimplantation mouse embryos, the Hippo signaling pathway plays a central role in regulating the fates of the trophectoderm (TE) and the inner cell mass (ICM). In early blastocysts with more than 32 cells, the Par-aPKC system controls polarization of the outer cells along the apicobasal axis, and cell polarity suppresses Hippo signaling. Inactivation of Hippo signaling promotes nuclear accumulation of a coactivator protein, Yap, leading to induction of TE-specific genes. However, whether similar mechanisms operate at earlier stages is not known. Here, we show that slightly different mechanisms operate in 16-cell stage embryos. Similar to 32-cell stage embryos, disruption of the Par-aPKC system activated Hippo signaling and suppressed nuclear Yap and Cdx2 expression in the outer cells. However, unlike 32-cell stage embryos, 16-cell stage embryos with a disrupted Par-aPKC system maintained apical localization of phosphorylated Ezrin/Radixin/Moesin (p-ERM), and the effects on Yap and Cdx2 were weak. Furthermore, normal 16-cell stage embryos often contained apolar cells in the outer position. In these cells, the Hippo pathway was strongly activated and Yap was excluded from the nuclei, thus resembling inner cells. Dissociated blastomeres of 8-cell stage embryos form polar-apolar couplets, which exhibit different levels of nuclear Yap, and the polar cell engulfed the apolar cell. These results suggest that cell polarization at the 16-cell stage is regulated by both Par-aPKC-dependent and -independent mechanisms. Asymmetric cell division is involved in cell polarity control, and cell polarity regulates cell positioning and most likely controls Hippo signaling. © The Authors Development, Growth & Differentiation published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Society of Developmental Biologists.

Intracellular integration of synthetic nanostructures with viable cells for controlled biochemical manipulation

NASA Astrophysics Data System (ADS)

McKnight, Timothy E.; Melechko, Anatoli V.; Griffin, Guy D.; Guillorn, Michael A.; Merkulov, Vladimir I.; Serna, Francisco; Hensley, Dale K.; Doktycz, Mitchel J.; Lowndes, Douglas H.; Simpson, Michael L.

2003-05-01

We demonstrate the integration of vertically aligned carbon nanofibre (VACNF) elements with the intracellular domains of viable cells for controlled biochemical manipulation. Deterministically synthesized VACNFs were modified with either adsorbed or covalently-linked plasmid DNA and were subsequently inserted into cells. Post insertion viability of the cells was demonstrated by continued proliferation of the interfaced cells and long-term (> 22 day) expression of the introduced plasmid. Adsorbed plasmids were typically desorbed in the intracellular domain and segregated to progeny cells. Covalently bound plasmids remained tethered to nanofibres and were expressed in interfaced cells but were not partitioned into progeny, and gene expression ceased when the nanofibre was no longer retained. This provides a method for achieving a genetic modification that is non-inheritable and whose extent in time can be directly and precisely controlled. These results demonstrate the potential of VACNF arrays as an intracellular interface for monitoring and controlling subcellular and molecular phenomena within viable cells for applications including biosensors, in vivo diagnostics, and in vivo logic devices.

Cell-controlled hybrid perfusion fed-batch CHO cell process provides significant productivity improvement over conventional fed-batch cultures.

PubMed

Hiller, Gregory W; Ovalle, Ana Maria; Gagnon, Matthew P; Curran, Meredith L; Wang, Wenge

2017-07-01

A simple method originally designed to control lactate accumulation in fed-batch cultures of Chinese Hamster Ovary (CHO) cells has been modified and extended to allow cells in culture to control their own rate of perfusion to precisely deliver nutritional requirements. The method allows for very fast expansion of cells to high density while using a minimal volume of concentrated perfusion medium. When the short-duration cell-controlled perfusion is performed in the production bioreactor and is immediately followed by a conventional fed-batch culture using highly concentrated feeds, the overall productivity of the culture is approximately doubled when compared with a highly optimized state-of-the-art fed-batch process. The technology was applied with near uniform success to five CHO cell processes producing five different humanized monoclonal antibodies. The increases in productivity were due to the increases in sustained viable cell densities. Biotechnol. Bioeng. 2017;114: 1438-1447. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Effect of spaceflight on natural killer cell activity

NASA Technical Reports Server (NTRS)

Rykova, Marina P.; Sonnenfeld, Gerald; Lesniak, A. T.; Taylor, Gerald R.; Meshkov, Dimitrii O.; Mandel, Adrian D.; Medvedev, Andrei E.; Berry, Wallace D.; Fuchs, Boris B.; Konstantinova, Irina V.

1992-01-01

The effects of spaceflight on immune cell function were determined in rats flown on Cosmos 2044. Control groups included vivarium, synchronous, and antiorthostatically suspended rats. The ability of natural killer cells to lyse two different target cell lines was determined. Spleen and bone marrow cells obtained from flight rats showed significantly inhibited cytotoxicity for YAC-1 target cells compared with cells from synchronous control rats. This could have been due to exposure of the rats to microgravity. Antiorthostatic suspension did not affect the level of cytotoxicity from spleen cells of suspended rats for YAC-1 cells. On the other hand, cells from rats flown in space showed no significant differences from vivarium and synchronous control rats in cytotoxicity for K-562 target cells. Binding of natural killer cells to K-562 target cells was unaffected by spaceflight. Antiorthostatic suspension resulted in higher levels of cytotoxicity from spleen cells for Cr-51-labeled K-562 cells. The results indicate differential effects of spaceflight on function of natural killer cells. This shows that spaceflight has selective effects on the immune response.

42 CFR 493.1265 - Standard: Virology.

Code of Federal Regulations, 2010 CFR

2010-10-01

... Systems § 493.1265 Standard: Virology. (a) When using cell culture to isolate or identify viruses, the laboratory must simultaneously incubate a cell substrate control or uninoculated cells as a negative control...

Microspectrofluorometry for metabolic control analysis and the study of organelle morphogenesis in cell differentiation and transformation

NASA Astrophysics Data System (ADS)

Hirschberg, Joseph G.; Kohen, Elli; Kohen, Cahide; Pinon, Raul

1994-02-01

Microspectrofluorometry has been used in conjunction with fluorescence micrography for metabolic control analysis in normal and genetically deficient human fibroblasts, as well as human melanoma cells. These studies point to the role of mitochondria as the `cell's policeman' with regard to metabolic control. Cytotoxic agents active on mitochondrial structure and function (i.e. anthralin, azelaic acid) produce an unleashing of extramitochondrial pathways characterized by large and out-of-control NAD(P)H transients elicited by microinjected substrates. An interesting aspect has been the demonstration of an active nuclear energy metabolism, by NAD(P)H fluorescence excited at 365 nm, which may help to link cell bioenergetics to gene expression in the eukaryotes by the use of DNA probes. The metabolic control analysis of cell bioenergetics has been extended to the pathways involved in the cell's handling of cytotoxic agents. Non invasive fluorescence equipment offers possibilities for diagnostics and therapeutics in dermatology. Structure and function studies can be carried out at considerably enhanced resolution and with on-line interpretation by introducing scanning nearfield optics microscopy (SNOM) and real-time interactive parameter experimentation control (RIPEC).

Surface micro- and nano-texturing of stainless steel by femtosecond laser for the control of cell migration.

PubMed

Martínez-Calderon, M; Manso-Silván, M; Rodríguez, A; Gómez-Aranzadi, M; García-Ruiz, J P; Olaizola, S M; Martín-Palma, R J

2016-11-02

The precise control over the interaction between cells and the surface of materials plays a crucial role in optimizing the integration of implanted biomaterials. In this regard, material surface with controlled topographic features at the micro- and nano-scales has been proved to affect the overall cell behavior and therefore the final osseointegration of implants. Within this context, femtosecond (fs) laser micro/nano machining technology was used in this work to modify the surface structure of stainless steel aiming at controlling cell adhesion and migration. The experimental results show that cells tend to attach and preferentially align to the laser-induced nanopatterns oriented in a specific direction. Accordingly, the laser-based fabrication method here described constitutes a simple, clean, and scalable technique which allows a precise control of the surface nano-patterning process and, subsequently, enables the control of cell adhesion, migration, and polarization. Moreover, since our surface-patterning approach does not involve any chemical treatments and is performed in a single step process, it could in principle be applied to most metallic materials.

Surface micro- and nano-texturing of stainless steel by femtosecond laser for the control of cell migration

PubMed Central

Martínez-Calderon, M.; Manso-Silván, M.; Rodríguez, A.; Gómez-Aranzadi, M.; García-Ruiz, J. P.; Olaizola, S. M.; Martín-Palma, R. J.

2016-01-01

The precise control over the interaction between cells and the surface of materials plays a crucial role in optimizing the integration of implanted biomaterials. In this regard, material surface with controlled topographic features at the micro- and nano-scales has been proved to affect the overall cell behavior and therefore the final osseointegration of implants. Within this context, femtosecond (fs) laser micro/nano machining technology was used in this work to modify the surface structure of stainless steel aiming at controlling cell adhesion and migration. The experimental results show that cells tend to attach and preferentially align to the laser-induced nanopatterns oriented in a specific direction. Accordingly, the laser-based fabrication method here described constitutes a simple, clean, and scalable technique which allows a precise control of the surface nano-patterning process and, subsequently, enables the control of cell adhesion, migration, and polarization. Moreover, since our surface-patterning approach does not involve any chemical treatments and is performed in a single step process, it could in principle be applied to most metallic materials. PMID:27805063

Rapid reconstitution of CMV-specific T-cells after stem-cell transplantation.

PubMed

Widmann, Thomas; Sester, Urban; Schmidt, Tina; Gärtner, Barbara C; Schubert, Jörg; Pfreundschuh, Michael; Sester, Martina

2018-04-13

As reconstitution of virus-specific T-cells is critical to control cytomegalovirus (CMV)-viremia following stem-cell transplantation (SCT), we characterized the dynamics in CMV-specific T-cell reconstitution after SCT. Cytomegalovirus-specific T-cells from 51 SCT-recipients were prospectively quantified and phenotypically characterised by intracellular cytokine-staining after specific stimulation and HLA class-I-specific pentamers using flow cytometry. Cytomegalovirus-specific CD4 T-cells reconstituted after a median of 2.3 (IQR, 2.0-3.0) weeks following autografting, and 4.0 (IQR, 3.0-5.6) weeks after allografting, with CMV-specific T-cells originating from donors and/or recipients. The time for reconstitution of CMV-specific CD4 and CD8 T-cells did not differ (P = .58). Factors delaying the time to initial reconstitution of CMV-specific CD4 T-cells included a negative recipient serostatus (P = .016) and CMV-viremia (P = .026). Percentages of CMV-specific CD4 T-cells significantly increased over time and reached a plateau after 90 days (P = .043). Relative CMV-specific CD4 T-cell levels remained higher in long-term transplant recipients compared with those in controls (P < .0001). However, due to persisting lymphopenia, absolute numbers of CMV-specific T-cells were similar as in controls. Cytomegalovirus-specific T-cells rapidly reconstitute after SCT and their percentages remain high in the long term. In the face of persistent lymphopenia, this results in similar absolute numbers of CMV-specific T-cells as in controls to ensure sufficient pathogen control. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Temperature and UV light affect the activity of marine cell-free enzymes

NASA Astrophysics Data System (ADS)

Thomson, Blair; Hepburn, Christopher David; Lamare, Miles; Baltar, Federico

2017-09-01

Microbial extracellular enzymatic activity (EEA) is the rate-limiting step in the degradation of organic matter in the oceans. These extracellular enzymes exist in two forms: cell-bound, which are attached to the microbial cell wall, and cell-free, which are completely free of the cell. Contrary to previous understanding, cell-free extracellular enzymes make up a substantial proportion of the total marine EEA. Little is known about these abundant cell-free enzymes, including what factors control their activity once they are away from their sites (cells). Experiments were run to assess how cell-free enzymes (excluding microbes) respond to ultraviolet radiation (UVR) and temperature manipulations, previously suggested as potential control factors for these enzymes. The experiments were done with New Zealand coastal waters and the enzymes studied were alkaline phosphatase (APase), β-glucosidase, (BGase), and leucine aminopeptidase (LAPase). Environmentally relevant UVR (i.e. in situ UVR levels measured at our site) reduced cell-free enzyme activities by up to 87 % when compared to controls, likely a consequence of photodegradation. This effect of UVR on cell-free enzymes differed depending on the UVR fraction. Ambient levels of UV radiation were shown to reduce the activity of cell-free enzymes for the first time. Elevated temperatures (15 °C) increased the activity of cell-free enzymes by up to 53 % when compared to controls (10 °C), likely by enhancing the catalytic activity of the enzymes. Our results suggest the importance of both UVR and temperature as control mechanisms for cell-free enzymes. Given the projected warming ocean environment and the variable UVR light regime, it is possible that there could be major changes in the cell-free EEA and in the enzymes contribution to organic matter remineralization in the future.

Neonatal isolation impairs neurogenesis in the dentate gyrus of the guinea pig.

PubMed

Rizzi, Simona; Bianchi, Patrizia; Guidi, Sandra; Ciani, Elisabetta; Bartesaghi, Renata

2007-01-01

In the current study we examined the effects of early isolation rearing on cell proliferation, survival and differentiation in the dentate gyrus of the guinea pig. Animals were assigned to either a standard (control) or an isolated environment a few days after birth (P5-P6), taking advantage of the precocious independence from maternal care of the guinea pig. On P14-P17 animals received one daily bromodeoxyuridine injection, to label dividing cells, and were sacrificed either on P18, to evaluate cell proliferation or on P45, to evaluate cell survival and differentiation. In P18 isolated animals we found a reduced cell proliferation (-35%) compared to controls and a lower expression of brain-derived neurotrophic factor (BDNF). Though in absolute terms P45 isolated animals had less surviving cells, they showed no differences in survival rate and phenotype percent distribution compared to controls. Looking at the location of the new neurons, we found that while in control animals 76% of them had migrated to the granule cell layer, in isolated animals only 55% of the new neurons had reached this layer. Examination of radial glia cells of P18 and P45 animals by vimentin immunohistochemistry showed that in isolated animals radial glia cells were reduced in density and had less and shorter processes. Granule cell count revealed that P45 isolated animals had less (-42%) granule cells than controls. Results show that isolation rearing reduces hippocampal cell proliferation, likely by reducing BDNF expression and hampers migration of the new neurons to the granule cell layer, likely by altering density/morphology of radial glia cells. The large reduction in granule cell number following isolation rearing emphasizes the role of environmental cues as relevant modulators of neurogenesis.

PP2ARts1 is a master regulator of pathways that control cell size

PubMed Central

Zapata, Jessica; Dephoure, Noah; MacDonough, Tracy; Yu, Yaxin; Parnell, Emily J.; Mooring, Meghan; Gygi, Steven P.; Stillman, David J.

2014-01-01

Cell size checkpoints ensure that passage through G1 and mitosis occurs only when sufficient growth has occurred. The mechanisms by which these checkpoints work are largely unknown. PP2A associated with the Rts1 regulatory subunit (PP2ARts1) is required for cell size control in budding yeast, but the relevant targets are unknown. In this paper, we used quantitative proteome-wide mass spectrometry to identify proteins controlled by PP2ARts1. This revealed that PP2ARts1 controls the two key checkpoint pathways thought to regulate the cell cycle in response to cell growth. To investigate the role of PP2ARts1 in these pathways, we focused on the Ace2 transcription factor, which is thought to delay cell cycle entry by repressing transcription of the G1 cyclin CLN3. Diverse experiments suggest that PP2ARts1 promotes cell cycle entry by inhibiting the repressor functions of Ace2. We hypothesize that control of Ace2 by PP2ARts1 plays a role in mechanisms that link G1 cyclin accumulation to cell growth. PMID:24493588

Automated microbeam observation environment for biological analysis—Custom portable environmental control applied to a vertical microbeam system

PubMed Central

England, Matthew J.; Bigelow, Alan W.; Merchant, Michael J.; Velliou, Eirini; Welch, David; Brenner, David J.; Kirkby, Karen J.

2018-01-01

Vertical Microbeams (VMB) are used to irradiate individual cells with low MeV energy ions. The irradiation of cells using VMBs requires cells to be removed from an incubator; this can cause physiological changes to cells because of the lower CO2 concentration, temperature and relative humidity outside of the incubator. Consequently, for experiments where cells require irradiation and observation for extended time periods, it is important to provide a controlled environment. The highly customised nature of the microscopes used on VMB systems means that there are no commercially available environmentally controlled microscope systems for VMB systems. The Automated Microbeam Observation Environment for Biological Analysis (AMOEBA) is a highly flexible modular environmental control system used to create incubator conditions on the end of a VMB. The AMOEBA takes advantage of the recent “maker” movement to create an open source control system that can be easily configured by the user to fit their control needs even beyond VMB applications. When applied to the task of controlling cell medium temperature, CO2 concentration and relative humidity on VMBs it creates a stable environment that allows cells to multiply on the end of a VMB over a period of 36 h, providing a low-cost (costing less than $2700 to build), customisable alternative to commercial time-lapse microscopy systems. AMOEBA adds the potential of VMBs to explore the long-term effects of radiation on single cells opening up new research areas for VMBs. PMID:29515291

Automated microbeam observation environment for biological analysis-Custom portable environmental control applied to a vertical microbeam system.

PubMed

England, Matthew J; Bigelow, Alan W; Merchant, Michael J; Velliou, Eirini; Welch, David; Brenner, David J; Kirkby, Karen J

2017-02-01

Vertical Microbeams (VMB) are used to irradiate individual cells with low MeV energy ions. The irradiation of cells using VMBs requires cells to be removed from an incubator; this can cause physiological changes to cells because of the lower CO 2 concentration, temperature and relative humidity outside of the incubator. Consequently, for experiments where cells require irradiation and observation for extended time periods, it is important to provide a controlled environment. The highly customised nature of the microscopes used on VMB systems means that there are no commercially available environmentally controlled microscope systems for VMB systems. The Automated Microbeam Observation Environment for Biological Analysis (AMOEBA) is a highly flexible modular environmental control system used to create incubator conditions on the end of a VMB. The AMOEBA takes advantage of the recent "maker" movement to create an open source control system that can be easily configured by the user to fit their control needs even beyond VMB applications. When applied to the task of controlling cell medium temperature, CO 2 concentration and relative humidity on VMBs it creates a stable environment that allows cells to multiply on the end of a VMB over a period of 36 h, providing a low-cost (costing less than $2700 to build), customisable alternative to commercial time-lapse microscopy systems. AMOEBA adds the potential of VMBs to explore the long-term effects of radiation on single cells opening up new research areas for VMBs.

Genetic Control of Wayward Pluripotent Stem Cells and Their Progeny after Transplantation

PubMed Central

Kiuru, Maija; Boyer, Julie L.; O’Connor, Timothy P.; Crystal, Ronald G.

2011-01-01

The proliferative capacity of pluripotent stem cells and their progeny brings a unique aspect to therapeutics, in that once a transplant is initiated the therapist no longer has control of the therapy. In the context of the recent FDA approval of a human ESC trial and report of a neuronal-stem-cell-derived tumor in a human trial, strategies need to be developed to control wayward pluripotent stem cells. Here, we focus on one approach: direct genetic modification of the cells prior to transplantation with genes that can prevent the adverse events and/or eliminate the transplanted cells and their progeny. PMID:19341619

Watch Out for the "Living Dead": Cell-Free Enzymes and Their Fate.

PubMed

Baltar, Federico

2017-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the "gatekeepers" of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell's fate. In contrast, cell-free enzymes belong to a kind of "living dead" realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go "beyond the living things," studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles.

Human Immunodeficiency Virus Type-1 Elite Controllers Maintain Low Co-Expression of Inhibitory Receptors on CD4+ T Cells.

PubMed

Noyan, Kajsa; Nguyen, Son; Betts, Michael R; Sönnerborg, Anders; Buggert, Marcus

2018-01-01

Human immunodeficiency virus type-1 (HIV-1) elite controllers (ELCs) represent a unique population that control viral replication in the absence of antiretroviral therapy (cART). It is well established that expression of multiple inhibitory receptors on CD8+ T cells is associated with HIV-1 disease progression. However, whether reduced co-expression of inhibitory receptors on CD4+ T cells is linked to natural viral control and slow HIV-1 disease progression remains undefined. Here, we report on the expression pattern of numerous measurable inhibitory receptors, associated with T cell exhaustion (programmed cell death-1, CTLA-4, and TIGIT), on different CD4+ T cell memory populations in ELCs and HIV-infected subjects with or without long-term cART. We found that the co-expression pattern of inhibitory receptors was significantly reduced in ELCs compared with HIV-1 cART-treated and viremic subjects, and similar to healthy controls. Markers associated with T cell exhaustion varied among different memory CD4+ T cell subsets and highest levels were found mainly on transitional memory T cells. CD4+ T cells co-expressing all inhibitory markers were positively correlated to T cell activation (CD38+ HLA-DR+) as well as the transcription factors Helios and FoxP3. Finally, clinical parameters such as CD4 count, HIV-1 viral load, and the CD4/CD8 ratio all showed significant associations with CD4+ T cell exhaustion. We demonstrate that ELCs are able to maintain lower levels of CD4+ T cell exhaustion despite years of ongoing viral replication compared with successfully cART-treated subjects. Our findings suggest that ELCs harbor a "healthy" state of inhibitory receptor expression on CD4+ T cells that might play part in maintenance of their control status.

Maternal retinoids control type 3 innate lymphoid cells and set the offspring immunity

NASA Astrophysics Data System (ADS)

van de Pavert, Serge A.; Ferreira, Manuela; Domingues, Rita G.; Ribeiro, Hélder; Molenaar, Rosalie; Moreira-Santos, Lara; Almeida, Francisca F.; Ibiza, Sales; Barbosa, Inês; Goverse, Gera; Labão-Almeida, Carlos; Godinho-Silva, Cristina; Konijn, Tanja; Schooneman, Dennis; O'Toole, Tom; Mizee, Mark R.; Habani, Yasmin; Haak, Esther; Santori, Fabio R.; Littman, Dan R.; Schulte-Merker, Stefan; Dzierzak, Elaine; Simas, J. Pedro; Mebius, Reina E.; Veiga-Fernandes, Henrique

2014-04-01

The impact of nutritional status during fetal life on the overall health of adults has been recognized; however, dietary effects on the developing immune system are largely unknown. Development of secondary lymphoid organs occurs during embryogenesis and is considered to be developmentally programmed. Secondary lymphoid organ formation depends on a subset of type 3 innate lymphoid cells (ILC3) named lymphoid tissue inducer (LTi) cells. Here we show that mouse fetal ILC3s are controlled by cell-autonomous retinoic acid (RA) signalling in utero, which pre-sets the immune fitness in adulthood. We found that embryonic lymphoid organs contain ILC progenitors that differentiate locally into mature LTi cells. Local LTi cell differentiation was controlled by maternal retinoid intake and fetal RA signalling acting in a haematopoietic cell-autonomous manner. RA controlled LTi cell maturation upstream of the transcription factor RORγt. Accordingly, enforced expression of Rorgt restored maturation of LTi cells with impaired RA signalling, whereas RA receptors directly regulated the Rorgt locus. Finally, we established that maternal levels of dietary retinoids control the size of secondary lymphoid organs and the efficiency of immune responses in the adult offspring. Our results reveal a molecular link between maternal nutrients and the formation of immune structures required for resistance to infection in the offspring.

Maternal retinoids control type 3 innate lymphoid cells and set the offspring immunity.

PubMed

van de Pavert, Serge A; Ferreira, Manuela; Domingues, Rita G; Ribeiro, Hélder; Molenaar, Rosalie; Moreira-Santos, Lara; Almeida, Francisca F; Ibiza, Sales; Barbosa, Inês; Goverse, Gera; Labão-Almeida, Carlos; Godinho-Silva, Cristina; Konijn, Tanja; Schooneman, Dennis; O'Toole, Tom; Mizee, Mark R; Habani, Yasmin; Haak, Esther; Santori, Fabio R; Littman, Dan R; Schulte-Merker, Stefan; Dzierzak, Elaine; Simas, J Pedro; Mebius, Reina E; Veiga-Fernandes, Henrique

2014-04-03

The impact of nutritional status during fetal life on the overall health of adults has been recognized; however, dietary effects on the developing immune system are largely unknown. Development of secondary lymphoid organs occurs during embryogenesis and is considered to be developmentally programmed. Secondary lymphoid organ formation depends on a subset of type 3 innate lymphoid cells (ILC3) named lymphoid tissue inducer (LTi) cells. Here we show that mouse fetal ILC3s are controlled by cell-autonomous retinoic acid (RA) signalling in utero, which pre-sets the immune fitness in adulthood. We found that embryonic lymphoid organs contain ILC progenitors that differentiate locally into mature LTi cells. Local LTi cell differentiation was controlled by maternal retinoid intake and fetal RA signalling acting in a haematopoietic cell-autonomous manner. RA controlled LTi cell maturation upstream of the transcription factor RORγt. Accordingly, enforced expression of Rorgt restored maturation of LTi cells with impaired RA signalling, whereas RA receptors directly regulated the Rorgt locus. Finally, we established that maternal levels of dietary retinoids control the size of secondary lymphoid organs and the efficiency of immune responses in the adult offspring. Our results reveal a molecular link between maternal nutrients and the formation of immune structures required for resistance to infection in the offspring.

The duration of mitosis and daughter cell size are modulated by nutrients in budding yeast

PubMed Central

2017-01-01

The size of nearly all cells is modulated by nutrients. Thus, cells growing in poor nutrients can be nearly half the size of cells in rich nutrients. In budding yeast, cell size is thought to be controlled almost entirely by a mechanism that delays cell cycle entry until sufficient growth has occurred in G1 phase. Here, we show that most growth of a new daughter cell occurs in mitosis. When the rate of growth is slowed by poor nutrients, the duration of mitosis is increased, which suggests that cells compensate for slow growth in mitosis by increasing the duration of growth. The amount of growth required to complete mitosis is reduced in poor nutrients, leading to a large reduction in cell size. Together, these observations suggest that mechanisms that control the extent of growth in mitosis play a major role in cell size control in budding yeast. PMID:28939614

Control of stem cell fate and function by engineering physical microenvironments

PubMed Central

Kshitiz; Park, Jinseok; Kim, Peter; Helen, Wilda; Engler, Adam J; Levchenko, Andre; Kim, Deok-Ho

2012-01-01

The phenotypic expression and function of stem cells are regulated by their integrated response to variable microenvironmental cues, including growth factors and cytokines, matrix-mediated signals, and cell-cell interactions. Recently, growing evidence suggests that matrix-mediated signals include mechanical stimuli such as strain, shear stress, substrate rigidity and topography, and these stimuli have a more profound impact on stem cell phenotypes than had previously been recognized, e.g. self-renewal and differentiation through the control of gene transcription and signaling pathways. Using a variety of cell culture models enabled by micro and nanoscale technologies, we are beginning to systematically and quantitatively investigate the integrated response of cells to combinations of relevant mechanobiological stimuli. This paper reviews recent advances in engineering physical stimuli for stem cell mechanobiology and discusses how micro- and nanoscale engineered platforms can be used to control stem cell niches environment and regulate stem cell fate and function. PMID:23077731

Regulatory iNKT cells lack PLZF expression and control Treg cell and macrophage homeostasis in adipose tissue

PubMed Central

Lynch, Lydia; Michelet, Xavier; Zhang, Sai; Brennan, Patrick J.; Moseman, Ashley; Lester, Chantel; Besra, Gurdyal; Vomhof-Dekrey, Emilie E.; Tighe, Mike; Koay, Hui-Fern; Godfrey, Dale I.; Leadbetter, Elizabeth A.; Sant’Angelo, Derek B.; von Andrian, Ulrich; Brenner, Michael B.

2015-01-01

iNKT cells are CD1d-restricted lipid-sensing innate T cells that express the transcription factor PLZF. iNKT cells accumulate in adipose tissue, where they are anti-inflammatory, but the factors that contribute to their anti-inflammatory nature, and their targets in adipose tissue are unknown. Here we report that adipose tissue iNKT cells have a unique transcriptional program and produce interleukin 2 (IL-2) and IL-10. Unlike other iNKT cells, they lack PLZF, but express the transcription factor E4BP4, which controls their IL-10 production. Adipose iNKT cells are a tissue resident population that induces an anti-inflammatory phenotype in macrophages and, through production of IL-2, controls the number, proliferation and suppressor function of adipose regulatory T (Treg) cells. Thus, adipose tissue iNKT cells are unique regulators of immune homeostasis in this tissue. PMID:25436972

T regulatory cells participate in the control of germinal centre reactions

PubMed Central

Alexander, Carla-Maria; Tygrett, Lorraine T; Boyden, Alexander W; Wolniak, Kristy L; Legge, Kevin L; Waldschmidt, Thomas J

2011-01-01

Germinal centre (GC) reactions are central features of T-cell-driven B-cell responses, and the site where antibody-producing cells and memory B cells are generated. Within GCs, a range of complex cellular and molecular events occur which are critical for the generation of high affinity antibodies. These processes require exquisite regulation not only to ensure the production of desired antibodies, but to minimize unwanted autoreactive or low affinity antibodies. To assess whether T regulatory (Treg) cells participate in the control of GC responses, immunized mice were treated with an anti-glucocorticoid-induced tumour necrosis factor receptor-related protein (GITR) monoclonal antibody (mAb) to disrupt Treg-cell activity. In anti-GITR-treated mice, the GC B-cell pool was significantly larger compared with control-treated animals, with switched GC B cells composing an abnormally high proportion of the response. Dysregulated GCs were also observed regardless of strain, T helper type 1 or 2 polarizing antigens, and were also seen after anti-CD25 mAb treatment. Within the spleens of immunized mice, CXCR5+ and CCR7− Treg cells were documented by flow cytometry and Foxp3+ cells were found within GCs using immunohistology. Final studies demonstrated administration of either anti-transforming growth factor-β or anti-interleukin-10 receptor blocking mAb to likewise result in dysregulated GCs, suggesting that generation of inducible Treg cells is important in controlling the GC response. Taken together, these findings indicate that Treg cells contribute to the overall size and quality of the humoral response by controlling homeostasis within GCs. PMID:21635248

An extensive program of periodic alternative splicing linked to cell cycle progression

PubMed Central

Dominguez, Daniel; Tsai, Yi-Hsuan; Weatheritt, Robert; Wang, Yang; Blencowe, Benjamin J; Wang, Zefeng

2016-01-01

Progression through the mitotic cell cycle requires periodic regulation of gene function at the levels of transcription, translation, protein-protein interactions, post-translational modification and degradation. However, the role of alternative splicing (AS) in the temporal control of cell cycle is not well understood. By sequencing the human transcriptome through two continuous cell cycles, we identify ~1300 genes with cell cycle-dependent AS changes. These genes are significantly enriched in functions linked to cell cycle control, yet they do not significantly overlap genes subject to periodic changes in steady-state transcript levels. Many of the periodically spliced genes are controlled by the SR protein kinase CLK1, whose level undergoes cell cycle-dependent fluctuations via an auto-inhibitory circuit. Disruption of CLK1 causes pleiotropic cell cycle defects and loss of proliferation, whereas CLK1 over-expression is associated with various cancers. These results thus reveal a large program of CLK1-regulated periodic AS intimately associated with cell cycle control. DOI: http://dx.doi.org/10.7554/eLife.10288.001 PMID:27015110

Bone regeneration with osteogenic matrix cell sheet and tricalcium phosphate: An experimental study in sheep.

PubMed

Kira, Tsutomu; Akahane, Manabu; Omokawa, Shohei; Shimizu, Takamasa; Kawate, Kenji; Onishi, Tadanobu; Tanaka, Yasuhito

2017-10-18

To determine the effects of a cell sheet created from sheep bone marrow and tricalcium phosphate (TCP) on osteogenesis. Bone marrow cells were harvested from a sheep and cultured in a minimal essential medium (MEM) containing ascorbic acid phosphate (AscP) and dexamethasone (Dex). After 2 wk, the formed osteogenic matrix cell sheet was lifted from the culture dish using a scraper. Additionally, harvested bone marrow cells were cultured in MEM only as a negative control group, and in MEM with AscP, Dex, and β-glycerophosphate as a positive control group. For in vitro evaluation, we measured the alkaline phosphatase (ALP) activity and osteocalcin (OC) content in the media of the cultured cells from each group. For in vivo analysis, a porous TCP ceramic was used as a scaffold. We prepared an experimental group comprising TCP scaffolds wrapped with the osteogenic matrix cell sheets and a control group consisting of the TCP scaffold only. The constructs were implanted subcutaneously into athymic rats and the cell donor sheep, and bone formation was confirmed by histology after 4 wk. In the in vitro part, the mean ALP activity was 0.39 ± 0.03 mg/well in the negative control group, 0.67 ± 0.04 mg/well in the sheet group, and 0.65 ± 0.07 mg/well in the positive control group. The mean OC levels were 1.46 ± 0.33 ng/well in the negative control group, 3.92 ± 0.16 ng/well in the sheet group, and 4.4 ± 0.47 ng/well in the positive control group, respectively. The ALP activity and OC levels were significantly higher in the cell sheet and positive control groups than in the negative control group ( P < 0.05). There was no significant difference in ALP activity or OC levels between the cell sheet group and the positive control group ( P > 0.05). TCP constructs wrapped with cell sheets prior to implantation showed bone formation, in contrast to TCP scaffolds alone, which exhibited poor bone formation when implanted, in the subcutaneous layer both in athymic rats and in the sheep. This technique for preparing highly osteoinductive TCP may promote regeneration in large bone defects.

9 CFR 113.300 - General requirements for live virus vaccines.

Code of Federal Regulations, 2010 CFR

2010-01-01

... control cells shall remain free of such fluorescence. (2) Serum neutralization test. The serum... fungi in accordance with the test provided in § 113.27. (2) Mycoplasma. Final container samples of... inoculated cells and uninoculated control cells. Cells shall be stained with fluorochrome conjugated specific...

Influence of zinc deficiency on cell-membrane fluidity in Jurkat, 3T3 and IMR-32 cells.

PubMed Central

Verstraeten, Sandra V; Zago, M Paola; MacKenzie, Gerardo G; Keen, Carl L; Oteiza, Patricia I

2004-01-01

We investigated whether zinc deficiency can affect plasma membrane rheology. Three cell lines, human leukaemia T-cells (Jurkat), rat fibroblasts (3T3) and human neuroblastoma cells (IMR-32), were cultured for 48 h in control medium, in zinc-deficient medium (1.5 microM zinc; 1.5 Zn), or in the zinc-deficient medium supplemented with 15 microM zinc (15 Zn). The number of viable cells was lower in the 1.5 Zn group than in the control and 15 Zn groups. The frequency of apoptosis was higher in the 1.5 Zn group than in the control and 15 Zn groups. Membrane fluidity was evaluated using the 6-(9-anthroyloxy)stearic acid and 16-(9-anthroyloxy)palmitic acid probes. Membrane fluidity was higher in 1.5 Zn cells than in the control cells; no differences were observed between control cells and 15 Zn cells. The effect of zinc deficiency on membrane fluidity at the water/lipid interface was associated with a higher phosphatidylserine externalization. The higher membrane fluidity in the hydrophobic region of the bilayer was correlated with a lower content of arachidonic acid. We suggest that the increased fluidity of the membrane secondary to zinc deficiency is in part due to a decrease in arachidonic acid content and the apoptosis-related changes in phosphatidylserine distribution. PMID:14629198

Osteoclasts control reactivation of dormant myeloma cells by remodelling the endosteal niche

PubMed Central

Lawson, Michelle A.; McDonald, Michelle M.; Kovacic, Natasa; Hua Khoo, Weng; Terry, Rachael L.; Down, Jenny; Kaplan, Warren; Paton-Hough, Julia; Fellows, Clair; Pettitt, Jessica A.; Neil Dear, T.; Van Valckenborgh, Els; Baldock, Paul A.; Rogers, Michael J.; Eaton, Colby L.; Vanderkerken, Karin; Pettit, Allison R.; Quinn, Julian M. W.; Zannettino, Andrew C. W.; Phan, Tri Giang; Croucher, Peter I.

2015-01-01

Multiple myeloma is largely incurable, despite development of therapies that target myeloma cell-intrinsic pathways. Disease relapse is thought to originate from dormant myeloma cells, localized in specialized niches, which resist therapy and repopulate the tumour. However, little is known about the niche, and how it exerts cell-extrinsic control over myeloma cell dormancy and reactivation. In this study, we track individual myeloma cells by intravital imaging as they colonize the endosteal niche, enter a dormant state and subsequently become activated to form colonies. We demonstrate that dormancy is a reversible state that is switched ‘on' by engagement with bone-lining cells or osteoblasts, and switched ‘off' by osteoclasts remodelling the endosteal niche. Dormant myeloma cells are resistant to chemotherapy that targets dividing cells. The demonstration that the endosteal niche is pivotal in controlling myeloma cell dormancy highlights the potential for targeting cell-extrinsic mechanisms to overcome cell-intrinsic drug resistance and prevent disease relapse. PMID:26632274

Hippo signaling controls cell cycle and restricts cell plasticity in planarians

PubMed Central

de Sousa, Nídia; Rodríguez-Esteban, Gustavo; Rojo-Laguna, Jose Ignacio; Saló, Emili

2018-01-01

The Hippo pathway plays a key role in regulating cell turnover in adult tissues, and abnormalities in this pathway are consistently associated with human cancers. Hippo was initially implicated in the control of cell proliferation and death, and its inhibition is linked to the expansion of stem cells and progenitors, leading to larger organ size and tumor formation. To understand the mechanism by which Hippo directs cell renewal and promotes stemness, we studied its function in planarians. These stem cell–based organisms are ideal models for the analysis of the complex cellular events underlying tissue renewal in the whole organism. hippo RNA interference (RNAi) in planarians decreased apoptotic cell death, induced cell cycle arrest, and could promote the dedifferentiation of postmitotic cells. hippo RNAi resulted in extensive undifferentiated areas and overgrowths, with no effect on body size or cell number. We propose an essential role for hippo in controlling cell cycle, restricting cell plasticity, and thereby preventing tumoral transformation. PMID:29357350

Surface engineering approaches to micropattern surfaces for cell-based assays.

PubMed

Falconnet, Didier; Csucs, Gabor; Grandin, H Michelle; Textor, Marcus

2006-06-01

The ability to produce patterns of single or multiple cells through precise surface engineering of cell culture substrates has promoted the development of cellular bioassays that provide entirely new insights into the factors that control cell adhesion to material surfaces, cell proliferation, differentiation and molecular signaling pathways. The ability to control shape and spreading of attached cells and cell-cell contacts through the form and dimension of the cell-adhesive patches with high precision is important. Commitment of stem cells to different specific lineages depends strongly on cell shape, implying that controlled microenvironments through engineered surfaces may not only be a valuable approach towards fundamental cell-biological studies, but also of great importance for the design of cell culture substrates for tissue engineering. Furthermore, cell patterning is an important tool for organizing cells on transducers for cell-based sensing and cell-based drug discovery concepts. From a material engineering standpoint, patterning approaches have greatly profited by combining microfabrication technologies, such as photolithography, with biochemical functionalization to present to the cells biological cues in spatially controlled regions where the background is rendered non-adhesive ("non-fouling") by suitable chemical modification. The focus of this review is on the surface engineering aspects of biologically motivated micropatterning of two-dimensional (flat) surfaces with the aim to provide an introductory overview and critical assessment of the many techniques described in the literature. In particular, the importance of non-fouling surface chemistries, the combination of hard and soft lithography with molecular assembly techniques as well as a number of less well known, but useful patterning approaches, including direct cell writing, are discussed.

Pectin methyl esterases and pectins in normal and hyperhydric shoots of carnation cultured in vitro.

PubMed

Saher, Shady; Piqueras, Abel; Hellin, Eladio; Olmos, Enrique

2005-02-01

Control and hyperhydric micropropagated plantlets from three carnation cultivars have been used to study their pectin composition and the activity of pectin methyl esterases (PMEs; EC 3.1.1.11). Pectins are a highly heterogeneous group of polymers that contribute to cell adhesion, cell wall architecture, and cell wall mechanical strength. Pectins control cell wall porosity and cell wall ionic status and are implicated in intercellular space development. The degree of esterification of pectins is controlled by the activity of cell wall PMEs; their different actions can affect the properties of the cell wall, which have been considered important with respect to controlling the development of hyperhydricity. The total pectins of hyperhydric leaves of the three varieties were significantly reduced in comparison with controls. The pectate fraction was significantly increased in hyperhydric leaves of all varieties while soluble pectins and protopectins were significantly lower. The PME activity of hyperhydric leaves was higher (4-10 times) compared to controls of the three varieties. Isoelectric focusing of PME isozymes revealed the presence of three isoforms; neutral PME activity was the major isozyme in control and hyperhydric leaves of the three varieties, whilst a decrease in the activity of the acidic isoforms was observed in hyperhydric leaves. The different PME activities could regulate some of the structural changes related to hyperhydricity in micropropagated carnation plants.

Presynaptic gain control by endogenous cotransmission of dopamine and GABA in the olfactory bulb.

PubMed

Vaaga, Christopher E; Yorgason, Jordan T; Williams, John T; Westbrook, Gary L

2017-03-01

In the olfactory bulb, lateral inhibition mediated by local juxtaglomerular interneurons has been proposed as a gain control mechanism, important for decorrelating odorant responses. Among juxtaglomerular interneurons, short axon cells are unique as dual-transmitter neurons that release dopamine and GABA. To examine their intraglomerular function, we expressed channelrhodopsin under control of the DAT-cre promoter and activated olfactory afferents within individual glomeruli. Optical stimulation of labeled cells triggered endogenous dopamine release as measured by cyclic voltammetry and GABA release as measured by whole cell GABA A receptor currents. Activation of short axon cells reduced the afferent presynaptic release probability via D 2 and GABA B receptor activation, resulting in reduced spiking in both mitral and external tufted cells. Our results suggest that short axon cells influence glomerular activity not only by direct inhibition of external tufted cells but also by inhibition of afferent inputs to external tufted and mitral cells. NEW & NOTEWORTHY Sensory systems, including the olfactory system, encode information across a large dynamic range, making synaptic mechanisms of gain control critical to proper function. Here we demonstrate that a dual-transmitter interneuron in the olfactory bulb controls the gain of intraglomerular afferent input via two distinct mechanisms, presynaptic inhibition as well as inhibition of a principal neuron subtype, and thereby potently controls the synaptic gain of afferent inputs. Copyright © 2017 the American Physiological Society.

The putative oncotarget CSN5 controls a transcription-uncorrelated p53-mediated autophagy implicated in cancer cell survival under curcumin treatment.

PubMed

Zhang, Qing-Yu; Jin, Rui; Zhang, Xian; Sheng, Ji-Po; Yu, Fang; Tan, Ren-Xiang; Pan, Ying; Huang, Jun-Jian; Kong, Ling-Dong

2016-10-25

Curcumin has shown promise as a safe and specific anticancer agent. The COP9 signalosome (CSN) component CSN5, a known specific target for curcumin, can control p53 stability by increasing its degradation through ubiquitin system. But the correlation of CSN5-controlled p53 to anticancer therapeutic effect of curcumin is currently unknown. Here we showed that CSN5-controlled p53 was transcriptional inactive and responsible for autophagy in human normal BJ cells and cancer HepG2 cells under curcumin treatment. Of note, CSN5-initiated cellular autophagy by curcumin treatment was abolished in p53-null HCT116p53-/- cancer cells, which could be rescued by reconstitution with wild-type p53 or transcription inactive p53 mutant p53R273H. Furthermore, CSN5-controlled p53 conferred a pro-survival autophagy in diverse cancer cells response to curcumin. Genetic p53 deletion, as well as autophagy pharmacological inhibition by chloroquine, significantly enhanced the therapeutic effect of curcumin on cancer cells in vitro and in vivo, but not normal cells. This study identifies a novel CSN5-controlled p53 in autophagy of human cells. The p53 expression state is a useful biomarker for predicting the anticancer therapeutic effect of curcumin. Therefore, the pharmacologic autophagy manipulation may benefit the ongoing anticancer clinical trials of curcumin.

BTG interacts with retinoblastoma to control cell fate in Dictyostelium.

PubMed

Conte, Daniele; MacWilliams, Harry K; Ceccarelli, Adriano

2010-03-12

In the genesis of many tissues, a phase of cell proliferation is followed by cell cycle exit and terminal differentiation. The latter two processes overlap: genes involved in the cessation of growth may also be important in triggering differentiation. Though conceptually distinct, they are often causally related and functional interactions between the cell cycle machinery and cell fate control networks are fundamental to coordinate growth and differentiation. A switch from proliferation to differentiation may also be important in the life cycle of single-celled organisms, and genes which arose as regulators of microbial differentiation may be conserved in higher organisms. Studies in microorganisms may thus contribute to understanding the molecular links between cell cycle machinery and the determination of cell fate choice networks. Here we show that in the amoebozoan D. discoideum, an ortholog of the metazoan antiproliferative gene btg controls cell fate, and that this function is dependent on the presence of a second tumor suppressor ortholog, the retinoblastoma-like gene product. Specifically, we find that btg-overexpressing cells preferentially adopt a stalk cell (and, more particularly, an Anterior-Like Cell) fate. No btg-dependent preference for ALC fate is observed in cells in which the retinoblastoma-like gene has been genetically inactivated. Dictyostelium btg is the only example of non-metazoan member of the BTG family characterized so far, suggesting that a genetic interaction between btg and Rb predated the divergence between dictyostelids and metazoa. While the requirement for retinoblastoma function for BTG antiproliferative activity in metazoans is known, an interaction of these genes in the control of cell fate has not been previously documented. Involvement of a single pathway in the control of mutually exclusive processes may have relevant implication in the evolution of multicellularity.

Tumorigenicity of MCF-7 human breast cancer cells lacking the p38α mitogen-activated protein kinase

PubMed Central

Mendoza, Rhone A; Moody, Emily E; Enriquez, Marlene I; Mejia, Sylvia M; Thordarson, Gudmundur

2011-01-01

We have generated cell lines with significantly reduced expression of the p38 mitogen-activated protein kinase (p38 MAPK), Min-p38 MAPK cells, and used these cells to investigate its role in tumorigenesis of breast cancer cells. MCF-7 cells were stably transfected with a plasmid producing small interfering RNA that inhibited the expression of p38 MAPK. Control cells were stably transfected with the same plasmid producing non-interfering RNA. The reduction in the p38 MAPK activity caused a significant increase in the expressions of the estrogen receptor-α (ERα) and the progesterone receptor, but eliminated the expression of the ERβ. Min-p38 MAPK cells showed an enhanced overall growth response to 17β-estradiol (E2), whereas growth hormone plus epidermal growth factor were largely ineffective growth stimulators in these cells compared to controls. Although the long-term net growth rate of the Min-p38 MAPK cells was increased in response to E2, their proliferation rate was not different from controls in short-term cultures. However, the Min-p38 MAPK cells did show a significant decreased rate of apoptosis after E2 treatment and a reduction in the basal phosphorylation of p53 tumor suppressor protein compared to controls. When the Min-p38 MAPK cells were xenografted into E2-treated athymic nude mice, their tumorigenicity was enhanced compared to control cells. Conclusions: increased tumorigenicity of Min-p38 MAPK cells was caused mainly by a decrease in apoptosis rate indicating that the lack of the p38 MAPK caused an imbalance to increase the ERα:ERβ ratio and a reduction in the activity of the p53 tumor suppressor protein. PMID:20974639

Tumorigenicity of MCF-7 human breast cancer cells lacking the p38α mitogen-activated protein kinase.

PubMed

Mendoza, Rhone A; Moody, Emily E; Enriquez, Marlene I; Mejia, Sylvia M; Thordarson, Gudmundur

2011-01-01

We have generated cell lines with significantly reduced expression of the p38 mitogen-activated protein kinase (p38 MAPK), Min-p38 MAPK cells, and used these cells to investigate p38 MAPK's role in tumorigenesis of breast cancer cells. MCF-7 cells were stably transfected with a plasmid producing small interfering RNA that inhibited the expression of p38 MAPK. Control cells were stably transfected with the same plasmid producing non-interfering RNA. The reduction in the p38 MAPK activity caused a significant increase in the expressions of estrogen receptor-α (ERα) and the progesterone receptor, but eliminated the expression of ERβ. Min-p38 MAPK cells showed an enhanced overall growth response to 17β-estradiol (E₂), whereas GH plus epidermal growth factor were largely ineffective growth stimulators in these cells compared to controls. Although the long-term net growth rate of the Min-p38 MAPK cells was increased in response to E₂, their proliferation rate was lower compared to controls in short-term cultures. However, the Min-p38 MAPK cells did show a significant decreased rate of apoptosis after E₂ treatment and a reduction in the basal phosphorylation of p53 tumor suppressor protein compared to controls. When the Min-p38 MAPK cells were xenografted into E₂-treated athymic nude mice, their tumorigenicity was enhanced compared to control cells. Increased tumorigenicity of Min-p38 MAPK cells was caused mainly by a decrease in the apoptosis rate indicating that the lack of the p38 MAPK caused an imbalance to increase the ERα:ERβ ratio and a reduction in the activity of the p53 tumor suppressor protein.

Charging system and method for multicell storage batteries

DOEpatents

Cox, Jay A.

1978-01-01

A battery-charging system includes a first charging circuit connected in series with a plurality of battery cells for controlled current charging. A second charging circuit applies a controlled voltage across each individual cell for equalization of the cells to the fully charged condition. This controlled voltage is determined at a level above the fully charged open-circuit voltage but at a sufficiently low level to prevent corrosion of cell components by electrochemical reaction. In this second circuit for cell equalization, a transformer primary receives closely regulated, square-wave voltage which is coupled to a plurality of equal secondary coil windings. Each secondary winding is connected in parallel to each cell of a series-connected pair of cells through half-wave rectifiers and a shared, intermediate conductor.

[Cytocompatibility of collagen membranes with bladder transitional cells of rabbit in vitro].

PubMed

Sun, Daodong; Song, Bo; Sun, Danning

2004-05-01

To evaluate the cytocompatibility of collagen membranes with transitional cells of rabbit in vitro and to discuss the possibility of the collagen membranes as urologic tissue engineering scaffolds. Primary cultured transitional cells isolated from New Zealand rabbits were implanted on collagen membranes at 1 x 10(5) cells/cm2. The changes of cell adhering were observed by inverted microscope and scanning electron microscope 2, 12 and 24 hours later. The experiment was divided into 4 groups: non-cell group (black control) culture medium group (negative control), extract medium from Polyvinyl chloride group(positive control) and extract medium from collagen membranes group(experimental group). The cells of generations 2 to 4 were implanted in 96-hole-plank at 1 x 10(4) cells every hole. And every group had 5 holes. Then absorption coefficient were detected at the wave length of 490 nm by MTT assay. Then the cytotoxicity and cytocompatibility were evaluated by comparison of the numbers of absorption coefficient. The bladder transitional cells began to adhere to the collagen membrane 2 hours after implanting, and the number of the adhered cells increased with time. The actual absorption coefficient of experimental groups was 0.590 +/- 0.024, 1.065 +/- 0.40 and 1.129 +/- 0.074 after 24, 72 and 120 hours. The actual absorption coefficient of negative control group was 0.639 +/- 0.068, 1.022 +/- 0.044 and 1.087 +/- 0.111. The actual absorption coefficient of positive control group was 0.302 +/- 0.029, 0.653 +/- 0.083 and 0.694 +/- 0.031. There was significant difference between the experimental group and positive control (P < 0.01), and no significant difference between the experimental group and negative control(P > 0.05). Collagen membrane has good cytocompatibility with transitional cells and no cytotoxicity. It can be used as scaffolds of urologic tissue engineering.

Lysosome-controlled efficient ROS overproduction against cancer cells with a high pH-responsive catalytic nanosystem

NASA Astrophysics Data System (ADS)

Fu, Jingke; Shao, Yiran; Wang, Liyao; Zhu, Yingchun

2015-04-01

Excess reactive oxygen species (ROS) have been proved to damage cancer cells efficiently. ROS overproduction is thus greatly desirable for cancer therapy. To date, ROS production is generally uncontrollable and outside cells, which always bring severe side-effects in the vasculature. Since most ROS share a very short half-life and primarily react close to their site of formation, it would be more efficient if excess ROS are controllably produced inside cancer cells. Herein, we report an efficient lysosome-controlled ROS overproduction via a pH-responsive catalytic nanosystem (FeOx-MSNs), which catalyze the decomposition of H2O2 to produce considerable ROS selectively inside the acidic lysosomes (pH 5.0) of cancer cells. After a further incorporation of ROS-sensitive TMB into the nanosystem (FeOx-MSNs-TMB), both a distinct cell labeling and an efficient death of breast carcinoma cells are obtained. This lysosome-controlled efficient ROS overproduction suggests promising applications in cancer treatments.Excess reactive oxygen species (ROS) have been proved to damage cancer cells efficiently. ROS overproduction is thus greatly desirable for cancer therapy. To date, ROS production is generally uncontrollable and outside cells, which always bring severe side-effects in the vasculature. Since most ROS share a very short half-life and primarily react close to their site of formation, it would be more efficient if excess ROS are controllably produced inside cancer cells. Herein, we report an efficient lysosome-controlled ROS overproduction via a pH-responsive catalytic nanosystem (FeOx-MSNs), which catalyze the decomposition of H2O2 to produce considerable ROS selectively inside the acidic lysosomes (pH 5.0) of cancer cells. After a further incorporation of ROS-sensitive TMB into the nanosystem (FeOx-MSNs-TMB), both a distinct cell labeling and an efficient death of breast carcinoma cells are obtained. This lysosome-controlled efficient ROS overproduction suggests promising applications in cancer treatments. Electronic supplementary information (ESI) available: Experimental section, supplementary figures and characterization of as-prepared compounds. See DOI: 10.1039/c5nr00706b

Transfusion of cell saver salvaged blood in neonates and infants undergoing open heart surgery significantly reduces RBC and coagulant product transfusions and donor exposures: results of a prospective, randomized, clinical trial.

PubMed

Cholette, Jill M; Powers, Karen S; Alfieris, George M; Angona, Ronald; Henrichs, Kelly F; Masel, Debra; Swartz, Michael F; Daugherty, L Eugene; Belmont, Kevin; Blumberg, Neil

2013-02-01

To evaluate whether transfusion of cell saver salvaged, stored at the bedside for up to 24 hrs, would decrease the number of postoperative allogeneic RBC transfusions and donor exposures, and possibly improve clinical outcomes. Prospective, randomized, controlled, clinical trial. Pediatric cardiac intensive care unit. Infants weighing less than 20 kg (n = 106) presenting for cardiac surgery with cardiopulmonary bypass. Subjects were randomized to a cell saver transfusion group where cell saver blood was available for transfusion up to 24 hrs after collection, or to a control group. Cell saver subjects received cell saver blood for volume replacement and/or RBC transfusions. Control subjects received crystalloid or albumin for volume replacement and RBCs for anemia. Blood product transfusions, donor exposures, and clinical outcomes were compared between groups. Children randomized to the cell saver group had significantly fewer RBC transfusions (cell saver: 0.19 ± 0.44 vs. control: 0.75 ± 1.2; p = 0.003) and coagulant product transfusions in the first 48 hrs post-op (cell saver: 0.09 ± 0.45 vs. control: 0.62 ± 1.4; p = 0.013), and significantly fewer donor exposures (cell saver: 0.60 ± 1.4 vs. control: 2.3 ± 4.8; p = 0.019). This difference persisted over the first week post-op, but did not reach statistical significance (cell saver: 0.64 ± 1.24 vs. control: 1.1 ± 1.4; p = 0.07). There were no significant clinical outcome differences. Cell saver blood can be safely stored at the bedside for immediate transfusion for 24 hrs after collection. Administration of cell saver blood significantly reduces the number of RBC and coagulant product transfusions and donor exposures in the immediate postoperative period. Reduction of blood product transfusions has the potential to reduce transfusion-associated complications and decrease postoperative morbidity. Larger studies are needed to determine whether this transfusion strategy will improve clinical outcomes.

Measurement of uterine natural killer cell percentage in the periimplantation endometrium from fertile women and women with recurrent reproductive failure: establishment of a reference range.

PubMed

Chen, Xiaoyan; Mariee, Najat; Jiang, Lingming; Liu, Yingyu; Wang, Chi Chiu; Li, Tin Chiu; Laird, Susan

2017-12-01

Uterine natural killer cells are the major leukocytes present in the periimplantation endometrium. Previous studies have found controversial differences in uterine natural killer cell percentage in women with recurrent reproductive failure compared with fertile controls. We sought to compare the uterine natural killer cell percentage in women with recurrent reproductive failure and fertile controls. This was a retrospective study carried out in university hospitals. A total of 215 women from 3 university centers participated in the study, including 97 women with recurrent miscarriage, 34 women with recurrent implantation failure, and 84 fertile controls. Endometrial biopsy samples were obtained precisely 7 days after luteinization hormone surge in a natural cycle. Endometrial sections were immunostained for CD56 and cell counting was performed by a standardized protocol. Results were expressed as percentage of positive uterine natural killer cell/total stromal cells. The median uterine natural killer cell percentage in Chinese ovulatory fertile controls in natural cycles was 2.5% (range 0.9-5.3%). Using 5th and 95th percentile to define the lower and upper limits of uterine natural killer cell percentage, the reference range was 1.2-4.5%. Overall, the groups with recurrent reproductive failure had significantly higher uterine natural killer cell percentage than the controls (recurrent miscarriage: median 3.2%, range 0.6-8.8%; recurrent implantation failure: median 3.1%, range 0.8-8.3%). However, there was a subset of both groups (recurrent miscarriage: 16/97; recurrent implantation failure: 6/34) that had lower uterine natural killer cell percentage compared to fertile controls. A reference range for uterine natural killer cell percentage in fertile women was established. Women with recurrent reproductive failure had uterine natural killer cell percentages both above and below the reference range. Copyright © 2017 Elsevier Inc. All rights reserved.

The Association of Peripheral Blood Regulatory T-Cell Concentrations With Epithelial Ovarian Cancer: A Brief Report.

PubMed

Cannioto, Rikki A; Sucheston-Campbell, Lara E; Hampras, Shalaka; Goode, Ellen L; Knutson, Keith; Ness, Roberta; Modugno, Francesmary; Wallace, Paul; Szender, J Brian; Mayor, Paul; Hong, Chi-Chen; Joseph, Janine M; Friel, Grace; Davis, Warren; Nesline, Mary; Eng, Kevin H; Edwards, Robert P; Kruszka, Bridget; Schmitt, Kristina; Odunsi, Kunle; Moysich, Kirsten B

2017-01-01

There is a mounting body of evidence demonstrating higher percentages of regulatory T (Treg) cells in the peripheral blood of patients with cancer in comparison to healthy controls, but there is a paucity of epidemiological literature characterizing circulating Treg cells among patients with epithelial ovarian cancer (EOC). To investigate the role of peripheral Treg cells in ovarian neoplasms, we conducted a case-control study to characterize circulating concentrations of Treg cells among patients with EOC, women with benign ovarian conditions, and healthy controls without a history of cancer. Participants were identified for inclusion due to their participation in the Data Bank and BioRepository program at Roswell Park Cancer Institute in Buffalo, NY. Patients included 71 women with a primary diagnosis of EOC and 195 women with a diagnosis of benign ovarian conditions. Controls included 101 age- and race-matched women without a history of cancer. Nonfasting, pretreatment peripheral blood levels of CD3+CD4+CD25+FOXP3+ Treg cells were measured using flow cytometric analyses and expressed as a percentage of total CD3+ cells and as a percentage of total CD3+CD4+ cells. Compared to healthy controls and women with benign ovarian conditions, patients with EOC had significantly higher frequency of Treg cells (P < 0.04). In multivariable logistic regression analyses using Treg frequency expressed as a percentage of CD+3 cells, we observed a significant positive association between Treg cell percentage and EOC risk, with each 1% increase associated with a 37% increased risk of EOC (odds ratio, 1.37; 95% confidence interval, 1.04-1.80). We observed a similar trend when Treg frequency was expressed as a percentage of CD3+CD+4 cells (odds ratio, 1.22; 95% confidence interval, 0.99-1.49). The current study provides support that peripheral Treg cell frequency is elevated in patients with EOC in comparison to women with benign ovarian conditions and healthy controls.

The association of peripheral blood regulatory T-cell concentrations with epithelial ovarian cancer: A brief report

PubMed Central

Hampras, Shalaka; Goode, Ellen L.; Knutson, Keith; Ness, Roberta; Modugno, Francesmary; Wallace, Paul; Szender, J. Brian; Mayor, Paul; Hong, Chi-Chen; Joseph, Janine M.; Friel, Grace; Davis, Warren; Nesline, Mary; Eng, Kevin H.; Edwards, Robert P.; Kruszka, Bridget; Schmitt, Kristina; Odunsi, Kunle; Moysich, Kirsten B.

2016-01-01

Objective There is a mounting body of evidence demonstrating higher percentages of regulatory T (Treg) cells in the peripheral blood of cancer patients in comparison to healthy controls, but there is a paucity of epidemiological literature characterizing circulating Treg cells among epithelial ovarian cancer (EOC) patients. To investigate the role of peripheral Treg cells in ovarian neoplasms, we conducted a case-control study to characterize circulating concentrations of Treg cells among EOC patients, women with benign ovarian conditions, and healthy controls without a history of cancer. Materials and Methods Participants were identified for inclusion due to their participation in the Data Bank and BioRepository program at Roswell Park Cancer Institute in Buffalo, NY. Patients included 71 women with a primary diagnosis of EOC and 195 women with a diagnosis of benign ovarian conditions. Controls included 101 age- and race-matched women without a history of cancer. Non-fasting, pre-treatment peripheral blood levels of CD3+CD4+CD25+FOXP3+ Treg cells were measured using flow cytometric analyses and expressed as a percentage of total CD3+ cells and as a percentage of total CD3+CD4+ cells. Results Compared to healthy controls and women with benign ovarian conditions, EOC patients had significantly higher frequency of Treg cells (p<0.04). In multivariable logistic regression analyses utilizing Treg frequency expressed as a percentage of CD+3 cells, we observed a significant positive association between Treg cell percentage and EOC risk, with each one percent increase associated with a 37% increased risk of EOC (OR=1.37, 95% CI: 1.04-1.80). We observed a similar trend when Treg frequency was expressed as a percentage of CD3+CD+4 cells (OR=1.22, 95% CI: 0.99-1.49). Conclusions The current study provides support that peripheral Treg cell frequency is elevated in EOC patients in comparison to women with benign ovarian conditions and healthy controls. PMID:27759594

STAT3 Knockdown Reduces Pancreatic Cancer Cell Invasiveness and Matrix Metalloproteinase-7 Expression in Nude Mice

PubMed Central

Huang, Ke jian; Wu, Wei dong; Jiang, Tao; Cao, Jun; Feng, Zhen zhong; Qiu, Zheng jun

2011-01-01

Aims Transducer and activator of transcription-3 (STAT3) plays an important role in tumor cell invasion and metastasis. The aim of the present study was to investigate the effects of STAT3 knockdown in nude mouse xenografts of pancreatic cancer cells and underlying gene expression. Methods A STAT3 shRNA lentiviral vector was constructed and infected into SW1990 cells. qRT-PCR and western immunoblot were performed to detect gene expression. Nude mouse xenograft assays were used to assess changes in phenotypes of these stable cells in vivo. HE staining was utilized to evaluate tumor cell invasion and immunohistochemistry was performed to analyze gene expression. Results STAT3 shRNA successfully silenced expression of STAT3 mRNA and protein in SW1990 cells compared to control cells. Growth rate of the STAT3-silenced tumor cells in nude mice was significantly reduced compared to in the control vector tumors and parental cells-generated tumors. Tumor invasion into the vessel and muscle were also suppressed in the STAT3-silenced tumors compared to controls. Collagen IV expression was complete and continuous surrounding the tumors of STAT3-silenced SW1990 cells, whereas collagen IV expression was incomplete and discontinuous surrounding the control tumors. Moreover, microvessel density was significantly lower in STAT3-silenced tumors than parental or control tumors of SW1990 cells. In addition, MMP-7 expression was reduced in STAT3-silenced tumors compared to parental SW1990 xenografts and controls. In contrast, expression of IL-1β and IgT7α was not altered. Conclusion These data clearly demonstrate that STAT3 plays an important role in regulation of tumor growth, invasion, and angiogenesis, which could be act by reducing MMP-7 expression in pancreatic cancer cells. PMID:21991388

In-Vivo Real-Time Control of Protein Expression from Endogenous and Synthetic Gene Networks

PubMed Central

Orabona, Emanuele; De Stefano, Luca; Ferry, Mike; Hasty, Jeff; di Bernardo, Mario; di Bernardo, Diego

2014-01-01

We describe an innovative experimental and computational approach to control the expression of a protein in a population of yeast cells. We designed a simple control algorithm to automatically regulate the administration of inducer molecules to the cells by comparing the actual protein expression level in the cell population with the desired expression level. We then built an automated platform based on a microfluidic device, a time-lapse microscopy apparatus, and a set of motorized syringes, all controlled by a computer. We tested the platform to force yeast cells to express a desired fixed, or time-varying, amount of a reporter protein over thousands of minutes. The computer automatically switched the type of sugar administered to the cells, its concentration and its duration, according to the control algorithm. Our approach can be used to control expression of any protein, fused to a fluorescent reporter, provided that an external molecule known to (indirectly) affect its promoter activity is available. PMID:24831205

Protein and chemotherapy profiling of extracellular vesicles harvested from therapeutic induced senescent triple negative breast cancer cells

PubMed Central

Kavanagh, E L; Lindsay, S; Halasz, M; Gubbins, L C; Weiner-Gorzel, K; Guang, M H Z; McGoldrick, A; Collins, E; Henry, M; Blanco-Fernández, A; Gorman, P O'; Fitzpatrick, P; Higgins, M J; Dowling, P; McCann, A

2017-01-01

Triple negative breast cancer (TNBC) is an aggressive subtype with relatively poor clinical outcomes and limited treatment options. Chemotherapy, while killing cancer cells, can result in the generation of highly chemoresistant therapeutic induced senescent (TIS) cells that potentially form stem cell niches resulting in metastases. Intriguingly, senescent cells release significantly more extracellular vesicles (EVs) than non-senescent cells. Our aim was to profile EVs harvested from TIS TNBC cells compared with control cells to identify a potential mechanism by which TIS TNBC cells maintain survival in the face of chemotherapy. TIS was induced and confirmed in Cal51 TNBC cells using the chemotherapeutic paclitaxel (PTX) (Taxol). Mass spectrometry (MS) analysis of EVs harvested from TIS compared with control Cal51 cells was performed using Ingenuity Pathway Analysis and InnateDB programs. We demonstrate that TIS Cal51 cells treated with 75 nM PTX for 7 days became senescent (senescence-associated β-galactosidase (SA-β-Gal) positive, Ki67-negative, increased p21 and p16, G2/M cell cycle arrest) and released significantly more EVs (P=0.0002) and exosomes (P=0.0007) than non-senescent control cells. Moreover, TIS cells displayed an increased expression of the multidrug resistance protein 1/p-glycoprotein. MS analysis demonstrated that EVs derived from senescent Cal51 cells contained 142 proteins with a significant increased fold change compared with control EVs. Key proteins included ATPases, annexins, tubulins, integrins, Rabs and insoluble senescence-associated secretory phenotype (SASP) factors. A fluorescent analogue of PTX (Flutax-2) allowed appreciation of the removal of chemotherapy in EVs from senescent cells. Treatment of TIS cells with the exosome biogenesis inhibitor GW4869 resulted in reduced SA-β-Gal staining (P=0.04). In summary, this study demonstrates that TIS cells release significantly more EVs compared with control cells, containing chemotherapy and key proteins involved in cell proliferation, ATP depletion, apoptosis and the SASP. These findings may partially explain why cancer senescent cells remain viable despite chemotherapeutic challenge. PMID:28991260

A new topology of fuel cell hybrid power source for efficient operation and high reliability

NASA Astrophysics Data System (ADS)

Bizon, Nicu

2011-03-01

This paper analyzes a new fuel cell Hybrid Power Source (HPS) topology having the feature to mitigate the current ripple of the fuel cell inverter system. In the operation of the inverter system that is grid connected or supplies AC motors in vehicle application, the current ripple normally appears at the DC port of the fuel cell HPS. Consequently, if mitigation measures are not applied, this ripple is back propagated to the fuel cell stack. Other features of the proposed fuel cell HPS are the Maximum Power Point (MPP) tracking, high reliability in operation under sharp power pulses and improved energy efficiency in high power applications. This topology uses an inverter system directly powered from the appropriate fuel cell stack and a controlled buck current source as low power source used for ripple mitigation. The low frequency ripple mitigation is based on active control. The anti-ripple current is injected in HPS output node and this has the LF power spectrum almost the same with the inverter ripple. Consequently, the fuel cell current ripple is mitigated by the designed active control. The ripple mitigation performances are evaluated by indicators that are defined to measure the mitigation ratio of the low frequency harmonics. In this paper it is shown that good performances are obtained by using the hysteretic current control, but better if a dedicated nonlinear controller is used. Two ways to design the nonlinear control law are proposed. First is based on simulation trials that help to draw the characteristic of ripple mitigation ratio vs. fuel cell current ripple. The second is based on Fuzzy Logic Controller (FLC). The ripple factor is up to 1% in both cases.

Reducing cell wall feruloylation by expression of a fungal ferulic acid esterase in Festuca arundinacea modifies plant growth, leaf morphology and the turnover of cell wall arabinoxylans

PubMed Central

Iyer, Prashanti R.; Buanafina, M. Fernanda; Shearer, Erica A.

2017-01-01

A feature of cell wall arabinoxylan in grasses is the presence of ferulic acid which upon oxidative coupling by the action of peroxidases forms diferuloyl bridges between formerly separated arabinoxylans. Ferulate cross-linking is suspected of playing various roles in different plant processes. Here we investigate the role of cell wall feruloyaltion in two major processes, that of leaf growth and the turnover of cell wall arabinoxylans on leaf senescence in tall fescue using plants in which the level of cell wall ferulates has been reduced by targeted expression of the Aspergillus niger ferulic acid esterase A (FAEA) to the apoplast or Golgi. Analysis of FAE expressing plants showed that all the lines had shorter and narrower leaves compared to control, which may be a consequence of the overall growth rate being lower and occurring earlier in FAE expressing leaves than in controls. Furthermore, the final length of epidermal cells was shorter than controls, indicating that their expansion was curtailed earlier than in control leaves. This may be due to the observations that the deposition of both ether and ester linked monomeric hydroxycinnamic acids and ferulate dimerization stopped earlier in FAE expressing leaves but at a lower level than controls, and hydroxycinnamic acid deposition started to slow down when peroxidase levels increased. It would appear therefore that one of the possible mechanisms for controlling overall leaf morphology such as leaf length and width in grasses, where leaf morphology is highly variable between species, may be the timing of hydroxycinnamic acid deposition in the expanding cell walls as they emerge from cell division into the elongation zone, controlled partially by the onset of peroxidase activity in this region. PMID:28934356

The metabolites in peripheral blood mononuclear cells showed greater differences between patients with impaired fasting glucose or type 2 diabetes and healthy controls than those in plasma.

PubMed

Kim, Minjoo; Kim, Minkyung; Han, Ji Yun; Lee, Sang-Hyun; Jee, Sun Ha; Lee, Jong Ho

2017-03-01

To determine differences between peripheral blood mononuclear cells and the plasma metabolites in patients with impaired fasting glucose or type 2 diabetes and healthy controls. In all, 65 nononobese patients (aged 30-70 years) with impaired fasting glucose or type 2 diabetes and 65 nonobese sex-matched healthy controls were included, and fasting peripheral blood mononuclear cell and plasma metabolomes were profiled. The diabetic or impaired fasting glucose patients showed higher circulating and peripheral blood mononuclear cell lipoprotein phospholipase A 2 activities, high-sensitivity C-reactive protein and tumour necrosis factor-α than controls. Compared with controls, impaired fasting glucose or diabetic subjects showed increases in 11 peripheral blood mononuclear cell metabolites: six amino acids (valine, leucine, methionine, phenylalanine, tyrosine and tryptophan), l-pyroglutamic acid, two fatty acid amides containing palmitic amide and oleamide and two lysophosphatidylcholines. In impaired fasting glucose or diabetic patients, peripheral blood mononuclear cell lipoprotein phospholipase A 2 positively associated with peripheral blood mononuclear cell lysophosphatidylcholines and circulating inflammatory markers, including tumour necrosis factor-α, high-sensitivity C-reactive protein and lipoprotein phospholipase A 2 activities. In plasma metabolites between patients and healthy controls, we observed significant increases in only three amino acids (proline, valine and leucine) and decreases in only five lysophosphatidylcholines. This study demonstrates significant differences in the peripheral blood mononuclear cell metabolome in patients with impaired fasting glucose or diabetes compared with healthy controls. These differences were greater than those observed in the plasma metabolome. These data suggest peripheral blood mononuclear cells as a useful tool to better understand the inflammatory pathophysiology of diabetes.

Systems Modeling of Molecular Mechanisms Controlling Cytokine-driven CD4+ T Cell Differentiation and Phenotype Plasticity

PubMed Central

Carbo, Adria; Hontecillas, Raquel; Kronsteiner, Barbara; Viladomiu, Monica; Pedragosa, Mireia; Lu, Pinyi; Philipson, Casandra W.; Hoops, Stefan; Marathe, Madhav; Eubank, Stephen; Bisset, Keith; Wendelsdorf, Katherine; Jarrah, Abdul; Mei, Yongguo; Bassaganya-Riera, Josep

2013-01-01

Differentiation of CD4+ T cells into effector or regulatory phenotypes is tightly controlled by the cytokine milieu, complex intracellular signaling networks and numerous transcriptional regulators. We combined experimental approaches and computational modeling to investigate the mechanisms controlling differentiation and plasticity of CD4+ T cells in the gut of mice. Our computational model encompasses the major intracellular pathways involved in CD4+ T cell differentiation into T helper 1 (Th1), Th2, Th17 and induced regulatory T cells (iTreg). Our modeling efforts predicted a critical role for peroxisome proliferator-activated receptor gamma (PPARγ) in modulating plasticity between Th17 and iTreg cells. PPARγ regulates differentiation, activation and cytokine production, thereby controlling the induction of effector and regulatory responses, and is a promising therapeutic target for dysregulated immune responses and inflammation. Our modeling efforts predict that following PPARγ activation, Th17 cells undergo phenotype switch and become iTreg cells. This prediction was validated by results of adoptive transfer studies showing an increase of colonic iTreg and a decrease of Th17 cells in the gut mucosa of mice with colitis following pharmacological activation of PPARγ. Deletion of PPARγ in CD4+ T cells impaired mucosal iTreg and enhanced colitogenic Th17 responses in mice with CD4+ T cell-induced colitis. Thus, for the first time we provide novel molecular evidence in vivo demonstrating that PPARγ in addition to regulating CD4+ T cell differentiation also plays a major role controlling Th17 and iTreg plasticity in the gut mucosa. PMID:23592971

Application of cell-based assays for toxicity characterization of complex wastewater matrices: Possible applications in wastewater recycle and reuse.

PubMed

Shrivastava, Preeti; Naoghare, Pravin K; Gandhi, Deepa; Devi, S Saravana; Krishnamurthi, Kannan; Bafana, Amit; Kashyap, Sanjay M; Chakrabarti, Tapan

2017-08-01

Exposure to pre-concentrated inlet or outlet STP wastewater extracts at different concentrations (0.001% to 1%) induced dose-dependent toxicity in MCF-7 cells, whereas drinking water extracts did not induce cytotoxicity in cells treated. GC-MS analysis revealed the occurrence of xenobiotic compounds (Benzene, Phthalate, etc.) in inlet/outlet wastewater extracts. Cells exposed to inlet/outlet extract showed elevated levels of reactive oxygen species (ROS: inlet: 186.58%, p<0.05, outlet, 147.8%, p<0.01) and loss of mitochondrial membrane potential (Δψm: inlet, 74.91%, p<0.01; outlet, 86.70%, p<0.05) compared to the control. These concentrations induced DNA damage (Tail length: inlet: 34.4%, p<0.05, outlet, 26.7%, p<0.05) in treated cells compared to the control (Tail length: 7.5%). Cell cycle analysis displayed drastic reduction in the G1 phase in treated cells (inlet, G1:45.0%; outlet, G1:58.3%) compared to the control (G1:67.3%). Treated cells showed 45.18% and 28.0% apoptosis compared to the control (1.2%). Drinking water extracts did not show any significant alterations with respect to ROS, Δψm, DNA damage, cell cycle and apoptosis compared to the control. Genes involved in cell cycle and apoptosis were found to be differentially expressed in cells exposed to inlet/outlet extracts. Herein, we propose cell-based toxicity assays to evaluate the efficacies of wastewater treatment and recycling processes. Copyright © 2017 Elsevier Inc. All rights reserved.

Cell viability test after laser guidance

NASA Astrophysics Data System (ADS)

Rosenbalm, Tabitha N.; Owens, Sarah; Bakken, Daniel; Gao, Bruce Z.

2006-02-01

To precisely control the position of multiple types of cells in a coculture for the study of cell-cell interactions, we have developed a laser micropatterning technique. The technique employs the optical forces generated by a weakly focused laser beam. In the beam's focal region, the optical force draws microparticles, such as cells, into the center of the beam, propels them along the beam axis, and guides them onto a target surface. Specific patterns are created through computercontrolled micromanipulation of the substrate relative to the laser beam. Preliminary data have demonstrated cell viability after laser guidance. This project was designed to systematically vary the controllable laser parameters, namely, intensity and exposure time of the laser on single cells, and thus determine the laser parameters that allow negligible cell damage with functional cellular position control. To accomplish this goal, embryonic day 7 (E7) chick forebrain neurons were cultured in 35 mm petri dishes. Control and test cells were selected one hour after cell placement to allow cell attachment. Test cells were subjected to the laser at the focal region. The experimental parameters were chosen as: wavelength - 800 nm, intensities - 100 mW, 200 mW, and 300 mW, and exposure times - 10 s and 60 s. Results were analyzed based on neurite outgrowth and the Live/Dead assay (Viability/Cytoxicity kit from Molecular Probes). No statistical difference (p >> 0.1, student t-test) in viability or function was found between the control neurons and those exposed to the laser. This confirms that laser guidance seems to be a promising method for cellular manipulation.

Establishing a Quality Control System for Stem Cell-Based Medicinal Products in China

PubMed Central

2015-01-01

Stem cell-based medicinal products (SCMPs) are emerging as novel therapeutic products. The success of its development depends on the existence of an effective quality control system, which is constituted by quality control technologies, standards, reference materials, guidelines, and the associated management system in accordance with regulatory requirements along product lifespan. However, a worldwide, effective quality control system specific for SCMPs is still far from established partially due to the limited understanding of stem cell sciences and lack of quality control technologies for accurately assessing the safety and biological effectiveness of SCMPs before clinical use. Even though, based on the existing regulations and current stem cell sciences and technologies, initial actions toward the goal of establishing such a system have been taken as exemplified by recent development of new “interim guidelines” for governing quality control along development of SCMPs and new development of the associated quality control technologies in China. In this review, we first briefly introduced the major institutions involved in the regulation of cell substrates and therapeutic cell products in China and the existing regulatory documents and technical guidelines used as critical references for developing the new interim guidelines. With focus only on nonhematopoietic stem cells, we then discussed the principal quality attributes of SCMPs as well as our thinking of proper testing approaches to be established with relevant evaluation technologies to ensure all quality requirements of SCMPs along different manufacturing processes and development stages. At the end, some regulatory and technical challenges were also discussed with the conclusion that combined efforts should be taken to promote stem cell regulatory sciences to establish the effective quality control system for SCMPs. PMID:25471126

Hydrogels with Spatially and Temporally Controlled Properties to Control Cellular Interactions

NASA Astrophysics Data System (ADS)

Burdick, Jason

2011-03-01

Stem cells (e.g., mesenchymal stem cells, MSCs) respond to many cues from their microenvironment, which may include chemical signals, mechanics, and topography. Importantly, these cues may be incorporated into scaffolding to control stem cell differentiation and optimize their ability to produce tissues in regenerative medicine. Despite the significant amount of work in this area, the materials have been primarily static and uniform. To this end, we have developed a sequential crosslinking process that relies on our ability to crosslinked functional biopolymers (e.g., methacrylated hyaluronic acid, HA) in two steps, namely a Michael-type addition reaction to partially consume reactive groups and then a light-initiated free-radical polymerization to further crosslink the material. With light exposure during the second step comes control over the material in space (via masks and lasers) and time (via intermittent light exposure). We are applying this technique for numerous applications. For example, when the HA hydrogels are crosslinked with MMP degradable peptides with thiol termini during the first step, a material that can be degraded by cells is obtained. However, cell-mediated degradation is obstructed with the introduction of kinetic chains during the second step, leading to spatially controlled cell degradability. Due to the influence of cellular spreading on MSC differentiation, we have controlled cell fates by controlling their spread ability, for instance towards osteoblasts in spread areas and adipocytes when cell remained rounded. We are also using the process of stiffening with time to investigate mechanically induced differentiation, particularly in materials with evolving mechanics. Overall, these advanced HA hydrogels provide us the opportunity to investigate diverse and controlled material properties on MSC interactions.

[Effect of G-CSF in vitro Stimulation on Distribution of Peripheral Lymphocyte Subsets in the Healthy Persons].

PubMed

Zhao, Sha-Sha; Fang, Shu; Zhu, Cheng-Ying; Wang, Li-Li; Gao, Chun-Ji

2018-02-01

To investigate the effect of granulocyte-colony stimulating factor (G-CSF) in vitro stimulation on the distribution of lymphocyte subset in healthy human. Peripheral blood mononuclear cells (PBMNCs) were collected from 8 healthy volunteers by density gradient centrifugation on Ficoll-Paque TM . In vitro 200 ng/ml G-CSF or 200 ng/ml G-CSF plus 10 µg/ml ConA directly act on PBMNCs, then the colleted cells were cultivated for 3 days. Lymphocyte subsets were stained with the corresponding fluoresce labeled antibodies and detected by flow cytometry. The levels of T cells in G-CSF group and G-CSF+ConA group were both higher than that in the control group (P<0.001, P<0.05). However, there were not significantly different in B cells and NK cells levels among the 3 groups. Furthermore, analysis of the effect of G-CSF on T cell subsets indicated that the levels of CD4 + T cells and CD8 + T cells in G-CSF group were both significantly higher than those in control group (P<0.01, P<0.05), Treg cells was not different between G-CSF and control group. Compared with the control group, the level of CD4 + T cells, CD8 + T cells and Treg cells in G-CSF+ConA group significantly increased (P<0.05, P<0.01, P<0.01). Analysis of G-CSF receptor (G-CSFR) expression showed that G-CSFR expression on T cells in G-CSF+ConA group dramatically increased, as compared with control group (P<0.01). The levels of CD4 + T cells and CD8 + T cells in healthy human peripheral blood can be increased by G-CSF stimulation. ConA can enhance the level of T cells and induce G-CSFR expression on T cells.

BTLA associates with increased Foxp3 expression in CD4(+) T cells in dextran sulfate sodium-induced colitis.

PubMed

Zhang, Han-Xian; Zhu, Bin; Fu, Xiao-Xia; Zeng, Jin-Cheng; Zhang, Jun-Ai; Wang, Wan-Dang; Kong, Bin; Xiang, Wen-Yu; Zhong, Jixin; Wang, Cong-Yi; Zheng, Xue-Bao; Xu, Jun-Fa

2015-01-01

Ulcerative colitis (UC) is an inflammatory bowel disease, and its pathogenesis involves a variety of genetic, environmental, and immunological factors such as T helper cells and their secreted cytokines. B and T lymphocyte attenuator (BTLA) is an immunoregulatory receptor that has a strong suppressive effect on T-cell function. However the role of BTLA in UC remains poorly understood. Here we demonstrated that the frequency of BTLA-expressing CD3(+) T cells, especially CD4(+) T cells, increased in blood and mucosa in mice with DSS-induced colitis. The frequency of Foxp3-expressing cells in BTLA+ CD4(+) T cell from lamina propria mononuclear cells (LPMCs) was much higher in DSS-treated mice than that in controls. Similarly, the proportion of IL-17+ cells in BTLA+ CD4(+) T cells from LPMCs in DSS-treated mice is much higher than that in controls, while no perceptible difference for the proportion of IFN-γ+ cells in BTLA+ CD4(+) T cells was noted between DSS-treated mice and controls. Treatment of mesalazine, an anti-ulcerative colitis drug, down-regulated Foxp3 and IL-17 expression in BTLA positive T cells along with attenuated severity for colitis. Our findings indicate that BTLA may be involved in the control of inflammatory responses through increasing Foxp3 expression, rather than attenuating IL-17 production, in DSS-induced colitis.

Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung

PubMed Central

Lange, Alexander W.; Sridharan, Anusha; Xu, Yan; Stripp, Barry R.; Perl, Anne-Karina; Whitsett, Jeffrey A.

2015-01-01

The Hippo/Yap pathway is a well-conserved signaling cascade that regulates cell proliferation and differentiation to control organ size and stem/progenitor cell behavior. Following airway injury, Yap was dynamically regulated in regenerating airway epithelial cells. To determine the role of Hippo signaling in the lung, the mammalian Hippo kinases, Mst1 and Mst2, were deleted in epithelial cells of the embryonic and mature mouse lung. Mst1/2 deletion in the fetal lung enhanced proliferation and inhibited sacculation and epithelial cell differentiation. The transcriptional inhibition of cell proliferation and activation of differentiation during normal perinatal lung maturation were inversely regulated following embryonic Mst1/2 deletion. Ablation of Mst1/2 from bronchiolar epithelial cells in the adult lung caused airway hyperplasia and altered differentiation. Inhibitory Yap phosphorylation was decreased and Yap nuclear localization and transcriptional targets were increased after Mst1/2 deletion, consistent with canonical Hippo/Yap signaling. YAP potentiated cell proliferation and inhibited differentiation of human bronchial epithelial cells in vitro. Loss of Mst1/2 and expression of YAP regulated transcriptional targets controlling cell proliferation and differentiation, including Ajuba LIM protein. Ajuba was required for the effects of YAP on cell proliferation in vitro. Hippo/Yap signaling regulates Ajuba and controls proliferation and differentiation of lung epithelial progenitor cells. PMID:25480985

CD28 T-cell costimulatory molecule expression in pemphigus vulgaris.

PubMed

Alecu, M; Ursaciuc, C; Surcel, M; Coman, G; Ciotaru, D; Dobre, M

2009-03-01

CD28 superfamily of immune costimulatory molecules could play an important role in autotolerance control. CD28 costimulation seems to be necessary for regulatory T cell (Treg) activation and successive suppressive activities involved in autoimmunity protection. This study investigates CD28 expression, especially inducible costimulator fraction, on T lymphocytes in pemphigus vulgaris (PV) patients. CD28 expression on T lymphocytes was assessed in 16 PV patients during acute attack. All patients and 10 healthy control subjects were tested for lymphocyte populations, T-cell subpopulations (T-CD4+, T-CD8+), Treg and CD28 expression on T-cell subpopulations. T, B and natural killer cells average values in PV patients were close to the control group values. Compared with control group, PV values showed lower Treg (2.2% compared with 4.7%), slightly decreased CD4+ CD28+ T cells (91% compared with 95%), higher CD4+ CD28- T cells (9% compared with 5%), decreased CD8+ CD28+ T cells (57% and 73%, respectively) and significantly enhanced CD8+ CD28- T cells (43% compared with 27%). These data suggest that Treg-mediated suppressor T-cell effects could be diminished in PV, together with an abnormal or ineffective subsequent helper T-cell suppression. CD28 high expression on helper T cells and low expression on suppressor T cells are arguments for a potential CD28 role in PV autoimmune response mechanism.

Regulation of Cell Diameter, For3p Localization, and Cell Symmetry by Fission Yeast Rho-GAP Rga4p

PubMed Central

Das, Maitreyi; Wiley, David J.; Medina, Saskia; Vincent, Helen A.; Larrea, Michelle; Oriolo, Andrea

2007-01-01

Control of cellular dimensions and cell symmetry are critical for development and differentiation. Here we provide evidence that the putative Rho-GAP Rga4p of Schizosaccharomyces pombe controls cellular dimensions. rga4Δ cells are wider in diameter and shorter in length, whereas Rga4p overexpression leads to reduced diameter of the growing cell tip. Consistent with a negative role in cell growth control, Rga4p protein localizes to the cell sides in a “corset” pattern, and to the nongrowing cell tips. Additionally, rga4Δ cells show an altered growth pattern similar to that observed in mutants of the formin homology protein For3p. Consistent with these observations, Rga4p is required for normal localization of For3p and for normal distribution of the actin cytoskeleton. We show that different domains of the Rga4p protein mediate diverse morphological functions. The C-terminal GAP domain mediates For3p localization to the cell tips and maintains cell diameter. Conversely, overexpression of the N-terminal LIM homology domain of Rga4p promotes actin cable formation in a For3p-dependent manner. Our studies indicate that Rga4p functionally interacts with For3p and has a novel function in the control of cell diameter and cell growth. PMID:17377067

Identification of Baicalin as an Immunoregulatory Compound by Controlling TH17 Cell Differentiation

PubMed Central

Chu, Yiwei; Li, Ming

2011-01-01

TH17 cells have been implicated in a growing list of inflammatory disorders. Antagonism of TH17 cells can be used for the treatment of inflammatory injury. Currently, very little is known about the natural compound controlling the differentiation of TH17 cells. Here, we showed that Baicalin, a compound isolated from a Chinese herb, inhibited TH17 cell differentiation both in vitro and in vivo. Baicalin might inhibit newly generated TH17 cells via reducing RORγt expression, and together with up-regulating Foxp3 expression to suppress RORγt-mediated IL-17 expression in established TH17 cells. In vivo treatment with Baicalin could inhibit TH17 cell differentiation, restrain TH17 cells infiltration into kidney, and protect MRL/lpr mice against nephritis. Our findings not only demonstrate that Baicalin could control TH17 cell differentiation but also suggest that Baicalin might be a promising therapeutic agent for the treatment of TH17 cells-mediated inflammatory diseases. PMID:21359178

Glycolysis controls the induction of human regulatory T cells by modulating the expression of FOXP3 exon 2 splicing variants

PubMed Central

De Rosa, Veronica; Galgani, Mario; Porcellini, Antonio; Colamatteo, Alessandra; Santopaolo, Marianna; Zuchegna, Candida; Romano, Antonella; De Simone, Salvatore; Procaccini, Claudio; La Rocca, Claudia; Carrieri, Pietro Biagio; Maniscalco, Giorgia Teresa; Salvetti, Marco; Buscarinu, Maria Chiara; Franzese, Adriana; Mozzillo, Enza; La Cava, Antonio; Matarese, Giuseppe

2016-01-01

Human regulatory T cells (Treg cells) that develop from conventional T cells (Tconv cells) following suboptimal stimulation via the T cell antigen receptor (TCR) (induced Treg cells (iTreg cells)) express the transcription factor Foxp3, are suppressive, and display an active proliferative and metabolic state. Here we found that the induction and suppressive function of iTreg cells tightly depended on glycolysis, which controlled Foxp3 splicing variants containing exon 2 (Foxp3-E2) through the glycolytic enzyme enolase-1. The Foxp3-E2–related suppressive activity of iTreg cells was altered in human autoimmune diseases, including multiple sclerosis and type 1 diabetes, and was associated with impaired glycolysis and signaling via interleukin 2. This link between glycolysis and Foxp3-E2 variants via enolase-1 shows a previously unknown mechanism for controlling the induction and function of Treg cells in health and in autoimmunity. PMID:26414764

Cyclic adenosine monophosphate modulates cell morphology and behavior of a cultured renal epithelial.

PubMed

Amsler, K

1990-07-01

The role of cyclic adenosine monophosphate (cAMP) dependent protein kinase (PKA) in modulating functions of differentiated renal cells is well established. Its importance in controlling their growth and differentiation is less clear. We have used somatic cell genetic techniques to probe the role of PKA in controlling morphology and behavior of a renal epithelial cell line, LLC-PK1, which acquires many properties characteristic of the renal proximal tubular cell. Mutants of this line altered in PKA activity have been isolated and their behavior compared to that of the parent line. The results indicate that PKA is involved, either directly or indirectly, in maintenance of cell morphology, cell-cell and cell-substratum interactions, density-dependent growth regulation, and expression of one function characteristic of the renal proximal tubular cell, Na-hexose symport. The relevance of these results to the role of PKA in controlling growth and differentiation of renal epithelial cells in vivo is discussed.

Temporal controls of the asymmetric cell division cycle in Caulobacter crescentus.

PubMed

Li, Shenghua; Brazhnik, Paul; Sobral, Bruno; Tyson, John J

2009-08-01

The asymmetric cell division cycle of Caulobacter crescentus is orchestrated by an elaborate gene-protein regulatory network, centered on three major control proteins, DnaA, GcrA and CtrA. The regulatory network is cast into a quantitative computational model to investigate in a systematic fashion how these three proteins control the relevant genetic, biochemical and physiological properties of proliferating bacteria. Different controls for both swarmer and stalked cell cycles are represented in the mathematical scheme. The model is validated against observed phenotypes of wild-type cells and relevant mutants, and it predicts the phenotypes of novel mutants and of known mutants under novel experimental conditions. Because the cell cycle control proteins of Caulobacter are conserved across many species of alpha-proteobacteria, the model we are proposing here may be applicable to other genera of importance to agriculture and medicine (e.g., Rhizobium, Brucella).

Panax Notoginseng Saponin Controls IL-17 Expression in Helper T Cells

PubMed Central

Wei, Jia-Ru; Wen, Xiaofeng; Bible, Paul W.; Li, Zhiyu; Nussenblatt, Robert B.

2017-01-01

Abstract Purpose: Panax Notoginseng, a traditional Chinese medicine, is known as an anti-inflammatory herb. However, the molecular mechanism by which it controls helper T cell mediated immune responses is largely unknown. Methods: Naive CD4+ T cells isolated from healthy donors, patients with Behcet's disease, and C57BL/6 mice were polarized into Th1, Th17, and Treg cells. Proliferation and cytokine expression were measured in these cells with the presence or absence of Panax Notoginseng saponins (PNS). Genomewide expression profiles of Th1, Th17, and Treg cells were assessed using Affymetrix microarray analysis. Results: We found that PNS control the proliferation and differentiation of Th17 cells by globally downregulating the expression of inflammatory cytokines and cell cycle genes. Conclusions: These findings demonstrated that PNS function as an anti-inflammatory agent through directly targeting Th17 cell mediated immune response. PMID:28051353

Fuel cell generator with fuel electrodes that control on-cell fuel reformation

DOEpatents

Ruka, Roswell J [Pittsburgh, PA; Basel, Richard A [Pittsburgh, PA; Zhang, Gong [Murrysville, PA

2011-10-25

A fuel cell for a fuel cell generator including a housing including a gas flow path for receiving a fuel from a fuel source and directing the fuel across the fuel cell. The fuel cell includes an elongate member including opposing first and second ends and defining an interior cathode portion and an exterior anode portion. The interior cathode portion includes an electrode in contact with an oxidant flow path. The exterior anode portion includes an electrode in contact with the fuel in the gas flow path. The anode portion includes a catalyst material for effecting fuel reformation along the fuel cell between the opposing ends. A fuel reformation control layer is applied over the catalyst material for reducing a rate of fuel reformation on the fuel cell. The control layer effects a variable reformation rate along the length of the fuel cell.

Course 6: Physics of Composite Cell Membrane and Actin Based Cytoskeleton

NASA Astrophysics Data System (ADS)

Sackmann, E.; Bausch, A. R.; Vonna, L.

1 Architecture of composite cell membranes 1.1 The lipid/protein bilayer is a multicomponent smectic phase with mosaic like architecture 1.2 The spectrin/actin cytoskeleton as hyperelastic cell stabilizer 1.3 The actin cortex: Architecture and function 2 Physics of the actin based cytoskeleton 2.1 Actin is a living semiflexible polymer 2.2 Actin network as viscoelastic body 2.3 Correlation between macroscopic viscoelasticity and molecular 3 Heterogeneous actin gels in cells and biological function 3.1 Manipulation of actin gels 3.2 Control of organization and function of actin cortex by cell signalling 4 Micromechanics and microrheometry of cells 5 Activation of endothelial cells: On the possibility of formation of stress fibers as phase transition of actin-network triggered by cell signalling pathways 6 On cells as adaptive viscoplastic bodies 7 Controll of cellular protrusions controlled by actin/myosin cortex

Life sciences, biotechnology, and microgravity

NASA Technical Reports Server (NTRS)

Hymer, W. C.; Hayes, C.; Grindeland, R.; Lanhan, J. W.; Morrison, D.

1987-01-01

Growth hormone (GH) studies on rats flown aboard Spacelab 3 are discussed, and evidence for the direct effect of microgravity on cell function is reviewed. SL-3 rat GH cells were found to experience a secretory lesion (they contained more hormone per cell, but released less per cell relative to controls). Pituitary cell culture experiments on the STS-8 mission showed that GH cells did not subsequently release as much hormone as did control cells, indicating a secretory lesion. Changes in bone and muscle noted in SL-3 rats are related to GH cell findings.

Specification and spatial arrangement of cells in the germline stem cell niche of the Drosophila ovary depend on the Maf transcription factor Traffic jam

PubMed Central

Panchal, Trupti; Chen, Xi; Poon, James; Kouptsova, Jane

2017-01-01

Germline stem cells in the Drosophila ovary are maintained by a somatic niche. The niche is structurally and functionally complex and contains four cell types, the escort, cap, and terminal filament cells and the newly identified transition cell. We find that the large Maf transcription factor Traffic jam (Tj) is essential for determining niche cell fates and architecture, enabling each niche in the ovary to support a normal complement of 2–3 germline stem cells. In particular, we focused on the question of how cap cells form. Cap cells express Tj and are considered the key component of a mature germline stem cell niche. We conclude that Tj controls the specification of cap cells, as the complete loss of Tj function caused the development of additional terminal filament cells at the expense of cap cells, and terminal filament cells developed cap cell characteristics when induced to express Tj. Further, we propose that Tj controls the morphogenetic behavior of cap cells as they adopted the shape and spatial organization of terminal filament cells but otherwise appeared to retain their fate when Tj expression was only partially reduced. Our data indicate that Tj contributes to the establishment of germline stem cells by promoting the cap cell fate, and controls the stem cell-carrying capacity of the niche by regulating niche architecture. Analysis of the interactions between Tj and the Notch (N) pathway indicates that Tj and N have distinct functions in the cap cell specification program. We propose that formation of cap cells depends on the combined activities of Tj and the N pathway, with Tj promoting the cap cell fate by blocking the terminal filament cell fate, and N supporting cap cells by preventing the escort cell fate and/or controlling the number of cap cell precursors. PMID:28542174

Air-Adapted Methanosarcina acetivorans Shows High Methane Production and Develops Resistance against Oxygen Stress

PubMed Central

Jasso-Chávez, Ricardo; Santiago-Martínez, M. Geovanni; Lira-Silva, Elizabeth; Pineda, Erika; Zepeda-Rodríguez, Armando; Belmont-Díaz, Javier; Encalada, Rusely; Saavedra, Emma; Moreno-Sánchez, Rafael

2015-01-01

Methanosarcina acetivorans, considered a strict anaerobic archaeon, was cultured in the presence of 0.4–1% O2 (atmospheric) for at least 6 months to generate air-adapted cells; further, the biochemical mechanisms developed to deal with O2 were characterized. Methane production and protein content, as indicators of cell growth, did not change in air-adapted cells respect to cells cultured under anoxia (control cells). In contrast, growth and methane production significantly decreased in control cells exposed for the first time to O2. Production of reactive oxygen species was 50 times lower in air-adapted cells versus control cells, suggesting enhanced anti-oxidant mechanisms that attenuated the O2 toxicity. In this regard, (i) the transcripts and activities of superoxide dismutase, catalase and peroxidase significantly increased; and (ii) the thiol-molecules (cysteine + coenzyme M-SH + sulfide) and polyphosphate contents were respectively 2 and 5 times higher in air-adapted cells versus anaerobic-control cells. Long-term cultures (18 days) of air-adapted cells exposed to 2% O2 exhibited the ability to form biofilms. These data indicate that M. acetivorans develops multiple mechanisms to contend with O2 and the associated oxidative stress, as also suggested by genome analyses for some methanogens. PMID:25706146

The glycoconjugate sugar residues of the sessile and motile cells in the thymus of normal and cyclosporin-A-treated rats: lectin histochemistry.

PubMed

Gheri, G; Gheri Bryk, S; Riccardi, R; Sgambati, E; Cirri Borghi, M B

2002-01-01

It is well known that cell surface glycoconjugates play a determinant role in cellular recognition, cell-to-cell adhesion and serve as receptor molecules. T-lymphocytes are in strict contact with the thymic epithelial cells, which control their process of maturation and proliferation. On the other hand the normal maturation of the epithelial cells is believed to be induced by T-lymphocytes. For these reasons we have studied the glycoconjugates saccharidic moieties of the sessile and motile cells in the thymus of normal male albino Wistar rats and their changes following cyclosporin-A treatment, using a battery of seven HRP-lectins. Cytochemical controls were performed for specificity of lectin-sugar reaction. Some sections were pre-treated with neuraminidase prior to staining with HRP-lectins. Our results have demonstrated, in the control rats, a large amount and a variety of terminal and subterminal oligosaccharides within and/or on the epithelial thymic cells and in macrophages. After cyclosporin-A treatment, among the thymic epithelial cells, the subcapsular, paraseptal and perivascular cells showed the loss of some sugar residues, which characterized the same cells in the intact thymus. Some hypotheses are reported on the role played by the glycoconjugate sugar residues in control and cyclosporin-A treated rats.

Immune cell populations within the duodenal mucosa of dogs with enteropathies.

PubMed

German, A J; Hall, E J; Day, M J

2001-01-01

The mucosal immune system may play a critical role in the pathogenesis of small intestinal enteropathies. The aim of the current study was to assess mucosal immune cell populations in dogs with inflammatory bowel disease (IBD), idiopathic antibiotic-responsive diarrhea (ARD), and adverse reactions to food (FR). Endoscopic biopsies were performed of the duodenum of dogs with these conditions and from a group of dogs without enteric disease. Additional control samples were collected after death from other dogs that did not have evidence of enteric disease. Immunohistochemistry and computer-aided morphometry were used to assess the distribution of immune cell subsets in both lamina propria and intestinal epithelium. Compared with controls, dogs with ARD had increased numbers of lamina propria immunoglobulin (Ig) A- plasma cells and CD4+ cells. More marked alterations were noted in dogs with IBD, with significant increases in lamina propria IgG+ plasma cells, T cells (CD3+), CD4+ cells, macrophages, and neutrophils, but with reduced mast cell numbers. Increased intraepithelial CD3+ T cells were also present in the dogs with IBD, compared with controls. However, lamina propria and epithelial populations were unaltered in dogs with FR when compared with controls. The altered mucosal immune cell populations observed in dogs with ARD or IBD may reflect an underlying immunologic pathogenesis in these disorders.

Mycobacteria Modify Their Cell Size Control under Sub-Optimal Carbon Sources

PubMed Central

Priestman, Miles; Thomas, Philipp; Robertson, Brian D.; Shahrezaei, Vahid

2017-01-01

The decision to divide is the most important one that any cell must make. Recent single cell studies suggest that most bacteria follow an “adder” model of cell size control, incorporating a fixed amount of cell wall material before dividing. Mycobacteria, including the causative agent of tuberculosis Mycobacterium tuberculosis, are known to divide asymmetrically resulting in heterogeneity in growth rate, doubling time, and other growth characteristics in daughter cells. The interplay between asymmetric cell division and adder size control has not been extensively investigated. Moreover, the impact of changes in the environment on growth rate and cell size control have not been addressed for mycobacteria. Here, we utilize time-lapse microscopy coupled with microfluidics to track live Mycobacterium smegmatis cells as they grow and divide over multiple generations, under a variety of growth conditions. We demonstrate that, under optimal conditions, M. smegmatis cells robustly follow the adder principle, with constant added length per generation independent of birth size, growth rate, and inherited pole age. However, the nature of the carbon source induces deviations from the adder model in a manner that is dependent on pole age. Understanding how mycobacteria maintain cell size homoeostasis may provide crucial targets for the development of drugs for the treatment of tuberculosis, which remains a leading cause of global mortality. PMID:28748182

Myocilin Regulates Cell Proliferation and Survival*

PubMed Central

Joe, Myung Kuk; Kwon, Heung Sun; Cojocaru, Radu; Tomarev, Stanislav I.

2014-01-01

Myocilin, a causative gene for open angle glaucoma, encodes a secreted glycoprotein with poorly understood functions. To gain insight into its functions, we produced a stably transfected HEK293 cell line expressing myocilin under an inducible promoter and compared gene expression profiles between myocilin-expressing and vector control cell lines by a microarray analysis. A significant fraction of differentially expressed genes in myocilin-expressing cells was associated with cell growth and cell death, suggesting that myocilin may have a role in the regulation of cell growth and survival. Increased proliferation of myocilin-expressing cells was demonstrated by the WST-1 proliferation assay, direct cell counting, and immunostaining with antibodies against Ki-67, a cellular proliferation marker. Myocilin-containing conditioned medium also increased proliferation of unmodified HEK293 cells. Myocilin-expressing cells were more resistant to serum starvation-induced apoptosis than control cells. TUNEL-positive apoptotic cells were dramatically decreased, and two apoptotic marker proteins, cleaved caspase 7 and cleaved poly(ADP-ribose) polymerase, were significantly reduced in myocilin-expressing cells as compared with control cells under apoptotic conditions. In addition, myocilin-deficient mesenchymal stem cells exhibited reduced proliferation and enhanced susceptibility to serum starvation-induced apoptosis as compared with wild-type mesenchymal stem cells. Phosphorylation of ERK1/2 and its upstream kinases, c-Raf and MEK, was increased in myocilin-expressing cells compared with control cells. Elevated phosphorylation of ERK1/2 was also observed in the trabecular meshwork of transgenic mice expressing 6-fold higher levels of myocilin when compared with their wild-type littermates. These results suggest that myocilin promotes cell proliferation and resistance to apoptosis via the ERK1/2 MAPK signaling pathway. PMID:24563482

Diversity in TAF proteomics: consequences for cellular differentiation and migration.

PubMed

Kazantseva, Jekaterina; Palm, Kaia

2014-09-19

Development is a highly controlled process of cell proliferation and differentiation driven by mechanisms of dynamic gene regulation. Specific DNA binding factors for establishing cell- and tissue-specific transcriptional programs have been characterised in different cell and animal models. However, much less is known about the role of "core transcription machinery" during cell differentiation, given that general transcription factors and their spatiotemporally patterned activity govern different aspects of cell function. In this review, we focus on the role of TATA-box associated factor 4 (TAF4) and its functional isoforms generated by alternative splicing in controlling lineage-specific differentiation of normal mesenchymal stem cells and cancer stem cells. In the light of our recent findings, induction, control and maintenance of cell differentiation status implies diversification of the transcription initiation apparatus orchestrated by alternative splicing.

Cosmos: 1989 immunology studies

NASA Technical Reports Server (NTRS)

Sonnenfeld, Gerald

1991-01-01

The effects of flight on Cosmos mission 2044 on leukocyte subset distribution and the sensitivity of bone marrow cells to colony stimulating factor-GM were determined. A parallel study with antiorthostatic suspension was also carried out. The study involved repetition and expansion of studies performed on Cosmos 1887. Spleen and bone marrow cells were obtained from flown, vivarium control, synchronous control, and suspended rats. The cells were stained with a series of monoclonal antibodies directed against rat leukocyte cell surface antigens. Control cells were stained with a monoclonal antibody directed against an irrelevant species or were unstained. Cells were then analyzed for fluorescence using a FACSCAN flow cytometer. Bone marrow cells were placed in culture with GM-CSF in McCoy's 5a medium and incubated for 5 days. Cultures were then evaluated for the number of colonies of 50 cells or greater.

Signaling-Dependent Control of Apical Membrane Size and Self-Renewal in Rosette-Stage Human Neuroepithelial Stem Cells.

PubMed

Medelnik, Jan-Philip; Roensch, Kathleen; Okawa, Satoshi; Del Sol, Antonio; Chara, Osvaldo; Mchedlishvili, Levan; Tanaka, Elly M

2018-06-05

In the developing nervous system, neural stem cells are polarized and maintain an apical domain facing a central lumen. The presence of apical membrane is thought to have a profound influence on maintaining the stem cell state. With the onset of neurogenesis, cells lose their polarization, and the concomitant loss of the apical domain coincides with a loss of the stem cell identity. Little is known about the molecular signals controlling apical membrane size. Here, we use two neuroepithelial cell systems, one derived from regenerating axolotl spinal cord and the other from human embryonic stem cells, to identify a molecular signaling pathway initiated by lysophosphatidic acid that controls apical membrane size and consequently controls and maintains epithelial organization and lumen size in neuroepithelial rosettes. This apical domain size increase occurs independently of effects on proliferation and involves a serum response factor-dependent transcriptional induction of junctional and apical membrane components. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Remotely controlled fusion of selected vesicles and living cells: a key issue review

NASA Astrophysics Data System (ADS)

Bahadori, Azra; Moreno-Pescador, Guillermo; Oddershede, Lene B.; Bendix, Poul M.

2018-03-01

Remote control over fusion of single cells and vesicles has a great potential in biological and chemical research allowing both transfer of genetic material between cells and transfer of molecular content between vesicles. Membrane fusion is a critical process in biology that facilitates molecular transport and mixing of cellular cytoplasms with potential formation of hybrid cells. Cells precisely regulate internal membrane fusions with the aid of specialized fusion complexes that physically provide the energy necessary for mediating fusion. Physical factors like membrane curvature, tension and temperature, affect biological membrane fusion by lowering the associated energy barrier. This has inspired the development of physical approaches to harness the fusion process at a single cell level by using remotely controlled electromagnetic fields to trigger membrane fusion. Here, we critically review various approaches, based on lasers or electric pulses, to control fusion between individual cells or between individual lipid vesicles and discuss their potential and limitations for present and future applications within biochemistry, biology and soft matter.

High-throughput microfluidics to control and measure signaling dynamics in single yeast cells

PubMed Central

Hansen, Anders S.; Hao, Nan; O'Shea, Erin K.

2015-01-01

Microfluidics coupled to quantitative time-lapse fluorescence microscopy is transforming our ability to control, measure, and understand signaling dynamics in single living cells. Here we describe a pipeline that incorporates multiplexed microfluidic cell culture, automated programmable fluid handling for cell perturbation, quantitative time-lapse microscopy, and computational analysis of time-lapse movies. We illustrate how this setup can be used to control the nuclear localization of the budding yeast transcription factor Msn2. Using this protocol, we generate oscillations of Msn2 localization and measure the dynamic gene expression response of individual genes in single cells. The protocol allows a single researcher to perform up to 20 different experiments in a single day, whilst collecting data for thousands of single cells. Compared to other protocols, the present protocol is relatively easy to adopt and higher-throughput. The protocol can be widely used to control and monitor single-cell signaling dynamics in other signal transduction systems in microorganisms. PMID:26158443

Control of cell respiration by nitric oxide in Ataxia Telangiectasia lymphoblastoid cells.

PubMed

Masci, Alessandra; Mastronicola, Daniela; Arese, Marzia; Piane, Maria; De Amicis, Andrea; Blanck, Thomas J J; Chessa, Luciana; Sarti, Paolo

2008-01-01

Ataxia Telangiectasia (AT) patients are particularly sensitive to oxidative-nitrosative stress. Nitric oxide (NO) controls mitochondrial respiration via the reversible inhibition of complex IV. The mitochondrial response to NO of AT lymphoblastoid cells was investigated. Cells isolated from three patients and three intrafamilial healthy controls were selected showing within each group a normal diploid karyotype and homogeneous telomere length. Different complex IV NO-inhibition patterns were induced by varying the electron flux through the respiratory chain, using exogenous cell membrane permeable electron donors. Under conditions of high electron flux the mitochondrial NO inhibition of respiration was greater in AT than in control cells (P< or =0.05). This property appears peculiar to AT, and correlates well to the higher concentration of cytochrome c detected in the AT cells. This finding is discussed on the basis of the proposed mechanism of reaction of NO with complex IV. It is suggested that the peculiar response of AT mitochondria to NO stress may be relevant to the mitochondrial metabolism of AT patients.

Effect of Removal of Spermatogonial Stem Cells (SSCs) from In Vitro Culture on Gene Expression of Niche Factors in Bovine

PubMed Central

Akbarinejad, Vahid; Tajik, Parviz; Movahedin, Mansoureh; Youssefi, Reza

2016-01-01

Background: Niche cells, regulating Spermatogonial Stem Cells (SSCs) fate are believed to have a reciprocal communication with SSCs. The present study was conducted to evaluate the effect of SSC elimination on the gene expression of Glial cell line-Derived Neurotrophic Factor (GDNF), Fibroblast Growth Factor 2 (FGF2) and Kit Ligand (KITLG), which are the main growth factors regulating SSCs development and secreted by niche cells, primarily Sertoli cells. Methods: Following isolation, bovine testicular cells were cultured for 12 days on extracellular matrix-coated plates. In the germ cell-removed group, the SSCs were removed from the in vitro culture using differential plating; however, in the control group, no intervention in the culture was performed. Colony formation of SSCs was evaluated using an inverted microscope. The gene expression of growth factors and spermatogonia markers were assessed using quantitative real time PCR. Results: SSCs colonies were developed in the control group but they were rarely observed in the germ cell-removed group; moreover, the expression of spermatogonia markers was detected in the control group while it was not observed in the germ cell-removed group, substantiating the success of SSCs removal. The expression of Gdnf and Fgf2 was greater in the germ cell-removed than control group (p<0.05), whereas the expression of Kitlg was lower in the germ cell-removed than control group (p< 0.05). Conclusion: In conclusion, the results revealed that niche cells respond to SSCs removal by upregulation of GDNF and FGF2, and downregulation of KITLG in order to stimulate self-renewal and arrest differentiation. PMID:27563426

Interactions between IGF-I, estrogen receptor-α (ERα), and ERβ in regulating growth/apoptosis of MCF-7 human breast cancer cells.

PubMed

Mendoza, Rhone A; Enriquez, Marlene I; Mejia, Sylvia M; Moody, Emily E; Thordarson, Gudmundur

2011-01-01

Understanding of the interactions between estradiol (E₂) and IGF-I is still incomplete. Cell lines derived from the MCF-7 breast cancer cells were generated with suppressed expression of the IGF-I receptor (IGF-IR), termed IGF-IR.low cells, by stable transfection using small interfering RNA (siRNA) expression vector. Vector for control cells carried sequence generating noninterfering RNA. Concomitant with reduction in the IGF-IR levels, the IGF-IR.low cells also showed a reduction in estrogen receptor α (ERα) and progesterone receptor expressions, and an elevation in the expression of ERβ. The number of the IGF-IR.low cells was reduced in response to IGF-I and human GH plus epidermal growth factor, but E₂ did not cause an increase in the number of the IGF-IR.low cells compared to controls. The proliferation rate of IGF-IR.low cells was only reduced in response to E₂ compared to controls, whereas their basal and hormone-stimulated apoptosis rate was increased. Phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) was increased in the IGF-IR.low cells after treatment with E₂, without affecting control cells. Furthermore, phosphorylation of the tumor suppressor protein p53 was elevated in the IGF-IR.low cells compared to the controls. In conclusion, suppressing IGF-IR expression decreased the level of ERα but increased the level of ERβ. Overall growth rate of the IGF-IR.low cells was reduced mostly through an increase in apoptosis without affecting proliferation substantially. We hypothesize that a decreased ERα:ERβ ratio triggered a rapid phosphorylation of p38 MAPK, which in turn phosphorylated the p53 tumor suppressor and accelerated apoptosis rate.

Experiment K-6-23. Effect of spaceflight on levels and function of immune cells

NASA Technical Reports Server (NTRS)

Mandel, A. D.; Sonnenfeld, G.; Berry, W.; Taylor, G.; Wellhausen, S. R.; Konstantinova, I.; Lesnyak, A.; Fuchs, B.

1990-01-01

Two different immunology experiments were performed on samples received from rats flown on Cosmos 1887. In the first experiment, rat bone marrow cells were examined in Moscow for their response to colony stimulating factor-M. In the second experiment, rat spleen and bone marrow cells were stained in Moscow with a variety of antibodies directed against cell surface antigenic markers. These cells were preserved and shipped to the United States where they were subjected to analysis on a flow cytometer. The results of the studies indicate that bone marrow cells from flown rats showed a decreased response to colony stimulating factor than did bone marrow cells from control rats. There was a higher percentage of spleen cells from flown rats staining positively for pan-T-cell, suppressor-T-cell and innate interleukin-2 receptor antigens than from control animals. In addition, a higher percentage of cells that appeared to be part of the myelogenous population of bone marrow cells from flown rats stained positively for surface immunoglobulin than did equivalent cells from control rats.

Cell confinement controls centrosome positioning and lumen initiation during epithelial morphogenesis

PubMed Central

Rodríguez-Fraticelli, Alejo E.; Auzan, Muriel; Alonso, Miguel A.; Bornens, Michel

2012-01-01

Epithelial organ morphogenesis involves sequential acquisition of apicobasal polarity by epithelial cells and development of a functional lumen. In vivo, cells perceive signals from components of the extracellular matrix (ECM), such as laminin and collagens, as well as sense physical conditions, such as matrix stiffness and cell confinement. Alteration of the mechanical properties of the ECM has been shown to promote cell migration and invasion in cancer cells, but the effects on epithelial morphogenesis have not been characterized. We analyzed the effects of cell confinement on lumen morphogenesis using a novel, micropatterned, three-dimensional (3D) Madin-Darby canine kidney cell culture method. We show that cell confinement, by controlling cell spreading, limits peripheral actin contractility and promotes centrosome positioning and lumen initiation after the first cell division. In addition, peripheral actin contractility is mediated by master kinase Par-4/LKB1 via the RhoA–Rho kinase–myosin II pathway, and inhibition of this pathway restores lumen initiation in minimally confined cells. We conclude that cell confinement controls nuclear–centrosomal orientation and lumen initiation during 3D epithelial morphogenesis. PMID:22965908

The Developmental Regulator SEEDSTICK Controls Structural and Mechanical Properties of the Arabidopsis Seed Coat

PubMed Central

Beauzamy, Léna; Caporali, Elisabetta; Koroney, Abdoul-Salam

2016-01-01

Although many transcription factors involved in cell wall morphogenesis have been identified and studied, it is still unknown how genetic and molecular regulation of cell wall biosynthesis is integrated into developmental programs. We demonstrate by molecular genetic studies that SEEDSTICK (STK), a transcription factor controlling ovule and seed integument identity, directly regulates PMEI6 and other genes involved in the biogenesis of the cellulose-pectin matrix of the cell wall. Based on atomic force microscopy, immunocytochemistry, and chemical analyses, we propose that structural modifications of the cell wall matrix in the stk mutant contribute to defects in mucilage release and seed germination under water-stress conditions. Our studies reveal a molecular network controlled by STK that regulates cell wall properties of the seed coat, demonstrating that developmental regulators controlling organ identity also coordinate specific aspects of cell wall characteristics. PMID:27624758

Changes in pituitary growth hormone cells prepared from rats flown on Spacelab 3

NASA Technical Reports Server (NTRS)

Grindeland, R.; Hymer, W. C.; Farrington, M.; Fast, T.; Hayes, C.; Motter, K.; Patil, L.; Vasques, M.

1987-01-01

The effect of exposure to microgravity on pituitary gland was investigated by examining cells isolated from anterior pituitaries of rats flown on the 7-day Spacelab 3 mission and, subsequently, cultured for 6 days. Compared with ground controls, flight cells contained more intracellular growth hormone (GH); however, the flight cells released less GH over the 6-day culture period and after implantation into hypophysectomized rats than did the control cells. Compared with control rats, glands from large rats (400 g) contained more somatotrophs (44 percent compared with 37 percent in control rats); small rats (200 g) showed no difference. No major differences were found in the somatotroph ultrastructure (by TEM) or in the pattern of the immunoactive GH variants. However, high-performance liquid chromatography fractionation of culture media indicated that flight cells released much less of a biologically active high-molecular weight GH variant, suggesting that space flight may lead to secretory dysfunction.

Reducing juvenile recidivism with cognitive training and a cell phone follow-up: an evaluation of the realvictory program.

PubMed

Burraston, Bert O; Cherrington, David J; Bahr, Stephen J

2012-02-01

The purpose of this research was to evaluate the effects of a cognitive training and cell phone intervention on the recidivism of 70 juvenile offenders. Median days to rearrest were 106 for the control group, 191 for the class-only group, and 278 for the class plus cell phone group. Using rearrest as the survival criterion, the survival ratios of the class-only and class plus cell phone groups were 2.64 and 2.94 times longer than the control group, respectively. After controlling for gender, prior arrests, and risk score, the Poisson regression indicated that the class-only and class plus cell phone groups were 51% lower in total arrests than the control group. These results suggest that cognitive training supplemented with a cell phone coach is an effective and cost-efficient intervention for reducing recidivism.

Innate myeloid cell TNFR1 mediates first line defence against primary Mycobacterium tuberculosis infection.

PubMed Central

Segueni, Noria; Benmerzoug, Sulayman; Rose, Stéphanie; Gauthier, Amandine; Bourigault, Marie-Laure; Reverchon, Flora; Philippeau, Amandine; Erard, François; Le Bert, Marc; Bouscayrol, Hélène; Wachter, Thierry; Garcia, Irène; Kollias, George; Jacobs, Muazzam; Ryffel, Bernhard; Quesniaux, Valerie F.J.

2016-01-01

TNF is crucial for controlling Mycobacterium tuberculosis infection and understanding how will help immunomodulating the host response. Here we assessed the contribution of TNFR1 pathway from innate myeloid versus T cells. We first established the prominent role of TNFR1 in haematopoietic cells for controlling M. tuberculosis in TNFR1 KO chimera mice. Further, absence of TNFR1 specifically on myeloid cells (M-TNFR1 KO) recapitulated the uncontrolled M. tuberculosis infection seen in fully TNFR1 deficient mice, with increased bacterial burden, exacerbated lung inflammation, and rapid death. Pulmonary IL-12p40 over-expression was attributed to a prominent CD11b+ Gr1high cell population in infected M-TNFR1 KO mice. By contrast, absence of TNFR1 on T-cells did not compromise the control of M. tuberculosis infection over 6-months. Thus, the protective TNF/TNFR1 pathway essential for controlling primary M. tuberculosis infection depends on innate macrophage and neutrophil myeloid cells, while TNFR1 pathway in T cells is dispensable. PMID:26931771

Galaptin Mediates the Effect of Hypergravity on Vascular Smooth Muscle cell (SMC) Adhesion to Laminin Containing Matrices

NASA Technical Reports Server (NTRS)

Enahora, Fatisha T.; Bosah, Francis N.; Harris-Hooker, Sandra; Sanford, Gary L.

1997-01-01

Galaptin, an endogenous beta-galactoside specific lectin, has been reported to bind to laminin and subsequently decrease the binding of SMC. Cellular function depend on cell:matrix interactions. Hypergravity (HGrav) affect a number of cellular functions, yet little is known about its affect on cell adhesion. We examined the possibility that galaptin mediates the effects of hypergravity on SMC adherence. Confluent primate aorta SMC cultures were subjected to Hgrav (centrifuged at 6G) for 24 and 48 hr. Cells were non-enzymatically dispersed, pretreated with antisense (AS-oligo) or control sense (SS-oligo) oligonucleotides to galaptin mRNA (0.01 micro g/ml), then seeded in uncoated or ECL-matrix coated plates. Adhesion of cells were monitored after 6 hr. HGrav increased adhesion by 100-300% compared to controls. AS-oligo decreased adhesion for both HGrav and control cells. SS-oligo did not affect adhesion for either HGrav or control cells. These studies show that HGrav affects cell adhesion and that galaptin expression is required for this effect.

A parallel row-based algorithm with error control for standard-cell replacement on a hypercube multiprocessor

NASA Technical Reports Server (NTRS)

Sargent, Jeff Scott

1988-01-01

A new row-based parallel algorithm for standard-cell placement targeted for execution on a hypercube multiprocessor is presented. Key features of this implementation include a dynamic simulated-annealing schedule, row-partitioning of the VLSI chip image, and two novel new approaches to controlling error in parallel cell-placement algorithms; Heuristic Cell-Coloring and Adaptive (Parallel Move) Sequence Control. Heuristic Cell-Coloring identifies sets of noninteracting cells that can be moved repeatedly, and in parallel, with no buildup of error in the placement cost. Adaptive Sequence Control allows multiple parallel cell moves to take place between global cell-position updates. This feedback mechanism is based on an error bound derived analytically from the traditional annealing move-acceptance profile. Placement results are presented for real industry circuits and the performance is summarized of an implementation on the Intel iPSC/2 Hypercube. The runtime of this algorithm is 5 to 16 times faster than a previous program developed for the Hypercube, while producing equivalent quality placement. An integrated place and route program for the Intel iPSC/2 Hypercube is currently being developed.

Imparting magnetic dipole heterogeneity to internalized iron oxide nanoparticles for microorganism swarm control

NASA Astrophysics Data System (ADS)

Kim, Paul Seung Soo; Becker, Aaron; Ou, Yan; Julius, Anak Agung; Kim, Min Jun

2015-03-01

Tetrahymena pyriformis is a single cell eukaryote that can be modified to respond to magnetic fields, a response called magnetotaxis. Naturally, this microorganism cannot respond to magnetic fields, but after modification using iron oxide nanoparticles, cells are magnetized and exhibit a constant magnetic dipole strength. In experiments, a rotating field is applied to cells using a two-dimensional approximate Helmholtz coil system. Using rotating magnetic fields, we characterize discrete cells' swarm swimming which is affected by several factors. The behavior of the cells under these fields is explained in detail. After the field is removed, relatively straight swimming is observed. We also generate increased heterogeneity within a population of cells to improve controllability of a swarm, which is explored in a cell model. By exploiting this straight swimming behavior, we propose a method to control discrete cells utilizing a single global magnetic input. Successful implementation of this swarm control method would enable teams of microrobots to perform a variety of in vitro microscale tasks impossible for single microrobots, such as pushing objects or simultaneous micromanipulation of discrete entities.

Femtosecond laser fabricated spike structures for selective control of cellular behavior.

PubMed

Schlie, Sabrina; Fadeeva, Elena; Koch, Jürgen; Ngezahayo, Anaclet; Chichkov, Boris N

2010-09-01

In this study we investigate the potential of femtosecond laser generated micrometer sized spike structures as functional surfaces for selective cell controlling. The spike dimensions as well as the average spike to spike distance can be easily tuned by varying the process parameters. Moreover, negative replications in soft materials such as silicone elastomer can be produced. This allows tailoring of wetting properties of the spike structures and their negative replicas representing a reduced surface contact area. Furthermore, we investigated material effects on cellular behavior. By comparing human fibroblasts and SH-SY5Y neuroblastoma cells we found that the influence of the material was cell specific. The cells not only changed their morphology, but also the cell growth was affected. Whereas, neuroblastoma cells proliferated at the same rate on the spike structures as on the control surfaces, the proliferation of fibroblasts was reduced by the spike structures. These effects can result from the cell specific adhesion patterns as shown in this work. These findings show a possibility to design defined surface microstructures, which could control cellular behavior in a cell specific manner.

Regional control of Drosophila gut stem cell proliferation: EGF establishes GSSC proliferative set point & controls emergence from quiescence.

PubMed

Strand, Marie; Micchelli, Craig A

2013-01-01

Adult stem cells vary widely in their rates of proliferation. Some stem cells are constitutively active, while others divide only in response to injury. The mechanism controlling this differential proliferative set point is not well understood. The anterior-posterior (A/P) axis of the adult Drosophila midgut has a segmental organization, displaying physiological compartmentalization and region-specific epithelia. These distinct midgut regions are maintained by defined stem cell populations with unique division schedules, providing an excellent experimental model with which to investigate this question. Here, we focus on the quiescent gastric stem cells (GSSCs) of the acidic copper cell region (CCR), which exhibit the greatest period of latency between divisions of all characterized gut stem cells, to define the molecular basis of differential stem cell activity. Our molecular genetic analysis demonstrates that the mitogenic EGF signaling pathway is a limiting factor controlling GSSC proliferation. We find that under baseline conditions, when GSSCs are largely quiescent, the lowest levels of EGF ligands in the midgut are found in the CCR. However, acute epithelial injury by enteric pathogens leads to an increase in EGF ligand expression in the CCR and rapid expansion of the GSSC lineage. Thus, the unique proliferative set points for gut stem cells residing in physiologically distinct compartments are governed by regional control of niche signals along the A/P axis.

Molecular control of brain size: Regulators of neural stem cell life, death and beyond

DOE Office of Scientific and Technical Information (OSTI.GOV)

Joseph, Bertrand; Hermanson, Ola, E-mail: ola.hermanson@ki.se

2010-05-01

The proper development of the brain and other organs depends on multiple parameters, including strictly controlled expansion of specific progenitor pools. The regulation of such expansion events includes enzymatic activities that govern the correct number of specific cells to be generated via an orchestrated control of cell proliferation, cell cycle exit, differentiation, cell death etc. Certain proteins in turn exert direct control of these enzymatic activities and thus progenitor pool expansion and organ size. The members of the Cip/Kip family (p21Cip1/p27Kip1/p57Kip2) are well-known regulators of cell cycle exit that interact with and inhibit the activity of cyclin-CDK complexes, whereas membersmore » of the p53/p63/p73 family are traditionally associated with regulation of cell death. It has however become clear that the roles for these proteins are not as clear-cut as initially thought. In this review, we discuss the roles for proteins of the Cip/Kip and p53/p63/p73 families in the regulation of cell cycle control, differentiation, and death of neural stem cells. We suggest that these proteins act as molecular interfaces, or 'pilots', to assure the correct assembly of protein complexes with enzymatic activities at the right place at the right time, thereby regulating essential decisions in multiple cellular events.« less

Unique roles of estrogen-dependent Pten control in epithelial cell homeostasis of mouse vagina.

PubMed

Miyagawa, S; Sato, M; Sudo, T; Yamada, G; Iguchi, T

2015-02-19

Numerous studies support a role of phosphatase and tensin homolog deleted from chromosome 10 (Pten) as a tumor suppressor gene that controls epithelial cell homeostasis to prevent tumor formation. Mouse vaginal epithelium cyclically exhibits cell proliferation and differentiation in response to estrogen and provides a unique model for analyzing homeostasis of stratified squamous epithelia. We analyzed vaginal epithelium-specific Pten conditional knockout (CKO) mice to provide new insights into Pten/phosphoinositide-3-kinase (PI3K)/Akt function. The vaginal epithelium of ovariectomized (OVX) mice (control) was composed of 1-2 layers of cuboidal cells, whereas OVX CKO mice exhibited epithelial hyperplasia in the suprabasal cells with increased cell mass and mucin production. This is possibly due to misactivation of mammalian target of rapamycin and mitogen-activated protein kinase. Intriguingly, estrogen administration to OVX Pten CKO mice induced stratification and keratinized differentiation in the vaginal epithelium, as in estrogen-treated controls. We found that Pten is exclusively expressed in the suprabasal cells in the absence of estrogens, whereas estrogen administration induced Pten expression in the basal cells. This suggests that Pten acts to prevent excessive cell proliferation as in the case of other squamous tissues. Thus, Pten exhibits a dual role on the control of vaginal homeostasis, depending on whether estrogens are present or absent. Our results provide new insights into how Pten functions in tissue homeostasis.

Self-regulating control of parasitic loads in a fuel cell power system

NASA Technical Reports Server (NTRS)

Vasquez, Arturo (Inventor)

2011-01-01

A fuel cell power system comprises an internal or self-regulating control of a system or device requiring a parasitic load. The internal or self-regulating control utilizes certain components and an interconnection scheme to produce a desirable, variable voltage potential (i.e., power) to a system or device requiring parasitic load in response to varying operating conditions or requirements of an external load that is connected to a primary fuel cell stack of the system. Other embodiments comprise a method of designing such a self-regulated control scheme and a method of operating such a fuel cell power system.

Stem cells from human exfoliated deciduous teeth (SHED) enhance wound healing and the possibility of novel cell therapy.

PubMed

Nishino, Yudai; Yamada, Yoichi; Ebisawa, Katsumi; Nakamura, Sayaka; Okabe, Kazuto; Umemura, Eri; Hara, Kenji; Ueda, Minoru

2011-05-01

In recent years, stem cells from human exfoliated deciduous teeth (SHED) have received attention as a novel stem cell source with multipotent potential. We examined the effect on wound-healing promotion with unique stem cells from deciduous teeth as a medical waste. An excisional wound-splinting mouse model was used and the effect of wound healing among SHED, human mesenchymal stromal cells (hMSCs), human fibroblasts (hFibro) and a control (phosphate-buffered saline; PBS) was evaluated by macroscopy, histology and enzyme-linked immunosorbent assay (ELISA), and the expression of hyaluronan (HA), which is related to wound healing, investigated. SHED and hMSCs accelerated wound healing compared with hFibro and the control. There was a statistically significant difference in wound healing area among hFibro, hMSCs and SHED compared with the control after day 5. At days 7 and 14 after cell transplantation, the histologic observation showed that transplanted PKH26-positive cells were surrounded by human HA binding protein, especially in hMSCs and SHED. HA expression volume values were 1558.41 ± 60.33 (control), 2092.75 ± 42.56 (hFibro), 2342.07 ± 188.10 (hMSCs) and 2314.85 ± 164.91 (SHED) ng/mg, respectively, and significantly higher in hMSCs and SHED compared with hFibro and control at days 7 and 14 (P < 0.05). Our results show that SHED hMSCs have similar effects of wound-healing promotion as hFibro and controls. This implies that SHED might offer a unique stem cell resource and the possibility of novel cell therapies for wound healing in the future.

siRNA blocking the RAS signalling pathway and inhibits the growth of oesophageal squamous cell carcinoma in nude mice.

PubMed

Wang, Xinjie; Zheng, Yuling; Fan, Qingxia; Zhang, Xudong; Shi, Yonggang

2014-12-01

The aim of this study was to study RAS-siRNA blocking RAS pathway and suppressing cell growth in human oesophageal squamous cell carcinoma in nude mice. The methods in this study was to construct RAS-siRNA expression vector, establish 40 oesophageal squamous cell carcinoma xenograft animal models and divided them into five groups: control group, siRNA control group, RAS-siRNA group, paclitaxel group and RAS-siRNA and paclitaxel group. We observed tumour growth in nude mice, studied histology by HE staining, tumour growth inhibition by TUNEL assay and detected the RAS, MAPK and cyclin D1 protein expression by immunohistochemistry and western blot. We have obtained the following results: (i) successfully established animal models; (ii) nude mice in each group after treatment inhibited tumour volume was significantly reduced compared with the control group (p < 0.05); (iii) compared with the control group, the number of apoptotic cells were significantly increased in the siRNA control group and the RAS-siRNA group, and the number of apoptosis cells in the paclitaxel and RAS-siRNA group is significantly most than the paclitaxel group and RAS-siRNA group (p < 0.05); and (iv) after treatment, RAS, MAPK and cyclin D1 expression in five groups was decreasing gradually. After adding paclitaxel, the protein expression in the paclitaxel and RAS-siRNA group was significantly lower than that of paclitaxel group, negative control and paclitaxel group (p < 0.05). We therefore conclude that RAS-siRNA can block the RAS signal transduction pathway, reduce the activity of tumour cells, arrest tumour cell cycle, promote apoptosis, inhibit cell proliferation and increase tumour cell sensitivity to chemotherapeutic drugs. Copyright © 2014 John Wiley & Sons, Ltd.

Increased peripheral blood CD4+ T cell responses to deamidated but not to native gliadin in children with coeliac disease

PubMed Central

Lammi, A; Arikoski, P; Vaarala, O; Kinnunen, T; Ilonen, J

2012-01-01

T cell recognition of gliadin from dietary gluten is essential for the pathogenesis of coeliac disease (CD). The aim of the present study was to analyse whether gliadin-specific T cells are detectable in the circulation of children with newly diagnosed coeliac disease by using a sensitive carboxfluorescein diacetate succinimidyl ester (CFSE) dilution method. Peripheral blood CD4+ T cell responses were analysed in 20 children at diagnosis of CD and compared to those in 64 healthy control children carrying the CD-associated human leucocyte antigen (HLA)-DQ2 or -DQ8 alleles. Deamidated gliadin (gTG)-specific T cells were detectable in the peripheral blood of more than half the children with CD (11 of 20, 55%) compared to 15 of 64 (23·4%) of the control children (P = 0·008). Proliferative responses to gTG were also significantly stronger in children with CD than in controls (P = 0·01). In contrast, T cells specific to native gliadin were detectable at comparable frequencies in children with CD (two of 19, 10·5%) and controls (13 of 64, 20·3%). gTG-specific T cells had a memory phenotype more often than those specific to native gliadin in children with CD (P = 0·02), whereas controls had similar percentages of memory cells in both stimulations. Finally, gTG-specific CD4+ T cells had a higher expression of the gut-homing molecule β7 integrin than those specific to the control antigen tetanus toxoid. Collectively, our current results demonstrate that the frequency of circulating memory CD4+ T cells specific to gTG but not native gliadin is increased in children with newly diagnosed CD. PMID:22471282

Increased peripheral blood CD4+ T cell responses to deamidated but not to native gliadin in children with coeliac disease.

PubMed

Lammi, A; Arikoski, P; Vaarala, O; Kinnunen, T; Ilonen, J

2012-05-01

T cell recognition of gliadin from dietary gluten is essential for the pathogenesis of coeliac disease (CD). The aim of the present study was to analyse whether gliadin-specific T cells are detectable in the circulation of children with newly diagnosed coeliac disease by using a sensitive carboxfluorescein diacetate succinimidyl ester (CFSE) dilution method. Peripheral blood CD4(+) T cell responses were analysed in 20 children at diagnosis of CD and compared to those in 64 healthy control children carrying the CD-associated human leucocyte antigen (HLA)-DQ2 or -DQ8 alleles. Deamidated gliadin (gTG)-specific T cells were detectable in the peripheral blood of more than half the children with CD (11 of 20, 55%) compared to 15 of 64 (23.4%) of the control children (P = 0.008). Proliferative responses to gTG were also significantly stronger in children with CD than in controls (P = 0.01). In contrast, T cells specific to native gliadin were detectable at comparable frequencies in children with CD (two of 19, 10.5%) and controls (13 of 64, 20.3%). gTG-specific T cells had a memory phenotype more often than those specific to native gliadin in children with CD (P = 0.02), whereas controls had similar percentages of memory cells in both stimulations. Finally, gTG-specific CD4(+) T cells had a higher expression of the gut-homing molecule β7 integrin than those specific to the control antigen tetanus toxoid. Collectively, our current results demonstrate that the frequency of circulating memory CD4(+) T cells specific to gTG but not native gliadin is increased in children with newly diagnosed CD. © 2012 The Authors;Clinical and Experimental Immunology © 2012 British Society for Immunology.

Efficient and reproducible mammalian cell bioprocesses without probes and controllers?

PubMed

Tissot, Stéphanie; Oberbek, Agata; Reclari, Martino; Dreyer, Matthieu; Hacker, David L; Baldi, Lucia; Farhat, Mohamed; Wurm, Florian M

2011-07-01

Bioprocesses for recombinant protein production with mammalian cells are typically controlled for several physicochemical parameters including the pH and dissolved oxygen concentration (DO) of the culture medium. Here we studied whether these controls are necessary for efficient and reproducible bioprocesses in an orbitally shaken bioreactor (OSR). Mixing, gas transfer, and volumetric power consumption (P(V)) were determined in both a 5-L OSR and a 3-L stirred-tank bioreactor (STR). The two cultivation systems had a similar mixing intensity, but the STR had a lower volumetric mass transfer coefficient of oxygen (k(L)a) and a higher P(V) than the OSR. Recombinant CHO cell lines expressing either tumor necrosis factor receptor as an Fc fusion protein (TNFR:Fc) or an anti-RhesusD monoclonal antibody were cultivated in the two systems. The 5-L OSR was operated in an incubator shaker with 5% CO(2) in the gas environment but without pH and DO control whereas the STR was operated with or without pH and DO control. Higher cell densities and recombinant protein titers were obtained in the OSR as compared to both the controlled and the non-controlled STRs. To test the reproducibility of a bioprocess in a non-controlled OSR, the two CHO cell lines were each cultivated in parallel in six 5-L OSRs. Similar cell densities, cell viabilities, and recombinant protein titers along with similar pH and DO profiles were achieved in each group of replicates. Our study demonstrated that bioprocesses can be performed in OSRs without pH or DO control in a highly reproducible manner, at least at the scale of operation studied here. Copyright © 2011 Elsevier B.V. All rights reserved.

Normal T-cell activation in elite controllers with preserved CD4+ T-cell counts.

PubMed

Bansal, Anju; Sterrett, Sarah; Erdmann, Nathan; Westfall, Andrew O; Dionne-Odom, Jodie; Overton, Edgar T; Goepfert, Paul A

2015-11-01

HIV elite controllers suppress HIV viremia without antiretroviral therapy (ART), yet previous studies demonstrated that elite controllers maintain an activated T-cell phenotype. Chronic immune activation has detrimental consequences and thus ART has been advocated for all elite controllers. However, elite controllers are not a clinically homogenous group. Since CD4% is among the best predictors of AIDS-related events, in the current study, we assessed whether this marker can be used to stratify elite controllers needing ART. Sixteen elite controllers were divided into two groups based on CD4% (EC > 40% and EC ≤40%), and T-cell subsets were analyzed for markers of memory/differentiation (CD45RA, CCR7, CD28), activation (CD38/HLA-DR), immunosenescence (CD57), costimulation (CD73, CD28) and exhaustion (PD-1, CD160, Tim-3). Monocyte subsets (CD14, CD16) were also analyzed and sCD14 levels were quantified using ELISA. In the EC group, expression of activation, exhaustion, and immunosensescence markers on T cells were significantly reduced compared with the EC group and similar to the seronegative controls. The EC group expressed higher levels of costimulatory molecules CD28 and CD73 and had lower levels of monocyte activation (HLA-DR expression) with a reduced frequency of inflammatory monocyte (CD14 CD16) subset. Furthermore, the EC group maintained a stable CD4% during a median follow-up of 6 years. Elite controllers with preserved CD4T cells (EC) have normal T-cell and monocyte phenotypes and therefore may have limited benefit from ART. CD4% can be an important marker for evaluating future studies aimed at determining the need for ART in this group of individuals.

Cell-free mitochondrial DNA copy number variation in head and neck squamous cell carcinoma: A study of non-invasive biomarker from Northeast India.

PubMed

Kumar, Manish; Srivastava, Shilpee; Singh, Seram Anil; Das, Anup Kumar; Das, Ganesh Chandra; Dhar, Bishal; Ghosh, Sankar Kumar; Mondal, Rosy

2017-10-01

Head and neck squamous cell carcinoma is the most commonly diagnosed cancer worldwide. The lifestyle, food habits, and customary practices manifest the Northeast Indian population toward higher susceptibility to develop head and neck squamous cell carcinoma. Here, we have investigated the association of smoke and smokeless tobacco, and alcohol with copy number variation of cell-free mitochondrial DNA and cell-free nuclear DNA in cases and controls. Cell-free DNA from plasma was isolated from 50 head and neck squamous cell carcinoma cases and 50 controls with informed written consent using QIAamp Circulating Nucleic Acid Kit. Real-time polymerase chain reaction was done for copy number variation in cell-free mitochondrial DNA and cell-free nuclear DNA. Receiver operating characteristic curve analysis was performed to evaluate the diagnostic application between the two study groups using clinicopathological parameters. The levels of cell-free nuclear DNA and cell-free mitochondrial DNA of cases in association with smoke and smokeless tobacco, alcohol with smoking (p < 0.05) were significantly higher (p < 0.01 and p < 0.001, respectively) than controls. Using receiver operating characteristic curve analysis between head and neck squamous cell carcinoma cases and controls, we distinguished cell-free mitochondrial DNA (cutoff: 19.84 raw Ct; sensitivity: 84%; specificity: 100%; p < 0.001) and cell-free nuclear DNA (cutoff: 463,282 genomic equivalent/mL; sensitivity: 53%; specificity: 87%; p < 0.001). The copy number variation in cases (cell-free nuclear DNA: 5451.66 genomic equivalent/mL and cell-free mitochondrial DNA: 29,103,476.15 genomic equivalent/mL) and controls (cell-free nuclear DNA: 1650.9 genomic equivalent/mL and cell-free mitochondrial DNA: 9,189,312.54 genomic equivalent/mL), respectively. Our result indicates that the cell-free mitochondrial DNA content is highly associated with smoke and smokeless tobacco, betel quid chewing, and alcohol which shows greater promises, holding the key characteristics of diagnostic biomarkers, that is, minimal invasiveness, high specificity, and sensitivity.

Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung.

PubMed

Lange, Alexander W; Sridharan, Anusha; Xu, Yan; Stripp, Barry R; Perl, Anne-Karina; Whitsett, Jeffrey A

2015-02-01

The Hippo/Yap pathway is a well-conserved signaling cascade that regulates cell proliferation and differentiation to control organ size and stem/progenitor cell behavior. Following airway injury, Yap was dynamically regulated in regenerating airway epithelial cells. To determine the role of Hippo signaling in the lung, the mammalian Hippo kinases, Mst1 and Mst2, were deleted in epithelial cells of the embryonic and mature mouse lung. Mst1/2 deletion in the fetal lung enhanced proliferation and inhibited sacculation and epithelial cell differentiation. The transcriptional inhibition of cell proliferation and activation of differentiation during normal perinatal lung maturation were inversely regulated following embryonic Mst1/2 deletion. Ablation of Mst1/2 from bronchiolar epithelial cells in the adult lung caused airway hyperplasia and altered differentiation. Inhibitory Yap phosphorylation was decreased and Yap nuclear localization and transcriptional targets were increased after Mst1/2 deletion, consistent with canonical Hippo/Yap signaling. YAP potentiated cell proliferation and inhibited differentiation of human bronchial epithelial cells in vitro. Loss of Mst1/2 and expression of YAP regulated transcriptional targets controlling cell proliferation and differentiation, including Ajuba LIM protein. Ajuba was required for the effects of YAP on cell proliferation in vitro. Hippo/Yap signaling regulates Ajuba and controls proliferation and differentiation of lung epithelial progenitor cells. © The Author (2014). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Spatial control of active CDC-42 during collective migration of hypodermal cells in Caenorhabditis elegans.

PubMed

Ouellette, Marie-Hélène; Martin, Emmanuel; Lacoste-Caron, Germain; Hamiche, Karim; Jenna, Sarah

2016-08-01

Collective epithelial cell migration requires the maintenance of cell-cell junctions while enabling the generation of actin-rich protrusions at the leading edge of migrating cells. Ventral enclosure of Caenorhabditis elegans embryos depends on the collective migration of anterior-positioned leading hypodermal cells towards the ventral midline where they form new junctions with their contralateral neighbours. In this study, we characterized the zygotic function of RGA-7/SPV-1, a CDC-42/Cdc42 and RHO-1/RhoA-specific Rho GTPase-activating protein, which controls the formation of actin-rich protrusions at the leading edge of leading hypodermal cells and the formation of new junctions between contralateral cells. We show that RGA-7 controls these processes in an antagonistic manner with the CDC-42's effector WSP-1/N-WASP and the CDC-42-binding proteins TOCA-1/2/TOCA1. RGA-7 is recruited to spatially distinct locations at junctions between adjacent leading cells, where it promotes the accumulation of clusters of activated CDC-42. It also inhibits the spreading of these clusters towards the leading edge of the junctions and regulates their accumulation and distribution at new junctions formed between contralateral leading cells. Our study suggests that RGA-7 controls collective migration and junction formation between epithelial cells by spatially restricting active CDC-42 within cell-cell junctions. © The Author (2015). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Twenty Four-Hour Exposure to a 0.12 THz Electromagnetic Field Does Not Affect the Genotoxicity, Morphological Changes, or Expression of Heat Shock Protein in HCE-T Cells.

PubMed

Koyama, Shin; Narita, Eijiro; Shimizu, Yoko; Shiina, Takeo; Taki, Masao; Shinohara, Naoki; Miyakoshi, Junji

2016-08-05

To investigate the cellular effects of terahertz (THz) exposure, human corneal epithelial (HCE-T) cells derived from human eye were exposed to 0.12 THz radiation at 5 mW/cm² for 24 h, then the genotoxicity, morphological changes, and heat shock protein (Hsp) expression of the cells were examined. There was no statistically significant increase in the micronucleus (MN) frequency of cells exposed to 0.12 THz radiation compared with sham-exposed controls and incubator controls, whereas the MN frequency of cells treated with bleomycin for 1 h (positive control) did increase significantly. Similarly, there were no significant morphological changes in cells exposed to 0.12 THz radiation compared to sham-exposed controls and incubator controls, and Hsp expression (Hsp27, Hsp70, and Hsp90α) was also not significantly different between the three treatments. These results indicate that exposure to 0.12 THz radiation using the present conditions appears to have no or very little effect on MN formation, morphological changes, and Hsp expression in cells derived from human eye.

Automatic Control of Gene Expression in Mammalian Cells.

PubMed

Fracassi, Chiara; Postiglione, Lorena; Fiore, Gianfranco; di Bernardo, Diego

2016-04-15

Automatic control of gene expression in living cells is paramount importance to characterize both endogenous gene regulatory networks and synthetic circuits. In addition, such a technology can be used to maintain the expression of synthetic circuit components in an optimal range in order to ensure reliable performance. Here we present a microfluidics-based method to automatically control gene expression from the tetracycline-inducible promoter in mammalian cells in real time. Our approach is based on the negative-feedback control engineering paradigm. We validated our method in a monoclonal population of cells constitutively expressing a fluorescent reporter protein (d2EYFP) downstream of a minimal CMV promoter with seven tet-responsive operator motifs (CMV-TET). These cells also constitutively express the tetracycline transactivator protein (tTA). In cells grown in standard growth medium, tTA is able to bind the CMV-TET promoter, causing d2EYFP to be maximally expressed. Upon addition of tetracycline to the culture medium, tTA detaches from the CMV-TET promoter, thus preventing d2EYFP expression. We tested two different model-independent control algorithms (relay and proportional-integral (PI)) to force a monoclonal population of cells to express an intermediate level of d2EYFP equal to 50% of its maximum expression level for up to 3500 min. The control input is either tetracycline-rich or standard growth medium. We demonstrated that both the relay and PI controllers can regulate gene expression at the desired level, despite oscillations (dampened in the case of the PI controller) around the chosen set point.

Regulatory T, natural killer T and γδ T cells in multiple sclerosis and chronic fatigue syndrome/myalgic encephalomyelitis: a comparison.

PubMed

Ramos, Sandra; Brenu, Ekua; Broadley, Simon; Kwiatek, Richard; Ng, Jennifer; Nguyen, Thao; Freeman, Susan; Staines, Donald; Marshall-Gradisnik, Sonya

2016-12-01

Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME), and Multiple Sclerosis (MS) may share some similarities in relation to reduced NK cell activity. It is likely that other cells such as regulatory T (Tregs), invariant Natural Killer T (iNKT) and gamma delta T (γδ T) cells may also be dysregulated in CFS/ME and MS. To evaluate and compare specific immune regulatory cells of patients with CFS/ME, patients with MS and healthy controls. Sixty three volunteers were included in this study: 24 were CFS/ME patients, 11 were MS patients and 27 were healthy controls. Blood samples were obtained from all participants for flow cytometry analysis of iNKT cells, Tregs and γδ T cell phenotypes. We observed a significant increase in Tregs in the CFS/ME group (p≤0.05) compared to the healthy control group. Total γδ and γδ2 T cells were significantly reduced in MS patients in comparison with the healthy control group. Conversely, CD4+iNKT percentage of iNKT, was significantly increased in the CFS/ME group compared with healthy controls and the double-negative iNKT percentage of iNKT significantly decreased compared with the healthy control group. This study has not identified any immunological disturbances that are common in both MS and CFS/ME patients. However, the differential expression of cell types between the conditions investigated suggests different pathways of disease. These differences need to be explored in further studies.

Force-controlled manipulation of single cells: from AFM to FluidFM.

PubMed

Guillaume-Gentil, Orane; Potthoff, Eva; Ossola, Dario; Franz, Clemens M; Zambelli, Tomaso; Vorholt, Julia A

2014-07-01

The ability to perturb individual cells and to obtain information at the single-cell level is of central importance for addressing numerous biological questions. Atomic force microscopy (AFM) offers great potential for this prospering field. Traditionally used as an imaging tool, more recent developments have extended the variety of cell-manipulation protocols. Fluidic force microscopy (FluidFM) combines AFM with microfluidics via microchanneled cantilevers with nano-sized apertures. The crucial element of the technology is the connection of the hollow cantilevers to a pressure controller, allowing their operation in liquid as force-controlled nanopipettes under optical control. Proof-of-concept studies demonstrated a broad spectrum of single-cell applications including isolation, deposition, adhesion and injection in a range of biological systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

Utilization of photoinduced charge-separated state of donor-acceptor-linked molecules for regulation of cell membrane potential and ion transport.

PubMed

Numata, Tomohiro; Murakami, Tatsuya; Kawashima, Fumiaki; Morone, Nobuhiro; Heuser, John E; Takano, Yuta; Ohkubo, Kei; Fukuzumi, Shunichi; Mori, Yasuo; Imahori, Hiroshi

2012-04-11

The control of ion transport across cell membranes by light is an attractive strategy that allows targeted, fast control of precisely defined events in the biological membrane. Here we report a novel general strategy for the control of membrane potential and ion transport by using charge-separation molecules and light. Delivery of charge-separation molecules to the plasma membrane of PC12 cells by a membranous nanocarrier and subsequent light irradiation led to depolarization of the membrane potential as well as inhibition of the potassium ion flow across the membrane. Photoregulation of the cell membrane potential and ion transport by using charge-separation molecules is highly promising for control of cell functions. © 2012 American Chemical Society

MreB Orientation Correlates with Cell Diameter in Escherichia coli.

PubMed

Ouzounov, Nikolay; Nguyen, Jeffrey P; Bratton, Benjamin P; Jacobowitz, David; Gitai, Zemer; Shaevitz, Joshua W

2016-09-06

Bacteria have remarkably robust cell shape control mechanisms. For example, cell diameter only varies by a few percent across a given population. The bacterial actin homolog, MreB, is necessary for establishment and maintenance of rod shape although the detailed properties of MreB that are important for shape control remained unknown. In this study, we perturb MreB in two ways: by treating cells with the polymerization-inhibiting drug A22 and by creating point mutants in mreB. These perturbations modify the steady-state diameter of cells over a wide range, from 790 ± 30 nm to 1700 ± 20 nm. To determine which properties of MreB are important for diameter control, we correlated structural characteristics of fluorescently tagged MreB polymers with cell diameter by simultaneously analyzing three-dimensional images of MreB and cell shape. Our results indicate that the helical pitch angle of MreB inversely correlates with the cell diameter of Escherichia coli. Other correlations between MreB and cell diameter are not found to be significant. These results demonstrate that the physical properties of MreB filaments are important for shape control and support a model in which MreB organizes the cell wall growth machinery to produce a chiral cell wall structure and dictate cell diameter. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Feasibility of mesenchymal stem cell culture expansion for a phase I clinical trial in multiple sclerosis.

PubMed

Planchon, Sarah M; Lingas, Karen T; Reese Koç, Jane; Hooper, Brittney M; Maitra, Basabi; Fox, Robert M; Imrey, Peter B; Drake, Kylie M; Aldred, Micheala A; Lazarus, Hillard M; Cohen, Jeffrey A

2018-01-01

Multiple sclerosis is an inflammatory, neurodegenerative disease of the central nervous system for which therapeutic mesenchymal stem cell transplantation is under study. Published experience of culture-expanding multiple sclerosis patients' mesenchymal stem cells for clinical trials is limited. To determine the feasibility of culture-expanding multiple sclerosis patients' mesenchymal stem cells for clinical use. In a phase I trial, autologous, bone marrow-derived mesenchymal stem cells were isolated from 25 trial participants with multiple sclerosis and eight matched controls, and culture-expanded to a target single dose of 1-2 × 10 6 cells/kg. Viability, cell product identity and sterility were assessed prior to infusion. Cytogenetic stability was assessed by single nucleotide polymorphism analysis of mesenchymal stem cells from 18 multiple sclerosis patients and five controls. One patient failed screening. Mesenchymal stem cell culture expansion was successful for 24 of 25 multiple sclerosis patients and six of eight controls. The target dose was achieved in 16-62 days, requiring two to three cell passages. Growth rate and culture success did not correlate with demographic or multiple sclerosis disease characteristics. Cytogenetic studies identified changes on one chromosome of one control (4.3%) after extended time in culture. Culture expansion of mesenchymal stem cells from multiple sclerosis patients as donors is feasible. However, culture time should be minimized for cell products designated for therapeutic administration.

Association between memory B-cells and clinical and immunological features of primary Sjögren's syndrome and Sicca patients.

PubMed

Barcelos, Filipe; Martins, Catarina; Papoila, Ana; Geraldes, Carlos; Cardigos, Joana; Nunes, Glória; Lopes, Teresa; Alves, Nuno; Vaz-Patto, José; Branco, Jaime; Borrego, Luís-Miguel

2018-06-01

B-cells play a pivotal role in primary Sjögren's syndrome (pSS) pathogenesis. We aim to (1) evaluate the distribution of B-lymphocyte subpopulations in pSS and Sicca patients, (2) establish cut-off points that discriminate pSS from controls, (3) evaluate the association between memory B-cells and phenotypic features in pSS. We included 57 pSS patients, 68 Sicca and 24 healthy controls. Circulating B-cells were characterized by flow cytometry as naïve and memory subsets and classified from Bm1 to Bm5. Compared to controls, pSS patients had lower percentages (29.5 vs 44.4%) and absolute numbers (47 vs 106 cells/µl) of memory B-cells. Through ROC curves, a cut-off of ≤ 58 total memory B-cells/µl yielded a specificity of 0.88 and a sensitivity of 0.60 for pSS, and was met by 59.6% of pSS patients, 38.8% of Sicca and 12.5% of controls. A cut-off of < 23.5 Switched-memory B-cells/µl yielded a specificity of 0.88 and a sensitivity of 0.54 and was met by 54.4% of pSS patients, 37.3% of Sicca and 12.5% of controls. In pSS, lower total memory B-cells count was associated with longer disease duration (14.3 vs 8.1 years, p = 0.006) and more active disease profile, as evaluated by the European League Against Rheumatism (EULAR) Sjögren's Syndrome Disease Activity Index (ESSDAI) (3.1 vs 1.4, p = 0.043). Decreased numbers of memory B-cells clearly discriminated pSS from controls and can also have prognostic value. It remains to be clarified whether Sicca patients with decreased memory B-cells represent pSS and if B-cell profiling could help in the diagnosis of pSS.

MicroRNAs: key regulators of stem cells.

PubMed

Gangaraju, Vamsi K; Lin, Haifan

2009-02-01

The hallmark of a stem cell is its ability to self-renew and to produce numerous differentiated cells. This unique property is controlled by dynamic interplays between extrinsic signalling, epigenetic, transcriptional and post-transcriptional regulations. Recent research indicates that microRNAs (miRNAs) have an important role in regulating stem cell self-renewal and differentiation by repressing the translation of selected mRNAs in stem cells and differentiating daughter cells. Such a role has been shown in embryonic stem cells, germline stem cells and various somatic tissue stem cells. These findings reveal a new dimension of gene regulation in controlling stem cell fate and behaviour.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

PubMed Central

Baltar, Federico

2018-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles. PMID:29354095

Gestational diabetes mellitus alters maternal and neonatal circulating endothelial progenitor cell subsets.

PubMed

Acosta, Juan C; Haas, David M; Saha, Chandan K; Dimeglio, Linda A; Ingram, David A; Haneline, Laura S

2011-03-01

The purpose of this study was to examine whether women with gestational diabetes mellitus (GDM) and their offspring have reduced endothelial progenitor cell subsets and vascular reactivity. Women with GDM, healthy control subjects, and their infants participated. Maternal blood and cord blood were assessed for colony-forming unit-endothelial cells and endothelial progenitor cell subsets with the use of polychromatic flow cytometry. Cord blood endothelial colony-forming cells were enumerated. Vascular reactivity was tested by laser Doppler imaging. Women with GDM had fewer CD34, CD133, CD45, and CD31 cells (circulating progenitor cells [CPCs]) at 24-32 weeks' gestation and 1-2 days after delivery, compared with control subjects. No differences were detected in colony-forming unit-endothelial cells or colony-forming unit-endothelial cells. In control subjects, CPCs were higher in the third trimester, compared with the postpartum period. Cord blood from GDM pregnancies had reduced CPCs. Vascular reactivity was not different between GDM and control subjects. The normal physiologic increase in CPCs during pregnancy is impaired in women with GDM, which may contribute to endothelial dysfunction and GDM-associated morbidities. Copyright © 2011 Mosby, Inc. All rights reserved.

An integrated system for synchronous culture of animal cells under controlled conditions.

PubMed

Mendoza-Pérez, Elena; Hernández, Vanessa; Palomares, Laura A; Serrato, José A

2016-01-01

The cell cycle has fundamental effects on cell cultures and their products. Tools to synchronize cultured cells allow the study of cellular physiology and metabolism at particular cell cycle phases. However, cells are most often arrested by methods that alter their homeostasis and are then cultivated in poorly controlled environments. Cell behavior could then be affected by the synchronization method and culture conditions used, and not just by the particular cell cycle phase under study. Moreover, only a few viable cells are recovered. Here, we designed an integrated system where a large number of cells from a controlled bioreactor culture is separated by centrifugal elutriation at high viabilities. In contrast to current elutriation methods, cells are injected directly from a bioreactor into an injection loop, allowing the introduction of a large number of cells into the separation chamber without stressful centrifugation. A low pulsation peristaltic pump increases the stability of the elutriation chamber. Using this approach, a large number of healthy cells at each cell cycle phase were obtained, allowing their direct inoculation into fully instrumented bioreactors. Hybridoma cells synchronized and cultured in this system behaved as expected for a synchronous culture.

Rac1/RhoA antagonism defines cell-to-cell heterogeneity during epidermal morphogenesis in nematodes

PubMed Central

Ouellette, Marie-Hélène

2016-01-01

The antagonism between the GTPases Rac1 and RhoA controls cell-to-cell heterogeneity in isogenic populations of cells in vitro and epithelial morphogenesis in vivo. Its involvement in the regulation of cell-to-cell heterogeneity during epidermal morphogenesis has, however, never been addressed. We used a quantitative cell imaging approach to characterize epidermal morphogenesis at a single-cell level during early elongation of Caenorhabditis elegans embryos. This study reveals that a Rac1-like pathway, involving the Rac/Cdc42 guanine-exchange factor β-PIX/PIX-1 and effector PAK1/PAK-1, and a RhoA-like pathway, involving ROCK/LET-502, control the remodeling of apical junctions and the formation of basolateral protrusions in distinct subsets of hypodermal cells. In these contexts, protrusions adopt lamellipodia or an amoeboid morphology. We propose that lamella formation may reduce tension building at cell–cell junctions during morphogenesis. Cell-autonomous antagonism between these pathways enables cells to switch between Rac1- and RhoA-like morphogenetic programs. This study identifies the first case of cell-to-cell heterogeneity controlled by Rac1/RhoA antagonism during epidermal morphogenesis. PMID:27821782

Control of autothermal reforming reactor of diesel fuel

NASA Astrophysics Data System (ADS)

Dolanc, Gregor; Pregelj, Boštjan; Petrovčič, Janko; Pasel, Joachim; Kolb, Gunther

2016-05-01

In this paper a control system for autothermal reforming reactor for diesel fuel is presented. Autothermal reforming reactors and the pertaining purification reactors are used to convert diesel fuel into hydrogen-rich reformate gas, which is then converted into electricity by the fuel cell. The purpose of the presented control system is to control the hydrogen production rate and the temperature of the autothermal reforming reactor. The system is designed in such a way that the two control loops do not interact, which is required for stable operation of the fuel cell. The presented control system is a part of the complete control system of the diesel fuel cell auxiliary power unit (APU).

HIV enteropathy: crypt stem and transit cell hyperproliferation induces villous atrophy in HIV/Microsporidia-infected jejunal mucosa.

PubMed

Batman, Philip A; Kotler, Donald P; Kapembwa, Moses S; Booth, Dawn; Potten, Christopher S; Orenstein, Jan M; Scally, Andrew J; Griffin, George E

2007-02-19

The study aim was to analyse the kinetics of stem and transit cells in the crypts of jejunal mucosa infected with HIV and Microsporidia. The size of villi, depth of crypts and proliferative activity of transit and stem cells in jejunal mucosa were measured using morphometric techniques. The surface area/volume ratio (S/V) of jejunal biopsies was estimated under light microscopy using a Weibel graticule. Crypt length was measured by counting enterocytes along the crypt side from the base to the villus junction, and the mean crypt length was calculated. The S/V and crypt lengths of the jejunal mucosa of 21 HIV and Microsporidia-infected test cases were compared with 14 control cases. The labelling index in relation to the crypt cell position of 10 of the test cases was analysed compared with 13 control cases. Differences were found in the S/V and crypt length, and there was a negative correlation between S/V and crypt length in test and control cases combined. Cell labelling indices fell into low and high proliferation groups. There were significant differences in labelling indices between low proliferation test cases and controls, between high proliferation test cases and controls, and between high and low proliferation test cases. Villous atrophy induced by HIV and Microsporidia is attributed to crypt cell hyperplasia and the encroachment of crypt cells onto villi. These infections induce crypt hypertrophy by stimulating cell mitosis predominantly in transit cells but also in stem cells. Increased stem cell proliferation occurs only in high proliferation cases.

MHC class II molecules control murine B cell responsiveness to lipopolysaccharide stimulation.

PubMed

Rodo, Joana; Gonçalves, Lígia A; Demengeot, Jocelyne; Coutinho, António; Penha-Gonçalves, Carlos

2006-10-01

LPS is a strong stimulator of the innate immune system and inducer of B lymphocyte activation. Two TLRs, TLR4 and RP105 (CD180), have been identified as mediators of LPS signaling in murine B cells, but little is known about genetic factors that are able to control LPS-induced cell activation. We performed a mouse genome-wide screen that aside from identifying a controlling locus mapping in the TLR4 region (logarithm of odds score, 2.77), also revealed that a locus closely linked to the MHC region (logarithm of odds score, 3.4) governed B cell responsiveness to LPS stimulation. Using purified B cells obtained from MHC congenic strains, we demonstrated that the MHC(b) haplotype is accountable for higher cell activation, cell proliferation, and IgM secretion, after LPS stimulation, when compared with the MHC(d) haplotype. Furthermore, B cells from MHC class II(-/-) mice displayed enhanced activation and proliferation in response to LPS. In addition, we showed that the MHC haplotype partially controls expression of RP105 (a LPS receptor molecule), following a pattern that resembles the LPS responsiveness phenotype. Together, our results strongly suggest that murine MHC class II molecules play a role in constraining the B cell response to LPS and that genetic variation at the MHC locus is an important component in controlling B cell responsiveness to LPS stimulation. This work raises the possibility that constraining of B cell responsiveness by MHC class II molecules may represent a functional interaction between adaptive and innate immune systems.

The duration of mitosis and daughter cell size are modulated by nutrients in budding yeast.

PubMed

Leitao, Ricardo M; Kellogg, Douglas R

2017-11-06

The size of nearly all cells is modulated by nutrients. Thus, cells growing in poor nutrients can be nearly half the size of cells in rich nutrients. In budding yeast, cell size is thought to be controlled almost entirely by a mechanism that delays cell cycle entry until sufficient growth has occurred in G1 phase. Here, we show that most growth of a new daughter cell occurs in mitosis. When the rate of growth is slowed by poor nutrients, the duration of mitosis is increased, which suggests that cells compensate for slow growth in mitosis by increasing the duration of growth. The amount of growth required to complete mitosis is reduced in poor nutrients, leading to a large reduction in cell size. Together, these observations suggest that mechanisms that control the extent of growth in mitosis play a major role in cell size control in budding yeast. © 2017 Leitao and Kellogg.

Crawling and turning in a minimal reaction-diffusion cell motility model: Coupling cell shape and biochemistry

NASA Astrophysics Data System (ADS)

Camley, Brian A.; Zhao, Yanxiang; Li, Bo; Levine, Herbert; Rappel, Wouter-Jan

2017-01-01

We study a minimal model of a crawling eukaryotic cell with a chemical polarity controlled by a reaction-diffusion mechanism describing Rho GTPase dynamics. The size, shape, and speed of the cell emerge from the combination of the chemical polarity, which controls the locations where actin polymerization occurs, and the physical properties of the cell, including its membrane tension. We find in our model both highly persistent trajectories, in which the cell crawls in a straight line, and turning trajectories, where the cell transitions from crawling in a line to crawling in a circle. We discuss the controlling variables for this turning instability and argue that turning arises from a coupling between the reaction-diffusion mechanism and the shape of the cell. This emphasizes the surprising features that can arise from simple links between cell mechanics and biochemistry. Our results suggest that similar instabilities may be present in a broad class of biochemical descriptions of cell polarity.

Transcription Factor Foxo1 Is a Negative Regulator of NK Cell Maturation and Function

PubMed Central

Deng, Youcai; Kerdiles, Yann; Chu, Jianhong; Yuan, Shunzong; Wang, Youwei; Chen, Xilin; Mao, Hsiaoyin; Zhang, Lingling; Zhang, Jianying; Hughes, Tiffany; Deng, Yafei; Zhang, Qi; Wang, Fangjie; Zou, Xianghong; Liu, Chang-Gong; Freud, Aharon G.; Li, Xiaohui; Caligiuri, Michael A; Vivier, Eric; Yu, Jianhua

2015-01-01

SUMMARY Little is known about the role of negative regulators in controlling natural killer (NK) cell development and effector functions. Foxo1 is a multifunctional transcription factor of the forkhead family. Using a mouse model of conditional deletion in NK cells, we found that Foxo1 negatively controlled NK cell differentiation and function. Immature NK cells expressed abundant Foxo1 and little Tbx21 relative to mature NK cells, but these two transcription factors reversed their expression as NK cells proceeded through development. Foxo1 promoted NK cell homing to lymph nodes through upregulating CD62L expression, and impaired late-stage maturation and effector functions by repressing Tbx21 expression. Loss of Foxo1 rescued the defect in late-stage NK cell maturation in heterozygous Tbx21+/− mice. Collectively, our data reveal a regulatory pathway by which the negative regulator Foxo1 and the positive regulator Tbx21 play opposing roles in controlling NK cell development and effector functions. PMID:25769609

Qualitative features of the HIV-specific CD8+ T-cell response associated with immunologic control.

PubMed

Hersperger, Adam R; Migueles, Stephen A; Betts, Michael R; Connors, Mark

2011-05-01

Over the past 2 years, a clearer picture has emerged regarding the properties of HIV-specific CD8+ T cells associated with immunologic control of HIV replication. These properties represent a potential mechanism by which rare patients might control HIV replication in the absence of antiretroviral therapy. This review addresses the background and recent findings that have lead to our current understanding of these mechanism(s). Patients with immunologic control of HIV are not distinguished by targeted specificities, or greater numbers or breadth of their HIV-specific CD8+ T-cell response. For this reason, recent work has focused greater attention on qualitative features of this response. The qualitative features most closely associated with immunologic control of HIV are related to the granule-exocytosis-mediated elimination of HIV-infected CD4 T cells. The ability of HIV-specific CD8+ T cells to increase their contents of proteins known to mediate cytotoxicity, such as granzyme B and perforin, appears to be a critical means by which HIV-specific cytotoxic capacity is regulated. Investigation from multiple groups has now focused upon HIV-specific CD8+ T-cell granule-exocytosis-mediated cytotoxicity as a correlate of immunologic control of HIV. In the near future, a more detailed understanding of the qualities associated with immunologic control may provide critical insights regarding the necessary features of a response that should be stimulated by immunotherapies or T-cell-based vaccines.

The putative oncotarget CSN5 controls a transcription-uncorrelated p53-mediated autophagy implicated in cancer cell survival under curcumin treatment

PubMed Central

Sheng, Ji-Po; Yu, Fang; Tan, Ren-Xiang; Pan, Ying; Huang, Jun-Jian; Kong, Ling-Dong

2016-01-01

Curcumin has shown promise as a safe and specific anticancer agent. The COP9 signalosome (CSN) component CSN5, a known specific target for curcumin, can control p53 stability by increasing its degradation through ubiquitin system. But the correlation of CSN5-controlled p53 to anticancer therapeutic effect of curcumin is currently unknown. Here we showed that CSN5-controlled p53 was transcriptional inactive and responsible for autophagy in human normal BJ cells and cancer HepG2 cells under curcumin treatment. Of note, CSN5-initiated cellular autophagy by curcumin treatment was abolished in p53-null HCT116p53−/− cancer cells, which could be rescued by reconstitution with wild-type p53 or transcription inactive p53 mutant p53R273H. Furthermore, CSN5-controlled p53 conferred a pro-survival autophagy in diverse cancer cells response to curcumin. Genetic p53 deletion, as well as autophagy pharmacological inhibition by chloroquine, significantly enhanced the therapeutic effect of curcumin on cancer cells in vitro and in vivo, but not normal cells. This study identifies a novel CSN5-controlled p53 in autophagy of human cells. The p53 expression state is a useful biomarker for predicting the anticancer therapeutic effect of curcumin. Therefore, the pharmacologic autophagy manipulation may benefit the ongoing anticancer clinical trials of curcumin. PMID:27626169

Patterned cortical tension mediated by N-cadherin controls cell geometric order in the Drosophila eye

PubMed Central

Chan, Eunice HoYee; Chavadimane Shivakumar, Pruthvi; Clément, Raphaël; Laugier, Edith; Lenne, Pierre-François

2017-01-01

Adhesion molecules hold cells together but also couple cell membranes to a contractile actomyosin network, which limits the expansion of cell contacts. Despite their fundamental role in tissue morphogenesis and tissue homeostasis, how adhesion molecules control cell shapes and cell patterns in tissues remains unclear. Here we address this question in vivo using the Drosophila eye. We show that cone cell shapes depend little on adhesion bonds and mostly on contractile forces. However, N-cadherin has an indirect control on cell shape. At homotypic contacts, junctional N-cadherin bonds downregulate Myosin-II contractility. At heterotypic contacts with E-cadherin, unbound N-cadherin induces an asymmetric accumulation of Myosin-II, which leads to a highly contractile cell interface. Such differential regulation of contractility is essential for morphogenesis as loss of N-cadherin disrupts cell rearrangements. Our results establish a quantitative link between adhesion and contractility and reveal an unprecedented role of N-cadherin on cell shapes and cell arrangements. DOI: http://dx.doi.org/10.7554/eLife.22796.001 PMID:28537220

Regulatory iNKT cells lack expression of the transcription factor PLZF and control the homeostasis of T(reg) cells and macrophages in adipose tissue.

PubMed

Lynch, Lydia; Michelet, Xavier; Zhang, Sai; Brennan, Patrick J; Moseman, Ashley; Lester, Chantel; Besra, Gurdyal; Vomhof-Dekrey, Emilie E; Tighe, Mike; Koay, Hui-Fern; Godfrey, Dale I; Leadbetter, Elizabeth A; Sant'Angelo, Derek B; von Andrian, Ulrich; Brenner, Michael B

2015-01-01

Invariant natural killer T cells (iNKT cells) are lipid-sensing innate T cells that are restricted by the antigen-presenting molecule CD1d and express the transcription factor PLZF. iNKT cells accumulate in adipose tissue, where they are anti-inflammatory, but the factors that contribute to their anti-inflammatory nature, as well as their targets in adipose tissue, are unknown. Here we found that iNKT cells in adipose tissue had a unique transcriptional program and produced interleukin 2 (IL-2) and IL-10. Unlike other iNKT cells, they lacked PLZF but expressed the transcription factor E4BP4, which controlled their IL-10 production. The adipose iNKT cells were a tissue-resident population that induced an anti-inflammatory phenotype in macrophages and, through the production of IL-2, controlled the number, proliferation and suppressor function of regulatory T cells (Treg cells) in adipose tissue. Thus, iNKT cells in adipose tissue are unique regulators of immunological homeostasis in this tissue.

Computer control of a microgravity mammalian cell bioreactor

NASA Technical Reports Server (NTRS)

Hall, William A.

1987-01-01

The initial steps taken in developing a completely menu driven and totally automated computer control system for a bioreactor are discussed. This bioreactor is an electro-mechanical cell growth system cell requiring vigorous control of slowly changing parameters, many of which are so dynamically interactive that computer control is a necessity. The process computer will have two main functions. First, it will provide continuous environmental control utilizing low signal level transducers as inputs and high powered control devices such as solenoids and motors as outputs. Secondly, it will provide continuous environmental monitoring, including mass data storage and periodic data dumps to a supervisory computer.

A programmable synthetic lineage-control network that differentiates human IPSCs into glucose-sensitive insulin-secreting beta-like cells

PubMed Central

Saxena, Pratik; Heng, Boon Chin; Bai, Peng; Folcher, Marc; Zulewski, Henryk; Fussenegger, Martin

2016-01-01

Synthetic biology has advanced the design of standardized transcription control devices that programme cellular behaviour. By coupling synthetic signalling cascade- and transcription factor-based gene switches with reverse and differential sensitivity to the licensed food additive vanillic acid, we designed a synthetic lineage-control network combining vanillic acid-triggered mutually exclusive expression switches for the transcription factors Ngn3 (neurogenin 3; OFF-ON-OFF) and Pdx1 (pancreatic and duodenal homeobox 1; ON-OFF-ON) with the concomitant induction of MafA (V-maf musculoaponeurotic fibrosarcoma oncogene homologue A; OFF-ON). This designer network consisting of different network topologies orchestrating the timely control of transgenic and genomic Ngn3, Pdx1 and MafA variants is able to programme human induced pluripotent stem cells (hIPSCs)-derived pancreatic progenitor cells into glucose-sensitive insulin-secreting beta-like cells, whose glucose-stimulated insulin-release dynamics are comparable to human pancreatic islets. Synthetic lineage-control networks may provide the missing link to genetically programme somatic cells into autologous cell phenotypes for regenerative medicine. PMID:27063289

The farnesoid-X-receptor in myeloid cells controls CNS autoimmunity in an IL-10-dependent fashion.

PubMed

Hucke, Stephanie; Herold, Martin; Liebmann, Marie; Freise, Nicole; Lindner, Maren; Fleck, Ann-Katrin; Zenker, Stefanie; Thiebes, Stephanie; Fernandez-Orth, Juncal; Buck, Dorothea; Luessi, Felix; Meuth, Sven G; Zipp, Frauke; Hemmer, Bernhard; Engel, Daniel Robert; Roth, Johannes; Kuhlmann, Tanja; Wiendl, Heinz; Klotz, Luisa

2016-09-01

Innate immune responses by myeloid cells decisively contribute to perpetuation of central nervous system (CNS) autoimmunity and their pharmacologic modulation represents a promising strategy to prevent disease progression in Multiple Sclerosis (MS). Based on our observation that peripheral immune cells from relapsing-remitting and primary progressive MS patients exhibited strongly decreased levels of the bile acid receptor FXR (farnesoid-X-receptor, NR1H4), we evaluated its potential relevance as therapeutic target for control of established CNS autoimmunity. Pharmacological FXR activation promoted generation of anti-inflammatory macrophages characterized by arginase-1, increased IL-10 production, and suppression of T cell responses. In mice, FXR activation ameliorated CNS autoimmunity in an IL-10-dependent fashion and even suppressed advanced clinical disease upon therapeutic administration. In analogy to rodents, pharmacological FXR activation in human monocytes from healthy controls and MS patients induced an anti-inflammatory phenotype with suppressive properties including control of effector T cell proliferation. We therefore, propose an important role of FXR in control of T cell-mediated autoimmunity by promoting anti-inflammatory macrophage responses.

Spatial control of translation repression and polarized growth by conserved NDR kinase Orb6 and RNA-binding protein Sts5.

PubMed

Nuñez, Illyce; Rodriguez Pino, Marbelys; Wiley, David J; Das, Maitreyi E; Chen, Chuan; Goshima, Tetsuya; Kume, Kazunori; Hirata, Dai; Toda, Takashi; Verde, Fulvia

2016-07-30

RNA-binding proteins contribute to the formation of ribonucleoprotein (RNP) granules by phase transition, but regulatory mechanisms are not fully understood. Conserved fission yeast NDR (Nuclear Dbf2-Related) kinase Orb6 governs cell morphogenesis in part by spatially controlling Cdc42 GTPase. Here we describe a novel, independent function for Orb6 kinase in negatively regulating the recruitment of RNA-binding protein Sts5 into RNPs to promote polarized cell growth. We find that Orb6 kinase inhibits Sts5 recruitment into granules, its association with processing (P) bodies, and degradation of Sts5-bound mRNAs by promoting Sts5 interaction with 14-3-3 protein Rad24. Many Sts5-bound mRNAs encode essential factors for polarized cell growth, and Orb6 kinase spatially and temporally controls the extent of Sts5 granule formation. Disruption of this control system affects cell morphology and alters the pattern of polarized cell growth, revealing a role for Orb6 kinase in the spatial control of translational repression that enables normal cell morphogenesis.

Clinical Control of HIV-1 by Cytotoxic T Cells Specific for Multiple Conserved Epitopes.

PubMed

Murakoshi, Hayato; Akahoshi, Tomohiro; Koyanagi, Madoka; Chikata, Takayuki; Naruto, Takuya; Maruyama, Rie; Tamura, Yoshiko; Ishizuka, Naoki; Gatanaga, Hiroyuki; Oka, Shinichi; Takiguchi, Masafumi

2015-05-01

Identification and characterization of CD8(+) T cells effectively controlling HIV-1 variants are necessary for the development of AIDS vaccines and for studies of AIDS pathogenesis, although such CD8(+) T cells have been only partially identified. In this study, we sought to identify CD8(+) T cells controlling HIV-1 variants in 401 Japanese individuals chronically infected with HIV-1 subtype B, in which protective alleles HLA-B*57 and HLA-B*27 are very rare, by using comprehensive and exhaustive methods. We identified 13 epitope-specific CD8(+) T cells controlling HIV-1 in Japanese individuals, though 9 of these epitopes were not previously reported. The breadths of the T cell responses to the 13 epitopes were inversely associated with plasma viral load (P = 2.2 × 10(-11)) and positively associated with CD4 count (P = 1.2 × 10(-11)), indicating strong synergistic effects of these T cells on HIV-1 control in vivo. Nine of these epitopes were conserved among HIV-1 subtype B-infected individuals, whereas three out of four nonconserved epitopes were cross-recognized by the specific T cells. These findings indicate that these 12 epitopes are strong candidates for antigens for an AIDS vaccine. The present study highlighted a strategy to identify CD8(+) T cells controlling HIV-1 and demonstrated effective control of HIV-1 by those specific for 12 conserved or cross-reactive epitopes. HLA-B*27-restricted and HLA-B*57-restricted cytotoxic T lymphocytes (CTLs) play a key role in controlling HIV-1 in Caucasians and Africans, whereas it is unclear which CTLs control HIV-1 in Asian countries, where HLA-B*57 and HLA-B*27 are very rare. A recent study showed that HLA-B*67:01 and HLA-B*52:01-C*12:02 haplotypes were protective alleles in Japanese individuals, but it is unknown whether CTLs restricted by these alleles control HIV-1. In this study, we identified 13 CTLs controlling HIV-1 in Japan by using comprehensive and exhaustive methods. They included 5 HLA-B*52:01-restricted and 3 HLA-B*67:01-restricted CTLs, suggesting that these CTLs play a predominant role in HIV-1 control. The 13 CTLs showed synergistic effects on HIV-1 control. Twelve out of these 13 epitopes were recognized as conserved or cross-recognized ones. These findings strongly suggest that these 12 epitopes are candidates for antigens for AIDS vaccines. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Clinical Control of HIV-1 by Cytotoxic T Cells Specific for Multiple Conserved Epitopes

PubMed Central

Murakoshi, Hayato; Akahoshi, Tomohiro; Koyanagi, Madoka; Chikata, Takayuki; Naruto, Takuya; Maruyama, Rie; Tamura, Yoshiko; Ishizuka, Naoki; Gatanaga, Hiroyuki; Oka, Shinichi

2015-01-01

ABSTRACT Identification and characterization of CD8+ T cells effectively controlling HIV-1 variants are necessary for the development of AIDS vaccines and for studies of AIDS pathogenesis, although such CD8+ T cells have been only partially identified. In this study, we sought to identify CD8+ T cells controlling HIV-1 variants in 401 Japanese individuals chronically infected with HIV-1 subtype B, in which protective alleles HLA-B*57 and HLA-B*27 are very rare, by using comprehensive and exhaustive methods. We identified 13 epitope-specific CD8+ T cells controlling HIV-1 in Japanese individuals, though 9 of these epitopes were not previously reported. The breadths of the T cell responses to the 13 epitopes were inversely associated with plasma viral load (P = 2.2 × 10−11) and positively associated with CD4 count (P = 1.2 × 10−11), indicating strong synergistic effects of these T cells on HIV-1 control in vivo. Nine of these epitopes were conserved among HIV-1 subtype B-infected individuals, whereas three out of four nonconserved epitopes were cross-recognized by the specific T cells. These findings indicate that these 12 epitopes are strong candidates for antigens for an AIDS vaccine. The present study highlighted a strategy to identify CD8+ T cells controlling HIV-1 and demonstrated effective control of HIV-1 by those specific for 12 conserved or cross-reactive epitopes. IMPORTANCE HLA-B*27-restricted and HLA-B*57-restricted cytotoxic T lymphocytes (CTLs) play a key role in controlling HIV-1 in Caucasians and Africans, whereas it is unclear which CTLs control HIV-1 in Asian countries, where HLA-B*57 and HLA-B*27 are very rare. A recent study showed that HLA-B*67:01 and HLA-B*52:01-C*12:02 haplotypes were protective alleles in Japanese individuals, but it is unknown whether CTLs restricted by these alleles control HIV-1. In this study, we identified 13 CTLs controlling HIV-1 in Japan by using comprehensive and exhaustive methods. They included 5 HLA-B*52:01-restricted and 3 HLA-B*67:01-restricted CTLs, suggesting that these CTLs play a predominant role in HIV-1 control. The 13 CTLs showed synergistic effects on HIV-1 control. Twelve out of these 13 epitopes were recognized as conserved or cross-recognized ones. These findings strongly suggest that these 12 epitopes are candidates for antigens for AIDS vaccines. PMID:25741000

The frequency and severity of epistaxis in children with sickle cell anaemia in eastern Uganda: a case-control study.

PubMed

Nardo-Marino, Amina; Williams, Thomas N; Olupot-Olupot, Peter

2017-01-01

There are a paucity of data on epistaxis as it pertains to sickle cell anaemia. Some case studies suggest epistaxis to be a significant complication in patients with sickle cell anaemia in sub-Saharan Africa; however, no robust studies have sought to establish the epidemiology or pathophysiology of this phenomenon. We conducted a case-control study with the aim of investigating the importance of epistaxis among children presenting with sickle cell anaemia at the Mbale Regional Referral Hospital in eastern Uganda. Cases were children aged 2-15 years with an existing diagnosis of laboratory confirmed sickle cell anaemia, while controls were children without sickle cell anaemia who were frequency matched to cases on the basis of age group and gender. The frequency and severity of epistaxis was assessed using a structured questionnaire developed specifically for this study. Odds ratios controlled for age group and gender were calculated using unconditional logistic regression. A total of 150 children were included, 73 children with sickle cell anaemia and 77 children without sickle cell anaemia. The overall prevalence of epistaxis among children with sickle cell anaemia and children without sickle cell anaemia was 32.9 and 23.4% respectively. The case-control odds ratios for epistaxis, recurrent epistaxis and severe epistaxis were, 1.6 (95%CI 0.8-3.4; p  = 0.2), 7.4 (1.6-34.5; 0.01), and 8.3 (1.0-69.8; 0.05) respectively. Our results suggest that in eastern Uganda, children with sickle cell anaemia experience epistaxis more frequently and with greater severity than children without sickle cell anaemia. Further studies are indicated to confirm this conclusion and investigate aetiology.

Designing a Binding Interface for Control of Cancer Cell Adhesion via 3D Topography and Metabolic Oligosaccharide Engineering

PubMed Central

Du, Jian; Che, Pao-Lin; Wang, Zhi-Yun; Aich, Udayanath; Yarema, Kevin J.

2011-01-01

This study combines metabolic oligosaccharide engineering (MOE), a technology where the glycocalyx of living cells is endowed with chemical features not normally found in sugars, with custom-designed three dimensional biomaterial substrates to enhance the adhesion of cancer cells and control their morphology and gene expression. Specifically, Ac5ManNTGc, a thiol-bearing analogue of N-acetyl-d-mannosamine (ManNAc) was used to introduce thiolated sialic acids into the glycocalyx of human Jurkat T-lymphoma derived cells. In parallel 2D films and 3D electrospun nanofibrous scaffolds were prepared from polyethersulfone (PES) and (as controls) left unmodified or aminated. Alternately, the materials were malemided or gold-coated to provide bioorthogonal binding partners for the thiol groups newly expressed on the cell surface. Cell attachment was modulated by both the topography of the substrate surface and by the chemical compatibility of the binding interface between the cell and the substrate; a substantial increase in binding for normally non-adhesive Jurkat line for 3D scaffold compared to 2D surfaces with an added degree of adhesion resulting from chemoselective binding to malemidede-derivatived or gold-coated surfaces. In addition, the morphology of the cells attached to the 3D scaffolds via MOE-mediated adhesion was dramatically altered and the expression of genes involved in cell adhesion changed in a time-dependent manner. This study showed that cell adhesion could be enhanced, gene expression modulated, and cell fate controlled by introducing the 3D topograhical cues into the growth substrate and by creating a glycoengineered binding interface where the chemistry of both the cell surface and biomaterials scaffold was controlled to facilitate a new mode of carbohydrate-mediated adhesion. PMID:21549424

The Outer Loop bioreactor: a case study of settlement monitoring and solids decomposition.

PubMed

Abichou, Tarek; Barlaz, Morton A; Green, Roger; Hater, Gary

2013-10-01

The Outer Loop landfill bioreactor (OLLB) located in Louisville, KY, USA has been in operation since 2000 and represents an opportunity to evaluate long-term bioreactor monitoring data at a full-scale operational landfill. Three types of landfill units were studied including a Control cell, a new landfill area that had a piping network installed as waste was being placed to support leachate recirculation (As-Built cell), and a conventional landfill that was modified to allow for liquid recirculation (Retrofit cell). The objective of this study is to summarize the results of settlement data and assess how these data relate to solids decomposition monitoring at the OLLB. The Retrofit cells started to settle as soon as liquids were introduced. The cumulative settlement during the 8years of monitoring varied from 60 to 100cm. These results suggest that liquid recirculation in the Retrofit cells caused a 5-8% reduction in the thickness of the waste column. The average long-term settlement in the As-Built and Control Cells was about 37% and 19%, respectively. The modified compression index (Cα(')) was 0.17 for the Control cells and 0.2-0.48 for the As-Built cells. While the As-Built cells exhibited greater settlement than the Control cells, the data do not support biodegradation as the only explanation. The increased settlement in the As-Built bioreactor cell appeared to be associated with liquid movement and not with biodegradation because both chemical (biochemical methane potential) and physical (moisture content) indicators of decomposition were similar in the Control and As-Built cells. The solids data are consistent with the concept that bioreactor operations accelerate the rate of decomposition, but not necessarily the cumulative loss of anaerobically degradable solids. Copyright © 2013 Elsevier Ltd. All rights reserved.

Genetic models rule out a major role of beta cell glycogen in the control of glucose homeostasis.

PubMed

Mir-Coll, Joan; Duran, Jordi; Slebe, Felipe; García-Rocha, Mar; Gomis, Ramon; Gasa, Rosa; Guinovart, Joan J

2016-05-01

Glycogen accumulation occurs in beta cells of diabetic patients and has been proposed to partly mediate glucotoxicity-induced beta cell dysfunction. However, the role of glycogen metabolism in beta cell function and its contribution to diabetes pathophysiology remain poorly understood. We investigated the function of beta cell glycogen by studying glucose homeostasis in mice with (1) defective glycogen synthesis in the pancreas; and (2) excessive glycogen accumulation in beta cells. Conditional deletion of the Gys1 gene and overexpression of protein targeting to glycogen (PTG) was accomplished by Cre-lox recombination using pancreas-specific Cre lines. Glucose homeostasis was assessed by determining fasting glycaemia, insulinaemia and glucose tolerance. Beta cell mass was determined by morphometry. Glycogen was detected histologically by periodic acid-Schiff's reagent staining. Isolated islets were used for the determination of glycogen and insulin content, insulin secretion, immunoblots and gene expression assays. Gys1 knockout (Gys1 (KO)) mice did not exhibit differences in glucose tolerance or basal glycaemia and insulinaemia relative to controls. Insulin secretion and gene expression in isolated islets was also indistinguishable between Gys1 (KO) and controls. Conversely, despite effective glycogen overaccumulation in islets, mice with PTG overexpression (PTG(OE)) presented similar glucose tolerance to controls. However, under fasting conditions they exhibited lower glycaemia and higher insulinaemia. Importantly, neither young nor aged PTG(OE) mice showed differences in beta cell mass relative to age-matched controls. Finally, a high-fat diet did not reveal a beta cell-autonomous phenotype in either model. Glycogen metabolism is not required for the maintenance of beta cell function. Glycogen accumulation in beta cells alone is not sufficient to trigger the dysfunction or loss of these cells, or progression to diabetes.

Sarcoidosis Th17 Cells are ESAT-6 Antigen Specific but Demonstrate Reduced IFN-γ Expression

PubMed Central

Richmond, Bradley W.; Ploetze, Kristen; Isom, Joan; Chambers-Harris, Isfahan; Braun, Nicole A.; Taylor, Thyneice; Abraham, Susamma; Mageto, Yolanda; Culver, Dan A.; Oswald-Richter, Kyra A.; Drake, Wonder P.

2013-01-01

Rationale Sarcoidosis is a granulomatous disease of unknown etiology. Many patients with sarcoidosis demonstrate antigen-specific immunity to mycobacterial virulence factors. Th-17 cells are crucial to the immune response in granulomatous inflammation, and have recently been shown to be present in greater numbers in the peripheral blood and bronchoalveolar lavage (BAL) fluid (BALF) of sarcoidosis patients than healthy controls. It is unclear whether Th-17 cells in sarcoidosis are specific for mycobacterial antigens, or whether they have similar functionality to control Th-17 cells. Methods Flow cytometry was used to determine the numbers of Th-17 cells present in the peripheral blood and BALF of patients with sarcoidosis, the percentage of Th-17 cells that were specific to the mycobacterial virulence factor ESAT-6, and as well as to assess IFN-γ expression in Th-17 cells following polyclonal stimulation. Results Patients with sarcoidosis had greater numbers of Th-17 cells in the peripheral blood and BALF than controls and produced significantly more extracellular IL-17A (p=0.03 and p=0.02, respectively). ESAT-6 specific Th-17 cells were present in both peripheral blood and BALF of sarcoidosis patients (p<0.001 and p=0.03, respectively). After polyclonal stimulation, Th-17 cells from sarcoidosis patients produced less IFN-γ than healthy controls. Conclusions Patients with sarcoidosis have mycobacterial antigen-specific Th-17 cells peripherally and in sites of active sarcoidosis involvement. Despite the Th1 immunophenotype of sarcoidosis immunology, the Th-17 cells have reduced IFN-γ expression, compared to healthy controls. This reduction in immunity may contribute to sarcoidosis pathogenesis. PMID:23073617

Multifunctional ferromagnetic disks for modulating cell function

PubMed Central

Vitol, Elina A.; Novosad, Valentyn; Rozhkova, Elena A.

2013-01-01

In this work, we focus on the methods for controlling cell function with ferromagnetic disk-shaped particles. We will first review the history of magnetically assisted modulation of cell behavior and applications of magnetic particles for studying physical properties of a cell. Then, we consider the biological applications of the microdisks such as the method for induction of cancer cell apoptosis, controlled drug release, hyperthermia and MRI imaging. PMID:23766544

Altered Gene Expression and Function of Peripheral Blood Natural Killer Cells in Children with Autism

PubMed Central

Enstrom, A M; Lit, L; Onore, C E; Gregg, J P; Hansen, R; Pessah, I N; Hertz-Picciotto, I; Van de Water, J A; Sharp, F R; Ashwood, P

2009-01-01

Immune related abnormalities have repeatedly been reported in autism spectrum disorders (ASD), including evidence of immune dysregulation and autoimmune phenomena. NK cells may play an important role in neurodevelopmental disorders such as ASD. Here we performed a gene expression screen and cellular functional analysis on peripheral blood obtained from 52 children with ASD and 27 typically developing control children enrolled in the case-control CHARGE study. RNA expression of NK cell receptors and effector molecules were significantly upregulated in ASD. Flow cytometric analysis of NK cells demonstrated increased production of perforin, granzyme B, and interferon gamma (IFNγ) under resting conditions in children with ASD (p<0.01). Following NK cell stimulation in the presence of K562 target cells, the cytotoxicity of NK cells was significantly reduced in ASD compared with controls (p<0.02). Furthermore, under similar stimulation conditions the presence of perforin, granzyme B, and IFNγ in NK cells from ASD children was significantly lower compared with controls (p<0.001). These findings suggest possible dysfunction of NK cells in children with ASD. Abnormalities in NK cells may represent a susceptibility factor in ASD and may predispose to the development of autoimmunity and/or adverse neuroimmune interactions during critical periods of development. PMID:18762240

The inhibition of apoptosis in EL4 lymphoma cells overexpressing growth hormone.

PubMed

Arnold, Robyn E; Weigent, Douglas A

2004-01-01

The antiapoptotic action of exogenous growth hormone (GH) has been reported for several lymphoid cell lines; however, the potential role of endogenous GH in apoptosis has not been thoroughly investigated. This study was designed to investigate the effects of endogenous GH on apoptosis induced by methyl methanesulfonate (MMS) in a T cell lymphoma overexpressing GH (GHo). The results of these experiments have shown that in EL4 lymphoma cells, overexpression of GH sustained viability after exposure to MMS compared to control cells. The extent of DNA fragmentation measured by ladder formation on agarose gels was reduced in GHo cells following treatment with MMS, when compared to control cells. Adding exogenous GH to control cells and treatment of GHo cells with antibodies to GH had no effect on MMS-induced DNA ladder formation. In further studies, DNA microarray analysis suggested a marked decrease in the constitutive expression of bax, BAD, and caspases 3, 8, and 9 in GHo cells compared to controls. In addition, after treatment with MMS, the activities of caspases 2, 3, 6, 8, and 9 were all lower than control in GHo cells. Western blot analysis detected an increase in Bcl-2 while the levels of nuclear factor kappa B (NFkappaB) remained unchanged in GHo cells. Treatment of EL4 cells with antisense deoxyoligonucleotides to GH and specific inhibitors of NFkappaB (SN-50) increased DNA fragmentation. GHo cells show increased levels of phosphorylated Akt and GSK-3, suggesting inactivation of this proapoptotic protein. The results, taken together with our previous data which showed increased nitric oxide formation in GHo cells, suggest a possible mechanism for the antiapoptotic effects of endogenous GH through the production of nitric oxide and support the idea that endogenous GH may play an important role in the survival of lymphocytes exposed to stressful stimuli. Copyright 2004 S. Karger AG, Basel

Flow cytometry analysis of T-cell subsets in cerebrospinal fluid of narcolepsy type 1 patients with long-lasting disease.

PubMed

Moresco, Monica; Lecciso, Mariangela; Ocadlikova, Darina; Filardi, Marco; Melzi, Silvia; Kornum, Birgitte Rahbek; Antelmi, Elena; Pizza, Fabio; Mignot, Emmanuel; Curti, Antonio; Plazzi, Giuseppe

2018-04-01

Type 1 narcolepsy (NT1) is a central hypersomnia linked to the destruction of hypocretin-producing neurons. A great body of genetic and epidemiological data points to likely autoimmune disease aetiology. Recent reports have characterized peripheral blood T-cell subsets in NT1, whereas data regarding the cerebrospinal fluid (CSF) immune cell composition are lacking. The current study aimed to characterize the T-cell and natural killer (NK) cell subsets in NT1 patients with long disease course. Immune cell subsets from CSF and peripheral blood mononuclear cell (PBMC) samples were analysed by flow cytometry in two age-balanced and sex-balanced groups of 14 NT1 patients versus 14 healthy controls. The frequency of CSF cell groups was compared with PBMCs. Non-parametric tests were used for statistical analyses. The NT1 patients did not show significant differences of CSF immune cell subsets compared to controls, despite a trend towards higher CD4 + terminally differentiated effector memory T cells. T cells preferentially displayed a memory phenotype in the CSF compared to PBMCs. Furthermore, a reduced frequency of CD4 + terminally differentiated effector memory T cells and an increased frequency of NK CD56 bright cells was observed in PBMCs from patients compared to controls. Finally, the ratio between CSF and peripheral CD4 + terminally differentiated effector memory T cells was two-fold increased in NT1 patients versus controls. Significant differences in PBMCs and in CSF/PBMC ratios of immune cell profile were found in NT1 patients compared to healthy controls. These differences might have arisen from the different HLA status, or be primary or secondary to hypocretin deficiency. Further functional studies in patients close to disease onset are required to understand NT1 pathophysiology. Copyright © 2017 Elsevier B.V. All rights reserved.

Human Keratinocytes That Express hTERT and Also Bypass a p16INK4a-Enforced Mechanism That Limits Life Span Become Immortal yet Retain Normal Growth and Differentiation Characteristics

PubMed Central

Dickson, Mark A.; Hahn, William C.; Ino, Yasushi; Ronfard, Vincent; Wu, Jenny Y.; Weinberg, Robert A.; Louis, David N.; Li, Frederick P.; Rheinwald, James G.

2000-01-01

Normal human cells exhibit a limited replicative life span in culture, eventually arresting growth by a process termed senescence. Progressive telomere shortening appears to trigger senescence in normal human fibroblasts and retinal pigment epithelial cells, as ectopic expression of the telomerase catalytic subunit, hTERT, immortalizes these cell types directly. Telomerase expression alone is insufficient to enable certain other cell types to evade senescence, however. Such cells, including keratinocytes and mammary epithelial cells, appear to require loss of the pRB/p16INK4a cell cycle control mechanism in addition to hTERT expression to achieve immortality. To investigate the relationships among telomerase activity, cell cycle control, senescence, and differentiation, we expressed hTERT in two epithelial cell types, keratinocytes and mesothelial cells, and determined the effect on proliferation potential and on the function of cell-type-specific growth control and differentiation systems. Ectopic hTERT expression immortalized normal mesothelial cells and a premalignant, p16INK4a-negative keratinocyte line. In contrast, when four keratinocyte strains cultured from normal tissue were transduced to express hTERT, they were incompletely rescued from senescence. After reaching the population doubling limit of their parent cell strains, hTERT+ keratinocytes entered a slow growth phase of indefinite length, from which rare, rapidly dividing immortal cells emerged. These immortal cell lines frequently had sustained deletions of the CDK2NA/INK4A locus or otherwise were deficient in p16INK4a expression. They nevertheless typically retained other keratinocyte growth controls and differentiated normally in culture and in xenografts. Thus, keratinocyte replicative potential is limited by a p16INK4a-dependent mechanism, the activation of which can occur independent of telomere length. Abrogation of this mechanism together with telomerase expression immortalizes keratinocytes without affecting other major growth control or differentiation systems. PMID:10648628

Cell size control and homeostasis in bacteria

NASA Astrophysics Data System (ADS)

Bradde, Serena; Taheri, Sattar; Sauls, John; Hill, Nobert; Levine, Petra; Paulsson, Johan; Vergassola, Massimo; Jun, Suckjoon

2015-03-01

How cells control their size is a fundamental question in biology. The mechanisms for sensing size, time, or a combination of the two are not supported by experimental evidence. By analysing distributions of size at division at birth and generation time of hundreds of thousands of Gram-negative E. coli and Gram-positive B. subtilis cells under a wide range of tightly controlled steady-state growth conditions, we are now in the position to validate different theoretical models. In this talk I will present all possible models in details and present a general mechanism that quantitatively explains all measurable aspects of growth and cell division at both population and single-cell levels.

Subcellular and supracellular mechanical stress prescribes cytoskeleton behavior in Arabidopsis cotyledon pavement cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sampathkumar, Arun; Krupinski, Pawel; Wightman, Raymond

Although it is a central question in biology, how cell shape controls intracellular dynamics largely remains an open question. Here, we show that the shape of Arabidopsis pavement cells creates a stress pattern that controls microtubule orientation, which then guides cell wall reinforcement. Live-imaging, combined with modeling of cell mechanics, shows that microtubules align along the maximal tensile stress direction within the cells, and atomic force microscopy demonstrates that this leads to reinforcement of the cell wall parallel to the microtubules. This feedback loop is regulated: cell-shape derived stresses could be overridden by imposed tissue level stresses, showing how competitionmore » between subcellular and supracellular cues control microtubule behavior. Furthermore, at the microtubule level, we identified an amplification mechanism in which mechanical stress promotes the microtubule response to stress by increasing severing activity. These multiscale feedbacks likely contribute to the robustness of microtubule behavior in plant epidermis.« less

Subcellular and supracellular mechanical stress prescribes cytoskeleton behavior in Arabidopsis cotyledon pavement cells

DOE PAGES

Sampathkumar, Arun; Krupinski, Pawel; Wightman, Raymond; ...

2014-04-16

Although it is a central question in biology, how cell shape controls intracellular dynamics largely remains an open question. Here, we show that the shape of Arabidopsis pavement cells creates a stress pattern that controls microtubule orientation, which then guides cell wall reinforcement. Live-imaging, combined with modeling of cell mechanics, shows that microtubules align along the maximal tensile stress direction within the cells, and atomic force microscopy demonstrates that this leads to reinforcement of the cell wall parallel to the microtubules. This feedback loop is regulated: cell-shape derived stresses could be overridden by imposed tissue level stresses, showing how competitionmore » between subcellular and supracellular cues control microtubule behavior. Furthermore, at the microtubule level, we identified an amplification mechanism in which mechanical stress promotes the microtubule response to stress by increasing severing activity. These multiscale feedbacks likely contribute to the robustness of microtubule behavior in plant epidermis.« less

Estradiol targets T cell signaling pathways in human systemic lupus.

PubMed

Walters, Emily; Rider, Virginia; Abdou, Nabih I; Greenwell, Cindy; Svojanovsky, Stan; Smith, Peter; Kimler, Bruce F

2009-12-01

The major risk factor for developing systemic lupus erythematosus (SLE) is being female. The present study utilized gene profiles of activated T cells from females with SLE and healthy controls to identify signaling pathways uniquely regulated by estradiol that could contribute to SLE pathogenesis. Selected downstream pathway genes (+/- estradiol) were measured by real time polymerase chain amplification. Estradiol uniquely upregulated six pathways in SLE T cells that control T cell function including interferon-alpha signaling. Measurement of interferon-alpha pathway target gene expression revealed significant differences (p= 0.043) in DRIP150 (+/- estradiol) in SLE T cell samples while IFIT1 expression was bimodal and correlated moderately (r= 0.55) with disease activity. The results indicate that estradiol alters signaling pathways in activated SLE T cells that control T cell function. Differential expression of transcriptional coactivators could influence estrogen-dependent gene regulation in T cell signaling and contribute to SLE onset and disease pathogenesis.

microRNAs as Pharmacological Targets in Endothelial Cell Function and Dysfunction

PubMed Central

Chamorro-Jorganes, Aránzazu; Araldi, Elisa; Suárez, Yajaira

2013-01-01

Endothelial cell dysfunction is a term which implies the dysregulation of normal endothelial cell functions, including impairment of the barrier functions, control of vascular tone, disturbance of proliferative, migratory and morphogenic capacities of endothelial cells, as well as control of leukocyte trafficking. MicroRNAs (miRNAs) are short non-coding RNAs that have emerged as critical regulators of gene expression acting predominantly at the post-transcriptional level. This review summarizes the latest insights in the identification of endothelial-specific miRNAs and their targets, as well as their roles in controlling endothelial cell functions in both autocrine and paracrine manner. In addition, we discuss the therapeutic potential for the treatment of endothelial cell dysfunction and associated vascular pathophysiological conditions. PMID:23603154

Focal adhesion kinase is involved in mechanosensing during fibroblast migration

NASA Technical Reports Server (NTRS)

Wang, H. B.; Dembo, M.; Hanks, S. K.; Wang, Y.

2001-01-01

Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase localized at focal adhesions and is believed to mediate adhesion-stimulated effects. Although ablation of FAK impairs cell movement, it is not clear whether FAK might be involved in the guidance of cell migration, a role consistent with its putative regulatory function. We have transfected FAK-null fibroblasts with FAK gene under the control of the tetracycline repression system. Cells were cultured on flexible polyacrylamide substrates for the detection of traction forces and the application of mechanical stimulation. Compared with control cells expressing wild-type FAK, FAK-null cells showed a decrease in migration speed and directional persistence. In addition, whereas FAK-expressing cells responded to exerted forces by reorienting their movements and forming prominent focal adhesions, FAK-null cells failed to show such responses. Furthermore, FAK-null cells showed impaired responses to decreases in substrate flexibility, which causes control cells to generate weaker traction forces and migrate away from soft substrates. Cells expressing Y397F FAK, which cannot be phosphorylated at a key tyrosine site, showed similar defects in migration pattern and force-induced reorientation as did FAK-null cells. However, other aspects of F397-FAK cells, including the responses to substrate flexibility and the amplification of focal adhesions upon mechanical stimulation, were similar to that of control cells. Our results suggest that FAK plays an important role in the response of migrating cells to mechanical input. In addition, phosphorylation at Tyr-397 is required for some, but not all, of the functions of FAK in cell migration.

Decoupling activation and exhaustion of B cells in spontaneous controllers of HIV infection

PubMed Central

Sciaranghella, Gaia; Tong, Neath; Mahan, Alison E.; Suscovich, Todd J.; Alter, Galit

2013-01-01

Objective To define the impact of chronic viremia and associated immune activation on B-cell exhaustion in HIV infection. Design Progressive HIV infection is marked by B-cell anergy and exhaustion coupled with dramatic hypergammaglobulinemia. Although both upregulation of CD95 and loss of CD21 have been used as markers of infection-associated B-cell dysfunction, little is known regarding the specific profiles of dysfunctional B cells and whether persistent viral replication and its associated immune activation play a central role in driving B-cell dysfunction. Methods Multiparameter flow cytometry was used to define the profile of dysfunctional B cells. The changes in the expression of CD21 and CD95 were tracked on B-cell subpopulations in patients with differential control of viral replication. Results Although the emergence of exhausted, CD21low tissue-like memory B cells followed similar patterns in both progressors and controllers, the frequency of CD21low activated memory B cells was lower in spontaneous controllers. Conclusion Our results suggest that the loss of CD21 and the upregulation of CD95 occur as separate events during the development of B-cell dysfunction. The loss of CD21 is a marker of B-cell exhaustion induced in the absence of appreciable viral replication, whereas the upregulation of CD95 is tightly linked to persistent viral replication and its associated immune activation. Thus, these dysfunctional profiles potentially represent two functionally distinct states within the B-cell compartment. PMID:23135171

Traditional Chinese medicine herbal mixture LQ arrests FUCCI-expressing HeLa cells in G₀/G₁ phase in 2D plastic, 2.5D Matrigel, and 3D Gelfoam culture visualized with FUCCI imaging.

PubMed

Zhang, Lei; Wu, Chengyu; Bouvet, Michael; Yano, Shuya; Hoffman, Robert M

2015-03-10

We used the fluorescence ubiquitination-based cell cycle indicator (FUCCI) to monitor cell cycle arrest after treatment of FUCCI-expressing HeLa cells (FUCCI-HeLa) with a traditional Chinese medicine (TCM) herbal mixture LQ, previously shown to have anti-tumor and anti-metastatic activity in mouse models. Paclitaxel was used as the positive control. In 2D monolayer culture, the untreated control had approximately 45% of the cells in S/G₂/M phase. In contrast, the LQ-treated cells (9 mg/ml) were mostly in the G₀/G₁ (>90%) after 72 hours. After treatment with paclitaxel (0.01 μm), for 72 hours, 95% of the cells were in S/G₂/M. In 2.5D Matrigel culture, the colonies in the untreated control group had 40% of the cells in S/G₂/M. LQ arrested the cells in G₀/G₁ after 72 hours. Paclitaxel arrested almost all the cells in S/G₂/M after 72 hours. In 3D Gelfoam culture, the untreated control culture had approximately 45% of cells in G₂/M. In contrast, the LQ-treated cells were mostly in G₀/G₁ phase (>80%) after 72 hours treatment. Paclitaxel resulted in 90% of the cells arrested in S/G₂/M after 72 hours. The present report suggests the non-toxic LQ has potential to maintain cancers in a quiescent state for long periods of time.

High-density mammalian cell cultures in stirred-tank bioreactor without external pH control.

PubMed

Xu, Sen; Chen, Hao

2016-08-10

Maintaining desired pH is a necessity for optimal cell growth and protein production. It is typically achieved through a two-sided pH control loop on the bioreactor controller. Here we investigated cell culture processes with minimum or no pH control and demonstrated that high-density mammalian cell cultures could be maintained for long-term protein production without pH control. The intrinsic interactions between pCO2, lactate, and pH were leveraged to maintain culture pH. Fed-batch cultures at the same lower pH limit of 6.75 but different upper pH limits (7.05, 7.30, 7.45, 7.65) were evaluated in the 3L bioreactors and comparable results were obtained. Neither CO2 sparging nor base addition was required to control pH in the pH range of 6.75-7.65. The impact of sparger configurations (drilled hole sparger vs. frit sparger) and scales (3L vs. 200L) on CO2 accumulation and culture pH was also demonstrated. The same principle was applied in two perfusion cultures with steady state cell densities at 42.5±3.3 or 68.3±6.0×10(6)cells/mL with low cell specific perfusion rates (15±2 to 23±3pL/cell/day), achieving up to 1.9±0.1g/L/day bioreactor productivity. Culture pH level in the 3L perfusion bioreactors was steadily maintained by controlling the residual lactate and pCO2 levels without the requirement of external pH control for up to 40days with consistent productivity and product quality. Furthermore, culture pH could be potentially modulated via adjusting residual glucose levels and CO2 stripping capability in perfusion cultures. To the best of our knowledge, this is the first time a systematic study was performed to evaluate the long-term cell cultivation and protein production in stirred-tank bioreactors without external pH control. Copyright © 2016 Elsevier B.V. All rights reserved.

Controlled ice nucleation in cryopreservation--a review.

PubMed

Morris, G John; Acton, Elizabeth

2013-04-01

We review here for the first time, the literature on control of ice nucleation in cryopreservation. Water and aqueous solutions have a tendency to undercool before ice nucleation occurs. Control of ice nucleation has been recognised as a critical step in the cryopreservation of embryos and oocytes but is largely ignored for other cell types. We review the processes of ice nucleation and crystal growth in the solution around cells and tissues during cryopreservation with an emphasis on non IVF applications. The extent of undercooling that is encountered during the cooling of various cryocontainers is defined and the methods that have been employed to control the nucleation of ice are examined. The effects of controlled ice nucleation on the structure of the sample and the outcome of cryopreservation of a range of cell types and tissues are presented and the physical events which define the cellular response are discussed. Nucleation of ice is the most significant uncontrolled variable in conventional cryopreservation leading to sample to sample variation in cell recovery, viability and function and should be controlled to allow standardisation of cryopreservation protocols for cells for biobanking, cell based assays or clinical application. This intervention allows a way of increasing viability of cells and reducing variability between samples and should be included as standard operating procedures are developed. Copyright © 2012 Elsevier Inc. All rights reserved.

Wnt/beta-Catenin, Foxa2, and CXCR4 Axis Controls Prostate Cancer Progression

DTIC Science & Technology

2014-07-01

NT1 cells that over-expressing Foxa2. The reason we used NT1 cells for the Foxa2 over-expressing experiments is that NT1 is an AR-expressing... cells . We have also established NT1 cells over-expressing a dominant active beta-catenin. We have characterized these cells . Our research found: 1...expression profiles of control NT1 , NT1 /Foxa2, and NT1 /beta-catenin cells Figure 1. We did RT-PCR to examine the expression of key

Expression of genomic AtCYCD2;1 in Arabidopsis induces cell division at smaller cell sizes: implications for the control of plant growth.

PubMed

Qi, Ruhu; John, Peter Crook Lloyd

2007-07-01

The Arabidopsis (Arabidopsis thaliana) CYCD2;1 gene introduced in genomic form increased cell formation in the Arabidopsis root apex and leaf, while generating full-length mRNA, raised CDK/CYCLIN enzyme activity, reduced G1-phase duration, and reduced size of cells at S phase and division. Other cell cycle genes, CDKA;1, CYCLIN B;1, and the cDNA form of CYCD2;1 that produced an aberrantly spliced mRNA, produced smaller or zero increases in CDK/CYCLIN activity and did not increase the number of cells formed. Plants with a homozygous single insert of genomic CYCD2;1 grew with normal morphology and without accelerated growth of root or shoot, not providing evidence that cell formation or CYCLIN D2 controls growth of postembryonic vegetative tissues. At the root apex, cells progressed normally from meristem to elongation, but their smaller size enclosed less growth and a 40% reduction in final size of epidermal and cortical cells was seen. Smaller elongated cell size inhibited endoreduplication, indicating a cell size requirement. Leaf cells were also smaller and more numerous during proliferation and epidermal pavement and palisade cells attained 59% and 69% of controls, whereas laminas reached normal size. Autonomous control of expansion was therefore not evident in abundant cell types that formed tissues of root or leaf. Cell size was reduced by a greater number formed in a tissue prior to cell and tissue expansion. Initiation and termination of expansion did not correlate with cell dimension or number and may be determined by tissue-wide signals acting across cellular boundaries.

Cell Fate Reprogramming by Control of Intracellular Network Dynamics

PubMed Central

Zañudo, Jorge G. T.; Albert, Réka

2015-01-01

Identifying control strategies for biological networks is paramount for practical applications that involve reprogramming a cell’s fate, such as disease therapeutics and stem cell reprogramming. Here we develop a novel network control framework that integrates the structural and functional information available for intracellular networks to predict control targets. Formulated in a logical dynamic scheme, our approach drives any initial state to the target state with 100% effectiveness and needs to be applied only transiently for the network to reach and stay in the desired state. We illustrate our method’s potential to find intervention targets for cancer treatment and cell differentiation by applying it to a leukemia signaling network and to the network controlling the differentiation of helper T cells. We find that the predicted control targets are effective in a broad dynamic framework. Moreover, several of the predicted interventions are supported by experiments. PMID:25849586

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

PubMed Central

Kalos, M; Zissler, J

1990-01-01

The prokaryote Myxococcus xanthus is a model for cell interactions important in multicellular behavior. We used the transposon TnphoA to specifically identify genes for cell-surface factors involved in cell interactions. From a library of 10,700 insertions of TnphoA, we isolated 36 that produced alkaline phosphatase activity. Three TnphoA insertions tagged cell motility genes, called cgl, which control the adventurous movement of cells. The products of the tagged cgl genes could function in trans upon other cells and were localized primarily in the cell envelope and extracellular space, consistent with TnphoA tagging genes for extracellular factors controlling motility. Images PMID:2172982

Cellular immune responses to HIV

NASA Astrophysics Data System (ADS)

McMichael, Andrew J.; Rowland-Jones, Sarah L.

2001-04-01

The cellular immune response to the human immunodeficiency virus, mediated by T lymphocytes, seems strong but fails to control the infection completely. In most virus infections, T cells either eliminate the virus or suppress it indefinitely as a harmless, persisting infection. But the human immunodeficiency virus undermines this control by infecting key immune cells, thereby impairing the response of both the infected CD4+ T cells and the uninfected CD8+ T cells. The failure of the latter to function efficiently facilitates the escape of virus from immune control and the collapse of the whole immune system.

Histone Methylation and Epigenetic Silencing in Breast Cancer

DTIC Science & Technology

2010-07-01

bars show relative luciferase expression levels (firefly versus Renilla control) in SKBR3 cells treated with a control non-targeted dsRNA (NT2) and red...luciferase expression levels (firefly versus Renilla control) in SKBR3 cells treated with a control non-targeted dsRNA (NT2) and red bars depict

Peripheral natural killer cytotoxicity and CD56(pos)CD16(pos) cells increase during early pregnancy in women with a history of recurrent spontaneous abortion.

PubMed

Emmer, P M; Nelen, W L; Steegers, E A; Hendriks, J C; Veerhoek, M; Joosten, I

2000-05-01

For diagnostic purposes we assessed peripheral natural killer (NK) cell cytotoxicity and NK and T cell numbers to assess their putative predictive value in recurrent spontaneous abortion (RSA). A total of 43 women with subsequent pregnancy, 37 healthy controls and 39 women successfully partaking in an in-vitro fertilization (IVF) procedure, were included in the study. We show that before pregnancy, levels of NK cytotoxicity and numbers of both single CD56(pos) and double CD56(pos)CD16(pos) cells were similar between RSA women and controls. But notably, within the RSA group, NK cell numbers of <12% were strongly associated with a subsequent pregnancy carried to term. Supplementation of folic acid led to an increase of single CD56(pos) cells, but cytotoxic function appeared unaffected. The expression pattern of killer inhibitory receptors on CD56(pos) cells was not different between patients and controls. A longitudinal study revealed that, compared with controls, in RSA women higher numbers of double CD56(pos)CD16(pos) cells were present during early pregnancy, paralleled by an increase in cytotoxic NK cell reactivity. The single CD56(pos) population decreased in number. In conclusion, the analysis of peripheral NK cell characteristics appears a suitable diagnostic tool in RSA. Immunomodulation aimed at NK cell function appears a promising therapeutic measure.

Prospective guidance in a free-swimming cell.

PubMed

Delafield-Butt, Jonathan T; Pepping, Gert-Jan; McCaig, Colin D; Lee, David N

2012-07-01

A systems theory of movement control in animals is presented in this article and applied to explaining the controlled behaviour of the single-celled Paramecium caudatum in an electric field. The theory-General Tau Theory-is founded on three basic principles: (i) all purposive movement entails prospectively controlling the closure of action-gaps (e.g. a distance gap when reaching, or an angle gap when steering); (ii) the sole informational variable required for controlling gaps is the relative rate of change of the gap (the time derivative of the gap size divided by the size), which can be directly sensed; and (iii) a coordinated movement is achieved by keeping the relative rates of change of gaps in a constant ratio. The theory is supported by studies of controlled movement in mammals, birds and insects. We now show for the first time that it is also supported by single-celled paramecia steering to the cathode in a bi-polar electric field. General Tau Theory is deployed to explain this guided steering by the cell. This article presents the first computational model of prospective perceptual control in a non-neural, single-celled system.

Low CD4+ T-cell levels and B-cell apoptosis in vertically HIV-exposed noninfected children and adolescents.

PubMed

Miyamoto, Maristela; Pessoa, Silvana D; Ono, Erika; Machado, Daisy M; Salomão, Reinaldo; Succi, Regina C de M; Pahwa, Savita; de Moraes-Pinto, Maria Isabel

2010-12-01

Lymphocyte subsets, activation markers and apoptosis were assessed in 20 HIV-exposed noninfected (ENI) children born to HIV-infected women who were or not exposed to antiretroviral (ARV) drugs during pregnancy and early infancy. ENI children and adolescents were aged 6-18 years and they were compared to 25 age-matched healthy non-HIV-exposed children and adolescents (Control). ENI individuals presented lower CD4(+) T cells/mm(3) than Control group (control: 1120.3 vs. ENI: 876.3; t-test, p = 0.030). ENI individuals had higher B-cell apoptosis than Control group (Control: 36.6%, ARV exposed: 82.3%, ARV nonexposed: 68.5%; Kruskal-Wallis, p < 0.05), but no statistical difference was noticed between those exposed and not exposed to ARV. Immune activation in CD4(+) T, CD8(+) T and in B cells was comparable in ENI and in Control children and adolescents. Subtle long-term immune alterations might persist among ENI individuals, but the clinical consequences if any are unknown, and these children require continued monitoring.

Increasing anthraquinone production by overexpression of 1-deoxy-D: -xylulose-5-phosphate synthase in transgenic cell suspension cultures of Morinda citrifolia.

PubMed

Quevedo, Carla; Perassolo, María; Alechine, Eugenia; Corach, Daniel; Giulietti, Ana María; Talou, Julián Rodriguez

2010-07-01

A Morinda citrifolia cell line was obtained by overexpresion of 1-deoxy-D: -xylulose 5-phosphate synthase (DXS) from Catharanthus roseus, a key enzyme of the metabolic pathway of anthraquinones (AQs). This cell line increased AQs production by about 24% compared to the control cell line. This transgenic cell line which carries dxs cDNA isolated from Catharanthus roseus, was achieved by direct transformation of cell suspension cultures of M. citrifolia using a hypervirulent Agrobacterium tumefaciens strain. The effects of the overexpression of the dxs gene also resulted in increased levels of dxs mRNA transcripts and DXS activity compared to the control cell line. In addition, total phenolics and phenylalanine ammonia-lyase activity were evaluated and were significantly higher in the transgenic line than in controls.

Scratch2 prevents cell cycle re-entry by repressing miR-25 in postmitotic primary neurons.

PubMed

Rodríguez-Aznar, Eva; Barrallo-Gimeno, Alejandro; Nieto, M Angela

2013-03-20

During the development of the nervous system the regulation of cell cycle, differentiation, and survival is tightly interlinked. Newly generated neurons must keep cell cycle components under strict control, as cell cycle re-entry leads to neuronal degeneration and death. However, despite their relevance, the mechanisms controlling this process remain largely unexplored. Here we show that Scratch2 is involved in the control of the cell cycle in neurons in the developing spinal cord of the zebrafish embryo. scratch2 knockdown induces postmitotic neurons to re-enter mitosis. Scratch2 prevents cell cycle re-entry by maintaining high levels of the cycle inhibitor p57 through the downregulation of miR-25. Thus, Scratch2 appears to safeguard the homeostasis of postmitotic primary neurons by preventing cell cycle re-entry.

HIV Controllers Exhibit Enhanced Frequencies of Major Histocompatibility Complex Class II Tetramer+ Gag-Specific CD4+ T Cells in Chronic Clade C HIV-1 Infection

PubMed Central

Laher, Faatima; Ranasinghe, Srinika; Porichis, Filippos; Mewalal, Nikoshia; Pretorius, Karyn; Ismail, Nasreen; Buus, Søren; Stryhn, Anette; Carrington, Mary; Walker, Bruce D.; Ndung'u, Thumbi

2017-01-01

ABSTRACT Immune control of viral infections is heavily dependent on helper CD4+ T cell function. However, the understanding of the contribution of HIV-specific CD4+ T cell responses to immune protection against HIV-1, particularly in clade C infection, remains incomplete. Recently, major histocompatibility complex (MHC) class II tetramers have emerged as a powerful tool for interrogating antigen-specific CD4+ T cells without relying on effector functions. Here, we defined the MHC class II alleles for immunodominant Gag CD4+ T cell epitopes in clade C virus infection, constructed MHC class II tetramers, and then used these to define the magnitude, function, and relation to the viral load of HIV-specific CD4+ T cell responses in a cohort of untreated HIV clade C-infected persons. We observed significantly higher frequencies of MHC class II tetramer-positive CD4+ T cells in HIV controllers than progressors (P = 0.0001), and these expanded Gag-specific CD4+ T cells in HIV controllers showed higher levels of expression of the cytolytic proteins granzymes A and B. Importantly, targeting of the immunodominant Gag41 peptide in the context of HLA class II DRB1*1101 was associated with HIV control (r = −0.5, P = 0.02). These data identify an association between HIV-specific CD4+ T cell targeting of immunodominant Gag epitopes and immune control, particularly the contribution of a single class II MHC-peptide complex to the immune response against HIV-1 infection. Furthermore, these results highlight the advantage of the use of class II tetramers in evaluating HIV-specific CD4+ T cell responses in natural infections. IMPORTANCE Increasing evidence suggests that virus-specific CD4+ T cells contribute to the immune-mediated control of clade B HIV-1 infection, yet there remains a relative paucity of data regarding the role of HIV-specific CD4+ T cells in shaping adaptive immune responses in individuals infected with clade C, which is responsible for the majority of HIV infections worldwide. Understanding the contribution of HIV-specific CD4+ T cell responses in clade C infection is particularly important for developing vaccines that would be efficacious in sub-Saharan Africa, where clade C infection is dominant. Here, we employed MHC class II tetramers designed to immunodominant Gag epitopes and used them to characterize CD4+ T cell responses in HIV-1 clade C infection. Our results demonstrate an association between the frequency of HIV-specific CD4+ T cell responses targeting an immunodominant DRB1*11-Gag41 complex and HIV control, highlighting the important contribution of a single class II MHC-peptide complex to the immune response against HIV-1 infections. PMID:28077659

Chronic heart failure and aging - effects of exercise training on endothelial function and mechanisms of endothelial regeneration: Results from the Leipzig Exercise Intervention in Chronic heart failure and Aging (LEICA) study.

PubMed

Sandri, Marcus; Viehmann, Manuel; Adams, Volker; Rabald, Kristin; Mangner, Norman; Höllriegel, Robert; Lurz, Philipp; Erbs, Sandra; Linke, Axel; Kirsch, Katharina; Möbius-Winkler, Sven; Thiery, Joachim; Teupser, Daniel; Hambrecht, Rainer; Schuler, Gerhard; Gielen, Stephan

2016-03-01

A reduction in number and function of endothelial progenitor cells (EPCs) occurs in both physiologic aging and chronic heart failure (CHF). We assessed whether disease and aging have additive effects on EPCs or whether beneficial effects of exercise training are diminished in old age. We randomized 60 patients with stable CHF and 60 referent controls to a training or a control group. To detect possible aging effects we included subjects below 55 (young) and above 65 years (older). Subjects in the training group exercised four times daily at 60% to 70% of VO2max for four weeks under supervision. At baseline and after the intervention the number and function of EPCs were assessed. As compared with young referent controls, older referent controls showed at baseline a reduced EPC number (young: 190 ± 37 CD34/KDR positive cells/ml blood; older: 131 ± 26 CD34/KDR positive cells/ml blood; p < 0.05) and function (young: 230 ± 41 migrated cells/1000 plated cells; older: 185 ± 28 cells/1000 plated cells; p < 0.05). In young and older CHF patients EPC-number (young: 85 ± 21 CD34/KDR positive cells/ml blood; older: 78 ± 20 CD34/KDR positive cells/ml blood) and EPC-function (young: 113 ± 26 cells/1000 plated cells; older: 120 ± 27 cells/1000 plated cells) were impaired. As a result of exercise training, EPC function improved by 24% in older referent controls (p < 0.05), while it remained unchanged in young training referent controls and controls respectively. In young and older patients with CHF four weeks of exercise training resulted in a significant improvement in EPC numbers and EPC function (young: number +66% function +43%; p < 0.05; older: number +69% function +36%; p < 0.05). These results were accompanied by a significant increase in flow mediated dilatation in the training groups of young/older CHF patients and in older referent controls. Four weeks of exercise training are effective in improving EPC number and EPC function in CHF patients. These training effects were not impaired among older patients, emphasizing the potentials of rehabilitation interventions in a patient group where CHF has a high prevalence. © The European Society of Cardiology 2015.

Piwi Is Required in Multiple Cell Types to Control Germline Stem Cell Lineage Development in the Drosophila Ovary

PubMed Central

Ma, Xing; Wang, Su; Do, Trieu; Song, Xiaoqing; Inaba, Mayu; Nishimoto, Yoshiya; Liu, Lu-ping; Gao, Yuan; Mao, Ying; Li, Hui; McDowell, William; Park, Jungeun; Malanowski, Kate; Peak, Allison; Perera, Anoja; Li, Hua; Gaudenz, Karin; Haug, Jeff; Yamashita, Yukiko; Lin, Haifan; Ni, Jian-quan; Xie, Ting

2014-01-01

The piRNA pathway plays an important role in maintaining genome stability in the germ line by silencing transposable elements (TEs) from fly to mammals. As a highly conserved piRNA pathway component, Piwi is widely expressed in both germ cells and somatic cells in the Drosophila ovary and is required for piRNA production in both cell types. In addition to its known role in somatic cap cells to maintain germline stem cells (GSCs), this study has demonstrated that Piwi has novel functions in somatic cells and germ cells of the Drosophila ovary to promote germ cell differentiation. Piwi knockdown in escort cells causes a reduction in escort cell (EC) number and accumulation of undifferentiated germ cells, some of which show active BMP signaling, indicating that Piwi is required to maintain ECs and promote germ cell differentiation. Simultaneous knockdown of dpp, encoding a BMP, in ECs can partially rescue the germ cell differentiation defect, indicating that Piwi is required in ECs to repress dpp. Consistent with its key role in piRNA production, TE transcripts increase significantly and DNA damage is also elevated in the piwi knockdown somatic cells. Germ cell-specific knockdown of piwi surprisingly causes depletion of germ cells before adulthood, suggesting that Piwi might control primordial germ cell maintenance or GSC establishment. Finally, Piwi inactivation in the germ line of the adult ovary leads to gradual GSC loss and germ cell differentiation defects, indicating the intrinsic role of Piwi in adult GSC maintenance and differentiation. This study has revealed new germline requirement of Piwi in controlling GSC maintenance and lineage differentiation as well as its new somatic function in promoting germ cell differentiation. Therefore, Piwi is required in multiple cell types to control GSC lineage development in the Drosophila ovary. PMID:24658126

Epithelial-mesenchymal transition transcription factors control pluripotent adult stem cell migration in vivo in planarians

PubMed Central

Abnave, Prasad; Aboukhatwa, Ellen; Kosaka, Nobuyoshi; Thompson, James; Hill, Mark A.

2017-01-01

Migration of stem cells underpins the physiology of metazoan animals. For tissues to be maintained, stem cells and their progeny must migrate and differentiate in the correct positions. This need is even more acute after tissue damage by wounding or pathogenic infection. Inappropriate migration also underpins metastasis. Despite this, few mechanistic studies address stem cell migration during repair or homeostasis in adult tissues. Here, we present a shielded X-ray irradiation assay that allows us to follow stem cell migration in planarians. We demonstrate the use of this system to study the molecular control of stem cell migration and show that snail-1, snail-2 and zeb-1 EMT transcription factor homologs are necessary for cell migration to wound sites and for the establishment of migratory cell morphology. We also observed that stem cells undergo homeostatic migration to anterior regions that lack local stem cells, in the absence of injury, maintaining tissue homeostasis. This requires the polarity determinant notum. Our work establishes planarians as a suitable model for further in-depth study of the processes controlling stem cell migration in vivo. PMID:28893948

Chrna2-Martinotti Cells Synchronize Layer 5 Type A Pyramidal Cells via Rebound Excitation

PubMed Central

Leão, Richardson N.; Edwards, Steven J.

2017-01-01

Martinotti cells are the most prominent distal dendrite–targeting interneurons in the cortex, but their role in controlling pyramidal cell (PC) activity is largely unknown. Here, we show that the nicotinic acetylcholine receptor α2 subunit (Chrna2) specifically marks layer 5 (L5) Martinotti cells projecting to layer 1. Furthermore, we confirm that Chrna2-expressing Martinotti cells selectively target L5 thick-tufted type A PCs but not thin-tufted type B PCs. Using optogenetic activation and inhibition, we demonstrate how Chrna2-Martinotti cells robustly reset and synchronize type A PCs via slow rhythmic burst activity and rebound excitation. Moreover, using optical feedback inhibition, in which PC spikes controlled the firing of surrounding Chrna2-Martinotti cells, we found that neighboring PC spike trains became synchronized by Martinotti cell inhibition. Together, our results show that L5 Martinotti cells participate in defined cortical circuits and can synchronize PCs in a frequency-dependent manner. These findings suggest that Martinotti cells are pivotal for coordinated PC activity, which is involved in cortical information processing and cognitive control. PMID:28182735

Design of a Single-Cell Positioning Controller Using Electroosmotic Flow and Image Processing

PubMed Central

Ay, Chyung; Young, Chao-Wang; Chen, Jhong-Yin

2013-01-01

The objective of the current research was not only to provide a fast and automatic positioning platform for single cells, but also improved biomolecular manipulation techniques. In this study, an automatic platform for cell positioning using electroosmotic flow and image processing technology was designed. The platform was developed using a PCI image acquisition interface card for capturing images from a microscope and then transferring them to a computer using human-machine interface software. This software was designed by the Laboratory Virtual Instrument Engineering Workbench, a graphical language for finding cell positions and viewing the driving trace, and the fuzzy logic method for controlling the voltage or time of an electric field. After experiments on real human leukemic cells (U-937), the success of the cell positioning rate achieved by controlling the voltage factor reaches 100% within 5 s. A greater precision is obtained when controlling the time factor, whereby the success rate reaches 100% within 28 s. Advantages in both high speed and high precision are attained if these two voltage and time control methods are combined. The control speed with the combined method is about 5.18 times greater than that achieved by the time method, and the control precision with the combined method is more than five times greater than that achieved by the voltage method. PMID:23698272

Differential expression of pro-inflammatory cytokines in intra-epithelial T cells between trachea and bronchi distinguishes severity of COPD.

PubMed

Hodge, Greg; Reynolds, Paul N; Holmes, Mark; Hodge, Sandra

2012-12-01

Measuring T-cell production of intracellular cytokines by flow cytometry enables specific monitoring of airway inflammation and response to therapies in chronic lung diseases including chronic obstructive pulmonary disease (COPD). We have previously shown that T cells in the airways of ex- and current- smoker COPD patients and healthy smokers produce increased T-cell pro-inflammatory cytokines IFNγ and TNFα versus healthy controls. However, we could not differentiate between COPD groups and smokers due to a high degree of inter-patient variability. To address this limitation, we hypothesized that intraepithelial T cells obtained from brushings of trachea may serve as an ideal intra-patient control compared with cells obtained from left and right bronchi. Production of intracellular cytokines by intraepithelial T-cells obtained from trachea and right and left bronchi from 26 individuals with COPD (16 with GOLD I and 10 with GOLD II-III disease), 11 healthy controls and 8 smokers was measured by flow cytometry. There was a significant increase in intraepithelial T-cell IFNγ and TNFα in both right and left bronchi of GOLD II-III COPD patients compared to cells obtained from the trachea. There were no changes in T cell pro-inflammatory cytokines between the bronchi and trachea from control subjects, GOLD I COPD patients or healthy smokers. There was a significant negative correlation between increased intraepithelial IFNγ and TNFα in bronchial brushing T-cells compared with tracheal T-cells, and compared with FEV1. Monitoring intracellular intra-epithelial T-cell cytokine production in bronchial brushings using autologous tracheal brushings as controls provides improves the sensitivity of the technique. Therapeutic targeting of these pro-inflammatory cytokines and assessing the effects of drugs on immune reactivity has the potential to reduce lung inflammation caused by intra-epithelial T cells in COPD. Copyright © 2012 Elsevier Ltd. All rights reserved.

Transplantation of dedifferentiated fat cell-derived micromass pellets contributed to cartilage repair in the rat osteochondral defect model.

PubMed

Shimizu, Manabu; Matsumoto, Taro; Kikuta, Shinsuke; Ohtaki, Munenori; Kano, Koichiro; Taniguchi, Hiroaki; Saito, Shu; Nagaoka, Masahiro; Tokuhashi, Yasuaki

2018-03-20

Mature adipocyte-derived dedifferentiated fat (DFAT) cells possesses the ability to proliferate effectively and the potential to differentiate into multiple linages of mesenchymal tissue; similar to adipose-derived stem cells (ASCs). The purpose of this study is to examine the effects of DFAT cell transplantation on cartilage repair in a rat model of osteochondral defects. Full-thickness osteochondral defects were created in the knees of Sprague-Dawley rats bilaterally. Cartilage-like micromass pellets were prepared from green fluorescent protein (GFP)-labeled rat DFAT cells and subsequently transplanted into the affected right knee of these rats. Defects in the left knee were used as a control. Macroscopic and microscopic changes of treated and control defects were evaluated up to 12 weeks post-treatment with DFAT cells. To observe the transplanted cells, sectioned femurs were immunostained for GFP and type II collagen. DFAT cells formed micromass pellets expressing characteristics of immature cartilage in vitro. In the DFAT cell-transplanted limbs, the defects were completely filled with white micromass pellets as early as 2 weeks post-treatment. These limbs became smooth at 4 weeks. Conversely, the defects in the control limbs were still not repaired by 4 weeks. Macroscopic ICRS scores at 2 and 4 weeks were significantly higher in the DFAT cells-transplanted limbs compared to those of the control limbs. The modified O'Driscol histological scores for the DFAT cell-transplanted limbs were significantly higher than those of the control limbs at corresponding time points. GFP-positive DAFT cells were detected in the transplanted area at 2 weeks but hardly visible at 12 weeks post-operation. Transplantation of DFAT cell-derived micromass pellets contribute to cartilage repair in a rat osteochondral defect model. DFAT cell transplantation may be a viable therapeutic strategy for the repair of osteochondral injuries. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Effect of Anger Patterns and Depression on Serum IgA and NK Cell Frequency.

PubMed

Farnam, Alireza; Majidi, Jafar; Nourazar, Seyyed Gholamreza; Ghojazadeh, Morteza; Movassaghpour, Aliakbar; Zolbanin, Saeedeh Majidi

2016-03-01

There are conflicting findings about relationship between depression and anger with immunological parameters. To investigate the relationship between anger patterns and immune system in depressed patients. Thirty-five patients with major depressive disorder were selected according to DSM-IV criteria. The Hamilton Depression Scale and Spielberger Anger questionnaires were used to determine severity of depression and "anger expression pattern", respectively. The control group without a previous history of mental illness was also selected. In the group of patients with moderate depression, serum IgA levels and NK cell percentage were measured. Mean differences of all types of "anger expression pattern", including; "state-trait anger", "anger expression out", "anger expression in", "anger control out" and "anger control in", between study and control groups, were statistically significant (p<0.05). Difference in mean serum levels of IgA in either group was not significant (p=0.9), but the mean difference was significant in terms of NK-cell percentage in both groups (p=0.04). There was no significant relationship between IgA levels and percentage of NK- cell with all types of "anger expression pattern" in both groups. Only in the control group, IgA had significant correlation with anger control out (p=0.04). Moderately depressed patients versus control group had higher Spielberger scores in all types of anger expression pattern except anger control-out and anger control-in. We found no evidence supporting the relationship between" anger expression pattern" and IgA levels and NK cell percentage; however, it seems that depression itself causes reduced number of NK cells and increased IgA levels.

TCPs, WUSs, and WINDs: families of transcription factors that regulate shoot meristem formation, stem cell maintenance, and somatic cell differentiation.

PubMed

Ikeda, Miho; Ohme-Takagi, Masaru

2014-01-01

In contrast to somatic mammalian cells, which cannot alter their fate, plant cells can dedifferentiate to form totipotent callus cells and regenerate a whole plant, following treatment with specific phytohormones. However, the regulatory mechanisms and key factors that control differentiation-dedifferentiation and cell totipotency have not been completely clarified in plants. Recently, several plant transcription factors that regulate meristem formation and dedifferentiation have been identified and include members of the TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP), WUSCHEL (WUS), and WOUND INDUCED DEDIFFERENTIATION (WIND1) families. WUS and WIND positively control plant cell totipotency, while TCP negatively controls it. Interestingly, TCP is a transcriptional activator that acts as a negative regulator of shoot meristem formation, and WUS is a transcriptional repressor that positively maintains totipotency of the stem cells of the shoot meristem. We describe here the functions of TCP, WUS, and WIND transcription factors in the regulation of differentiation-dedifferentiation by positive and negative transcriptional regulators.

A Theoretical Solid Oxide Fuel Cell Model for Systems Controls and Stability Design

NASA Technical Reports Server (NTRS)

Kopasakis, George; Brinson, Thomas; Credle, Sydni

2008-01-01

As the aviation industry moves toward higher efficiency electrical power generation, all electric aircraft, or zero emissions and more quiet aircraft, fuel cells are sought as the technology that can deliver on these high expectations. The hybrid solid oxide fuel cell system combines the fuel cell with a micro-turbine to obtain up to 70% cycle efficiency, and then distributes the electrical power to the loads via a power distribution system. The challenge is to understand the dynamics of this complex multidiscipline system and the design distributed controls that take the system through its operating conditions in a stable and safe manner while maintaining the system performance. This particular system is a power generation and a distribution system, and the fuel cell and micro-turbine model fidelity should be compatible with the dynamics of the power distribution system in order to allow proper stability and distributed controls design. The novelty in this paper is that, first, the case is made why a high fidelity fuel cell mode is needed for systems control and stability designs. Second, a novel modeling approach is proposed for the fuel cell that will allow the fuel cell and the power system to be integrated and designed for stability, distributed controls, and other interface specifications. This investigation shows that for the fuel cell, the voltage characteristic should be modeled but in addition, conservation equation dynamics, ion diffusion, charge transfer kinetics, and the electron flow inherent impedance should also be included.

Reduced response to Epstein–Barr virus antigens by T-cells in systemic lupus erythematosus patients

PubMed Central

Draborg, Anette Holck; Jacobsen, Søren; Westergaard, Marie; Mortensen, Shila; Larsen, Janni Lisander; Houen, Gunnar; Duus, Karen

2014-01-01

Objective Epstein–Barr virus (EBV) has for long been associated with systemic lupus erythematosus (SLE). In this study, we investigated the levels of latent and lytic antigen EBV-specific T-cells and antibodies in SLE patients. Methods T cells were analyzed by flow cytometry and antibodies were analyzed by enzyme-linked immunosorbent assay. Results SLE patients showed a significantly reduced number of activated (CD69) T-cells upon ex vivo stimulation with EBV nuclear antigen (EBNA) 1 or EBV early antigen diffuse (EBV-EA/D) in whole blood samples compared with healthy controls. Also, a reduced number of T-cells from SLE patients were found to produce interferon-γ upon stimulation with these antigens. Importantly, responses to a superantigen were normal in SLE patients. Compared with healthy controls, SLE patients had fewer EBV-specific T-cells but higher titres of antibodies against EBV. Furthermore, an inverse correlation was revealed between the number of lytic antigen EBV-specific T-cells and disease activity of the SLE patients, with high-activity SLE patients having fewer T-cells than low-activity SLE patients. Conclusions These results indicate a limited or a defective EBV-specific T-cell response in SLE patients, which may suggest poor control of EBV infection in SLE with an immune reaction shift towards a humoral response in an attempt to control viral reactivation. A role for decreased control of EBV as a contributing agent in the development or exacerbation of SLE is proposed. PMID:25396062

In-vivo neutrophil migration and nitroblue tetrazolium reduction in sickle cell disease.

PubMed

Akinyanju, O O

1985-01-01

In order to determine the contribution of neutrophil malfunction to the phenomenon of enhanced susceptibility of sickle cell disease patients to bacterial infection, the in-vivo neutrophil migration capacity in 23 sickle cell patients and in 14 normal controls; and the neutrophil reduction of nitroblue tetrazolium dye in 74 sickle cell patients and in 78 normal controls were studied. Secondarily the usefulness of the NBT test in distinguishing between osteomyelitis and uncomplicated bone pain was examined. No impairment of neutrophil migratory capacity was evident as no significant difference was observed between the mean migrated neutrophil count in the sickle cell subjects (1.99 X 10(9)/1) and that in normal controls (2.08 X 10(9)/1). The mean NBT scores were 19.9 +/- 8.9% in non-infected controls and 41.3 +/- 14.6% in infected controls (P less than 0.001). In sickle cell disease they were 23.6 +/- 6% in steady state subjects, 29.2 +/- 16.4% in sterile painful crises, 42.9 +/- 15% in non-osteomyelitic bacterial infection (P less than 0.001) and 18.9 +/- 4.2% during osteomyelitis. Thus all sickle cell subjects apart from those with osteomyelitis showed significant increases in the NBT scores during bacterial infection. The low score in sickle cell osteomyelitis is possibly associated with a relative neutrophil phagocytic defect which requires further elucidation. The NBT test was not useful in distinguishing uncomplicated painful crisis from early osteomyelitis in sickle cell disease.

Proliferation of protease-enriched mast cells in sarcoptic skin lesions of raccoon dogs.

PubMed

Noviana, D; W Harjanti, D; Otsuka, Y; Horii, Y

2004-07-01

Skin sites, tongue, lung, liver, jejunum and rectum from two raccoon dogs with Sarcoptes scabiei infestation and five normal (control) raccoon dogs were examined in terms of the distribution, proteoglycan properties and protease activity of mast cells. Infestation with S. scabiei caused a significant increase in the number of dermal mast cells. While the number of mast cells (average +/- standard deviation) in specimens of skin from the dorsum, dorsal neck, dorsal hind foot and dorsal fore foot was 40.0 +/- 19.8/mm2 in control animals, it was 236.1 +/- 58.9/mm2 in the skin of mange-infested animals. Histochemical analysis revealed the glycosaminoglycan, heparin, within the mast cells of all organs examined in both control and affected animals. Enzyme-histochemical detection of serine proteases demonstrated an increase in mast-cell-specific protease activity (i.e., chymase and tryptase) in the skin of infested animals. The percentage of mast cells demonstrating chymase activity was 53.0 +/- 27.4% in control animals and 73.8 +/- 19.4% in mite-infested animals. The corresponding results for tryptase activity were 53.5 +/- 25.2% and 89.4 +/- 9.8%. Increases in mast cell chymase or tryptase activity, or both, were also observed within other organs of the infected animals, but the total number of mast cells found at such sites (with the exception of liver and ventrolateral pinna) did not differ from those of control animals. Copyright 2004 Elsevier Ltd.

Clinical performance of stem cell therapy in patients with acute-on-chronic liver failure: a systematic review and meta-analysis.

PubMed

Xue, Ran; Meng, Qinghua; Dong, Jinling; Li, Juan; Yao, Qinwei; Zhu, Yueke; Yu, Hongwei

2018-05-10

Stem cell therapy has been applied in the treatment of acute-on-chronic liver failure (ACLF). However, its clinical efficiency is still debatable. The aim of this systematic review and meta-analysis is to evaluate the clinical efficiency of stem cell therapy in the treatment of ACLF. The Cochrane Library, OVID, EMBASE, and PUBMED were searched to December 2017. Both randomized and non-randomized studies, assessing stem cell therapy in patients with ACLF, were included. The outcome measures were total bilirubin (TBIL), alanine transaminase (ALT), international normalized ratio (INR), albumin (ALB), and the model for end-stage liver disease (MELD) score. The quality of evidence was assessed by GRADEpro. Four randomized controlled trials and six non-randomized controlled trials were included. The TBIL levels significantly decreased at 1-, 3-, 12-month after the stem cell therapy (p = 0.0008; p = 0.04; p = 0.007). The ALT levels decreased significantly compared with the control group in the short-term (p < 0.00001). There was no obvious change in the INR level compared with the control groups (p = 0.64). The ALB levels increased markedly as compared with the control groups (p < 0.0001). The significant difference can be found in MELD score between stem cell therapy and control groups (p = 0.008). Further subgroup analysis for 3-month clinical performance according to the stem cell types have also been performed. This study suggests that the clinical outcomes of stem cell therapy were satisfied in patients with ACLF in the short-term. MSCs may be better than BM-MNCs in the stem cells transplantation of ACLF. However, more attention should focus on clinical trials in large-volume centers.

Dact2 is expressed in the developing ureteric bud/collecting duct system of the kidney and controls morphogenetic behavior of collecting duct cells.

PubMed

Lee, Wen-Chin; Hough, Melinda T; Liu, Weijia; Ekiert, Robert; Lindström, Nils O; Hohenstein, Peter; Davies, Jamie A

2010-10-01

The overall pattern of the developing kidney is set in large part by the developing ureteric bud/collecting duct system, and dysgenesis of this system accounts for a variety of clinically significant renal diseases. Understanding how the behavior of cells in the developing ureteric bud/collecting duct is controlled is therefore important to understanding the normal and abnormal kidney. Dact proteins have recently been identified as cytoplasmic regulators of intracellular signaling. Dact1 inhibits Wnt signaling, and Dact2 inhibits transforming growth factor (TGF)-β signaling. Here, we report that Dact2 is expressed in developing and adult mouse kidneys, specifically in the ureteric bud/collecting duct epithelium, a structure whose morphogenesis is controlled partially by TGF-β. When small interfering RNA is used to knock down Dact2 expression in collecting duct cells, they show some constitutive phospho-Smad2, undetectable in controls, and elevated phospho-Smad2 in response to TGF-β. They also show defective migration and, in a monolayer wound-healing assay, they fail to assemble a leading edge "cable" of actomyosin and advance instead as a disorganized mass of lamellipodium-bearing cells. This effect is seriously exacerbated by exogenous TGF-β, although control cells tolerate it well. In three-dimensional culture, Dact2 knockdown cells form cysts and branching tubules, but the outlines of the cysts made by knockdown cells are ragged rather than smooth and the branching tubules are decorated with many fine spikes not seen in controls. These data suggest Dact2 plays a role in regulating morphogenesis by renal collecting duct cells, probably by protecting cells from overly strong TGF-β pathway activation.

Increased TERC gene copy number and cells in senescence in primary sclerosing cholangitis compared to colitis and control patients.

PubMed

Laish, Ido; Katz, Hila; Sulayev, Yael; Liberman, Meytal; Naftali, Timna; Benjaminov, Fabiana; Stein, Assaf; Kitay-Cohen, Yona; Biron-Shental, Tal; Konikoff, Fred; Amiel, Aliza

2013-10-25

Primary sclerosing cholangitis (PSC) is a chronic cholestatic disorder that involves inflammatory and fibrotic changes in the bile ducts. Up to 80% of patients have concomitant inflammatory bowel disease (IBD) with colitis. PSC patients are predisposed to develop hepatobiliary, colonic and other extrahepatic malignancies, probably related to inflammatory processes that might promote carcinogenesis. Telomerase is an enzyme complex that lengthens telomeres and has enhanced expression in numerous malignancies. In this study, we evaluated the TERC gene copy number, the proportion of cells in senescence and the amount of fragmentation in the senescent state. Fluorescence in situ hybridization (FISH) for the TERC gene was applied to lymphocytes retrieved from PSC (N=19), colitis (N=20) and healthy control patients (N=20) to determine the TERC copy number. On the same FISH slides, cells stained with DAPI were also analyzed for senescence-associated heterochromatin foci (SAHF) status, including the number of cells with fragments and the number of SAHF fragments in each cell. A higher TERC gene copy number was observed in cells from PSC patients compared to colitis and control group patients. It was also higher in the colitis than in the control group. Significantly more cells in the senescent state and more fragmentation in each cell were observed in the PSC group compared to colitis and control groups. The TERC gene copy number and the number of cells in the senescent state were increased in PSC patients compared to the colitis and control groups. These findings are probably related to the genetic instability parameters that reflect the higher tendency of this patient group to develop malignancies. © 2013.

The ratio of cancer cells to stroma after induction therapy in the treatment of non-small cell lung cancer.

PubMed

Goto, Masaki; Naito, Masahito; Saruwatari, Koichi; Hisakane, Kakeru; Kojima, Motohiro; Fujii, Satoshi; Kuwata, Takeshi; Ochiai, Atsushi; Nomura, Shogo; Aokage, Keiju; Hishida, Tomoyuki; Yoshida, Junji; Yokoi, Kohei; Tsuboi, Masahiro; Ishii, Genichiro

2017-02-01

Induction therapy induces degenerative changes of various degrees in both cancerous and non-cancerous cells of non-small cell lung cancer (NSCLC). The effect of induction therapy on histological characteristics, in particular the ratio of residual cancer cells to non-cancerous components, is unknown. Seventy-four NSCLC patients treated with induction therapy followed by surgery were enrolled. Residual cancer cells were identified using anti-pan-cytokeratin antibody (AE1/AE3). We analyzed and quantified the following three factors via digital image analysis; (1) the tumor area containing cancer cells and non-cancerous components (TA), (2) the total area of AE1/AE3 positive cancer cells (TACC), (3) the percentage of TACC to TA (%TACC). These factors were also analyzed in a matched control group (surgery alone, n = 80). The median TACC of the induction therapy group was significantly lower than that of the control group (p < 0.01). In addition, the median %TACC of the induction therapy group (5.9 %) was significantly lower than that of the control group (58.6 %) (p < 0.01). TACC had a strong positive correlation with TA in the control group (r = 0.93), but not in the induction therapy group. Conversely, TACC had a strong positive correlation with %TACC in the induction therapy group (r = 0.95), but not in the control group. Unlike the control group, the smaller the total area of residual cancer cells, the higher residual tumor contained non-cancerous components in the induction group, which may be the characteristic histological feature of NSCLC after induction therapy.

Characterization of dendritic cells in lip and oral cavity squamous cell carcinoma.

PubMed

Costa, Nádia Lago; Gonçalves, Andréia Souza; Martins, Allisson Filipe Lopes; Arantes, Diego Antônio Costa; Silva, Tarcília Aparecida; Batista, Aline Carvalho

2016-07-01

There may be differences in the antitumor immunity induced by dendritic cells (DCs) during the development of squamous cell carcinoma (SCC) located in the lip rather than in the oral cavity. The aim of this study was to evaluate the number of immature and mature DCs in SCC and potentially malignant disorders of the oral cavity and lip. Immunohistochemistry was used to identify the number (cells/mm(2) ) of immature (CD1a(+) ) or mature (CD83(+) ) DCs in samples of oral cavity SCC (OCSCC) (n = 39), lip SCC (LSCC) (n = 23), leukoplakia (LK) (n = 21), actinic cheilitis (AC) (n = 13), and normal mucosa of the oral cavity (OC control, n = 12) and the lip (lip control, n = 11). The number of CD1a(+) cells tended to be higher in the OC control samples compared with the LK (P = 0.04) and OCSCC (P = 0.21). Unlike, this cell population was lower in the lip control than in AC or LSCC (P < 0.05). The number of CD83(+) cells was increased in the LSCC samples compared with the AC and lip control (P = 0.0001) and in OCSCC compared with both the LK (P = 0.001) and OC control (P = 0.0001) samples. LSCC showed an elevated number of CD1a(+) and CD83(+) cells compared with OCSCC (P = 0.03). The population of mature DCs was lower than the population of immature DCs in all of the tested groups (P < 0.05). There were a greater number of both mature and immature DC populations in the LSCC samples than in the OCSCC, which could contribute to establishing a more effective immune antitumor response for this neoplasm. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

[A preliminary study for the effect of nano hydroxyapatite on human adipose-derived mesenchymal stem cells mixture 3D bio-printing].

PubMed

Song, Y; Wang, X F; Wang, Y G; Dong, F; Lv, P J

2016-10-18

To study the effect of nano hydroxyapatite on human adipose-derived mesenchymal stem cells(hASCs) mixture 3D bio-printing for cells' proliferation and osteogenesis. P5 hASCs were used as seed cells, 10 g/L nano hydroxyapatite was added into the cell-sodium alginate-gelatin mixture (concentration: 20 g/L sodium alginate, 80 g/L gelatin; cell density: 1×10 6 /mL), then the mixture was printed by 3D bio-printer as the experimental group. And the cell-sodium alginate-gelatin mixture without nano hydroxyapatite was printed as the control group. Respectively, both the experimental and control groups were detected by microscope, CCK-8, Western blot and PCR at certain time pointsafter being printed, whose cells' proliferation and osteogenic differentiation were analyzed. The microscopic observation and CCK-8 results showed that the cells of the experimental group and the control group both had a good proliferation 24 h and 7 d after being printed. The Western blot results showed that 14 d after printing, the expression of Runt-related transcription factor 2 (RUNX2) had no statistical difference between the experimental group and control group. The PCR results showed that 14 d after printing, the expression of osteogenesis-related genes (RUNX2, osterix, and osteocalcin) was significantly higher in the experimental group than in the control group. Nano hydroxyapatite can increase osteogenic differentiation of the hASCs mixture after bio-printing, in which the cells still have a good proliferation.

Presence of Epstein-Barr virus-infected B lymphocytes with thyrotropin receptor antibodies on their surface in Graves' disease patients and in healthy individuals.

PubMed

Nagata, Keiko; Higaki, Katsumi; Nakayama, Yuji; Miyauchi, Hiromi; Kiritani, Yui; Kanai, Kyosuke; Matsushita, Michiko; Iwasaki, Takeshi; Sugihara, Hirotsugu; Kuwamoto, Satoshi; Kato, Masako; Murakami, Ichiro; Nanba, Eiji; Kimura, Hiroshi; Hayashi, Kazuhiko

2014-05-01

Graves' disease is an autoimmune hyperthyroidism caused by thyrotropin receptor antibodies (TRAbs). Because Epstein-Barr virus (EBV) persists in B cells and is occasionally reactivated, we hypothesized that EBV contributes to TRAbs production in Graves' disease patients by stimulating the TRAbs-producing B cells. In order for EBV to stimulate antibody-producing cells, EBV must be present in those cells but that have not yet been observed. We examined whether EBV-infected (EBV(+)) B cells with TRAbs on their surface (TRAbs(+)) as membrane immunoglobulin were present in peripheral blood of Graves' disease patients. We analyzed cultured or non-cultured peripheral blood mononuclear cells (PBMCs) from 13 patients and 11 healthy controls by flow-cytometry and confocal laser microscopy, and confirmed all cultured PBMCs from 8 patients really had TRAbs(+) EBV(+) double positive cells. We unexpectedly detected TRAbs(+) cells in all healthy controls, and TRAbs(+) EBV(+) double positive cells in all cultured PBMC from eight healthy controls. The frequency of TRAbs(+) cells in cultured PBMCs was significantly higher in patients than in controls (p = 0.021). In this study, we indicated the presence of EBV-infected B lymphocytes with TRAbs on their surface, a possible player of the production of excessive TRAbs, the causative autoantibody for Graves' disease. This is a basic evidence for our hypothesis that EBV contributes to TRAbs production in Graves' disease patients. Our results further suggest that healthy controls have the potential for TRAbs production. This gives us an important insight into the pathogenesis of Graves' disease.

Presence of Epstein–Barr virus-infected B lymphocytes with thyrotropin receptor antibodies on their surface in Graves’ disease patients and in healthy individuals

PubMed Central

Nagata, Keiko; Higaki, Katsumi; Nakayama, Yuji; Miyauchi, Hiromi; Kiritani, Yui; Kanai, Kyosuke; Matsushita, Michiko; Iwasaki, Takeshi; Sugihara, Hirotsugu; Kuwamoto, Satoshi; Kato, Masako; Murakami, Ichiro; Nanba, Eiji; Kimura, Hiroshi; Hayashi, Kazuhiko

2014-01-01

Abstract Graves’ disease is an autoimmune hyperthyroidism caused by thyrotropin receptor antibodies (TRAbs). Because Epstein–Barr virus (EBV) persists in B cells and is occasionally reactivated, we hypothesized that EBV contributes to TRAbs production in Graves’ disease patients by stimulating the TRAbs-producing B cells. In order for EBV to stimulate antibody-producing cells, EBV must be present in those cells but that have not yet been observed. We examined whether EBV-infected (EBV(+)) B cells with TRAbs on their surface (TRAbs(+)) as membrane immunoglobulin were present in peripheral blood of Graves’ disease patients. We analyzed cultured or non-cultured peripheral blood mononuclear cells (PBMCs) from 13 patients and 11 healthy controls by flow-cytometry and confocal laser microscopy, and confirmed all cultured PBMCs from 8 patients really had TRAbs(+) EBV(+) double positive cells. We unexpectedly detected TRAbs(+) cells in all healthy controls, and TRAbs(+) EBV(+) double positive cells in all cultured PBMC from eight healthy controls. The frequency of TRAbs(+) cells in cultured PBMCs was significantly higher in patients than in controls (p = 0.021). In this study, we indicated the presence of EBV-infected B lymphocytes with TRAbs on their surface, a possible player of the production of excessive TRAbs, the causative autoantibody for Graves’ disease. This is a basic evidence for our hypothesis that EBV contributes to TRAbs production in Graves’ disease patients. Our results further suggest that healthy controls have the potential for TRAbs production. This gives us an important insight into the pathogenesis of Graves’ disease. PMID:24467196

Multivalent ligands control stem cell behaviour in vitro and in vivo

NASA Astrophysics Data System (ADS)

Conway, Anthony; Vazin, Tandis; Spelke, Dawn P.; Rode, Nikhil A.; Healy, Kevin E.; Kane, Ravi S.; Schaffer, David V.

2013-11-01

There is broad interest in designing nanostructured materials that can interact with cells and regulate key downstream functions. In particular, materials with nanoscale features may enable control over multivalent interactions, which involve the simultaneous binding of multiple ligands on one entity to multiple receptors on another and are ubiquitous throughout biology. Cellular signal transduction of growth factor and morphogen cues (which have critical roles in regulating cell function and fate) often begins with such multivalent binding of ligands, either secreted or cell-surface-tethered to target cell receptors, leading to receptor clustering. Cellular mechanisms that orchestrate ligand-receptor oligomerization are complex, however, so the capacity to control multivalent interactions and thereby modulate key signalling events within living systems is currently very limited. Here, we demonstrate the design of potent multivalent conjugates that can organize stem cell receptors into nanoscale clusters and control stem cell behaviour in vitro and in vivo. The ectodomain of ephrin-B2, normally an integral membrane protein ligand, was conjugated to a soluble biopolymer to yield multivalent nanoscale conjugates that potently induce signalling in neural stem cells and promote their neuronal differentiation both in culture and within the brain. Super-resolution microscopy analysis yielded insights into the organization of the receptor-ligand clusters at the nanoscale. We also found that synthetic multivalent conjugates of ephrin-B1 strongly enhance human embryonic and induced pluripotent stem cell differentiation into functional dopaminergic neurons. Multivalent bioconjugates are therefore powerful tools and potential nanoscale therapeutics for controlling the behaviour of target stem cells in vitro and in vivo.

Mind-controlled transgene expression by a wireless-powered optogenetic designer cell implant.

PubMed

Folcher, Marc; Oesterle, Sabine; Zwicky, Katharina; Thekkottil, Thushara; Heymoz, Julie; Hohmann, Muriel; Christen, Matthias; Daoud El-Baba, Marie; Buchmann, Peter; Fussenegger, Martin

2014-11-11

Synthetic devices for traceless remote control of gene expression may provide new treatment opportunities in future gene- and cell-based therapies. Here we report the design of a synthetic mind-controlled gene switch that enables human brain activities and mental states to wirelessly programme the transgene expression in human cells. An electroencephalography (EEG)-based brain-computer interface (BCI) processing mental state-specific brain waves programs an inductively linked wireless-powered optogenetic implant containing designer cells engineered for near-infrared (NIR) light-adjustable expression of the human glycoprotein SEAP (secreted alkaline phosphatase). The synthetic optogenetic signalling pathway interfacing the BCI with target gene expression consists of an engineered NIR light-activated bacterial diguanylate cyclase (DGCL) producing the orthogonal second messenger cyclic diguanosine monophosphate (c-di-GMP), which triggers the stimulator of interferon genes (STING)-dependent induction of synthetic interferon-β promoters. Humans generating different mental states (biofeedback control, concentration, meditation) can differentially control SEAP production of the designer cells in culture and of subcutaneous wireless-powered optogenetic implants in mice.

Geometric effects in microfluidics on heterogeneous cell stress using an Eulerian-Lagrangian approach.

PubMed

Warren, K M; Mpagazehe, J N; LeDuc, P R; Higgs, C F

2016-02-07

The response of individual cells at the micro-scale in cell mechanics is important in understanding how they are affected by changing environments. To control cell stresses, microfluidics can be implemented since there is tremendous control over the geometry of the devices. Designing microfluidic devices to induce and manipulate stress levels on biological cells can be aided by computational modeling approaches. Such approaches serve as an efficient precursor to fabricating various microfluidic geometries that induce predictable levels of stress on biological cells, based on their mechanical properties. Here, a three-dimensional, multiphase computational fluid dynamics (CFD) modeling approach was implemented for soft biological materials. The computational model incorporates the physics of the particle dynamics, fluid dynamics and solid mechanics, which allows us to study how stresses affect the cells. By using an Eulerian-Lagrangian approach to treat the fluid domain as a continuum in the microfluidics, we are conducting studies of the cells' movement and the stresses applied to the cell. As a result of our studies, we were able to determine that a channel with periodically alternating columns of obstacles was capable of stressing cells at the highest rate, and that microfluidic systems can be engineered to impose heterogenous cell stresses through geometric configuring. We found that when using controlled geometries of the microfluidics channels with staggered obstructions, we could increase the maximum cell stress by nearly 200 times over cells flowing through microfluidic channels with no obstructions. Incorporating computational modeling in the design of microfluidic configurations for controllable cell stressing could help in the design of microfludic devices for stressing cells such as cell homogenizers.

MLF1 interacting protein: a potential gene therapy target for human prostate cancer?

PubMed

Zhang, Lei; Ji, Guoqing; Shao, Yuzhang; Qiao, Shaoyi; Jing, Yuming; Qin, Rongliang; Sun, Huiming; Shao, Chen

2015-02-01

Here, we investigated the role of one gene that has been previously associated with human prostate carcinoma cells-myelodysplasia/myeloid leukemia factor 1 interacting protein (MLF1IP)-in order to better ascertain its role in human prostate carcinogenesis. The prostate cancer cell line PC-3 was lentivirally transfected to silence endogenous MLF1IP gene expression, which was confirmed by real-time quantitative PCR (RT-qPCR). Cellomics ArrayScan VTI imaging and MTT assays were conducted to assess cell proliferation. Cell cycle phase arrest and apoptosis were assayed by flow cytometry. Colony formation was assessed by fluorescence microscopy. MLF1IP gene expression was also analyzed by RT-qPCR in sixteen prostate cancer tissue samples and six healthy control prostate tissue samples from human patients. Cell proliferation was significantly inhibited in MLF1IP-silenced cells relative to control cells. G1 phase, S and G2/M phase cell counts were not significantly changed in MLF1IP-silenced cells relative to control cells. Apoptosis was significantly increased in MLF1IP-silenced cells, while MLF1IP-silenced cells displayed a significantly reduced number of cell colonies, compared to control cells. The 16 human prostate cancer tissue samples revealed no clear upregulation or downregulation in MLF1IP gene expression. MLF1IP significantly promotes prostate cancer cell proliferation and colony formation and significantly inhibits apoptosis without affecting cell cycle phase arrest. Further study is required to conclusively determine whether MLF1IP is upregulated in human prostate cancer tumors and to determine the precise cellular mechanism(s) for MLF1IP in prostate carcinogenesis.

How do fission yeast cells grow and connect growth to the mitotic cycle?

PubMed

Sveiczer, Ákos; Horváth, Anna

2017-05-01

To maintain size homeostasis in a unicellular culture, cells should coordinate growth to the division cycle. This is achieved via size control mechanisms (also known as size checkpoints), i.e. some events during the mitotic cycle supervene only if the cell has reached a critical size. Rod-shaped cells like those of fission yeast are ideal model organisms to study these checkpoints via time-lapse microphotography. By applying this method, once we can analyse the growth process between two consecutive divisions at a single (or even at an 'average') cellular level, moreover, we can also position the size checkpoint(s) at the population level. Finally, any of these controls can be abolished in appropriate cell cycle mutants, either in steady-state or in induction synchronised cultures. In the latter case, we produce abnormally oversized cells, and microscopic experiments with them clearly show the existence of a critical size above which the size checkpoint ceases (becomes cryptic). In this review, we delineate the development of our knowledge both on the growth mode of fission yeast and on the operating size control(s) during its mitotic cycle. We finish these historical stories with our recent findings, arguing that three different size checkpoints exist in the fission yeast cell cycle, namely in late G1, in mid G2 and in late G2, which has been concluded by analysing these controls in several cell cycle mutants.

Vitamin E Intake and Risk of Renal Cell Carcinoma: A Meta-Analysis of 7 Case-Control Studies.

PubMed

Shang, Yonggang; Yi, Shanhong; Cui, Dong; Han, Guangwei; Liu, Chengcheng

2015-07-01

Vitamin E intake may reduce the risk of renal cell carcinoma, but the results were inconsistent. Hence, we conducted a meta-analysis to assess the association between dietary vitamin E intake and the risk of renal cell carcinoma. We searched PubMed to identify the relevant case-control studies up to June 2014. Reference lists of retrieved articles were also reviewed. Odds ratios and corresponding 95% confidence intervals were used to estimate the association between dietary vitamin E intake and the risk of renal cell carcinoma. We identified 7 case-control studies regarding dietary vitamin E intake and risk of renal cell carcinoma, involving 5789 cases and 14866 controls. The odds ratio of renal cell carcinoma for the highest compared with the lowest dietary vitamin E intake was 0.75 (95% confidence interval: 0.59-0.91), and heterogeneity was observed across studies. The association between dietary vitamin E intake and the risk of renal cell carcinoma was not significantly differed by gender, but this association were inconsistent in the North American and European populations. Our study provided a evidence that there was a significant inverse association of dietary vitamin E intake with risk of renal cell carcinoma. However, this finding was based on the case-control studies, more well-designed cohort studies are needed. Copyright © 2015 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.

[MAPK signaling pathways involved in aluminum-induced apoptosis and necroptosis in SH-SY5Y cells].

PubMed

Jia, Xiaofang; Zhang, Qinli; Niu, Qiao

2014-11-01

To explore the role of MAPK signaling pathway in apoptosis and necroptosis induced by aluminum in SH-SY5Y cells. To imitate neural cell death induced by aluminium, AlCl3 x 6H2O (4 mmol/L) was used to treat SH-SY5Y cells. Necrostatin-1 (Nec-1,60 μmol/L), the specific inhibitor for necroptosis, and zVAD-fmk (20 μmol/L), the specific inhibitor for apoptosis, were added into cultures for inhibiting the occurrence of necroptosis and apoptosis. CCK-8 was performed to measure cell viability, flow cytometry was used to test the difference of apoptosis rate and necrosis rate between groups, and western-blot was used to detect the change of MAPK protein. Compared with blank control group, solvent control group, Nec-1 control group and zVAD-fmk control group, cell viabiligy of Al(3+) exposed group, Al(3+) plus Nec-1 group and Al(3+) plus zVAD-fmk group decreaced (P < 0.05). Compared with Al(3+) exposed group, cell viability of Al(3+) plus Nec-1 group and Al(3+) plus zVAD-fmk group increased (P < 0.05). Necrotic rate and apoptotic rate in Al(3+) exposed group, Al(3+) plus Nec-1 group and Al(3+) plus zVAD-fmk group obviously increased compared with blank control group, solvent control group, Nec-1 control group and zVAD-fmk control group (P < 0.05). Compared with Al(3+) exposed group, necrotic and apaptotic rate of Al(3+) plus zVAD-fmk group and Al(3+) plus Nec-1 group were statistically significant decreased (P < 0.05). Compared with blank control group, solvent control group, Nec-1 control group and zVAD-fmk control group, expression of p-p38 in Al(3+) exposed group, Al(3+) plus Nec-1 group and Al(3+) plus zVAD-fmk group increased obviously (P < 0.05), and expression of p-ERK decreased significantly (P < 0.05). Compared with Al(3+) exposed group, expression of p-p38 decreased (P < 0.05), but p-ERK increased in Al(3+) plus Nec-1 group (P < 0.05). The ERK and p38 MAPK signaling pathways are involved in aluminum-induced necroptosis in SH-SY5Y cells, but only ERK signaling pathway is involved in aluminum-induced apoptosis, and JNK signaling pathway is not involved in aluminum-induced cell death.

Calbindin and parvalbumin are early markers of non-mitotically regenerating hair cells in the bullfrog vestibular otolith organs

NASA Technical Reports Server (NTRS)

Steyger, P. S.; Burton, M.; Hawkins, J. R.; Schuff, N. R.; Baird, R. A.

1997-01-01

Earlier studies have demonstrated hair cell regeneration in the absence of cell proliferation, and suggested that supporting cells could phenotypically convert into hair cells following hair cell loss. Because calcium-binding proteins are involved in gene up-regulation, cell growth, and cell differentiation, we wished to determine if these proteins were up-regulated in scar formations and regenerating hair cells following gentamicin treatment. Calbindin and parvalbumin immunolabeling was examined in control or gentamicin-treated (GT) bullfrog saccular and utricular explants cultured for 3 days in amphibian culture medium or amphibian culture medium supplemented with aphidicolin, a blocker of nuclear DNA replication in eukaryotic cells. In control cultures, calbindin and parvalbumin immunolabeled the hair bundles and, less intensely, the cell bodies of mature hair cells. In GT or mitotically-blocked GT (MBGT) cultures, calbindin and parvalbumin immunolabeling was also seen in the hair bundles, cuticular plates, and cell bodies of hair cells with immature hair bundles. Thus, these antigens were useful markers for both normal and regenerating hair cells. Supporting cell immunolabeling was not seen in control cultures nor in the majority of supporting cells in GT cultures. In MBGT cultures, calbindin and parvalbumin immunolabeling was up-regulated in the cytosol of single supporting cells participating in scar formations and in supporting cells with hair cell-like characteristics. These data provide further evidence that non-mitotic hair cell regeneration in cultures can be accomplished by the conversion of supporting cells into hair cells.

Two Beam Energy Exchange in Hybrid Liquid Crystal Cells with Photorefractive Field Controlled Boundary Conditions (Postprint)

DTIC Science & Technology

2016-09-12

AFRL-RX-WP-JA-2017-0209 TWO BEAM ENERGY EXCHANGE IN HYBRID LIQUID CRYSTAL CELLS WITH PHOTOREFRACTIVE FIELD CONTROLLED BOUNDARY...estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the... CRYSTAL CELLS WITH PHOTOREFRACTIVE FIELD CONTROLLED BOUNDARY CONDITIONS (POSTPRINT) 5a. CONTRACT NUMBER FA8650-16-D-5402-0001 5b. GRANT

Process for control of cell division

NASA Technical Reports Server (NTRS)

Cone, C. D., Jr. (Inventor)

1977-01-01

A method of controlling mitosis of biological cells was developed, which involved inducing a change in the intracellular ionic hierarchy accompanying the cellular electrical transmembrane potential difference (Esubm) of the cells. The ionic hierarchy may be varied by imposing changes on the relative concentrations of Na(+), K(+) and Cl(-), or by directly imposing changes in the physical Esubm level across the cell surface.

A Hypergravity Environment Induced by Centrifugation Alters Plant Cell Proliferation and Growth in an Opposite Way to Microgravity

NASA Astrophysics Data System (ADS)

Manzano, Ana I.; Herranz, Raúl; van Loon, Jack J. W. A.; Medina, F. Javier

2012-12-01

Seeds of Arabidopsis thaliana were exposed to hypergravity environments (2 g and 6 g) and germinated during centrifugation. Seedlings grew for 2 and 4 days before fixation. In all cases, comparisons were performed against an internal (subjected to rotational vibrations and other factors of the machine) and an external control at 1 g. On seedlings grown in hypergravity the total length and the root length were measured. The cortical root meristematic cells were analyzed to investigate the alterations in cell proliferation, which were quantified by counting the number of cells per millimeter in the specific cell files, and cell growth, which were appraised through the rate of ribosome biogenesis, assessed by morphological and morphometrical parameters of the nucleolus. The expression of cyclin B1, a key regulator of entry in mitosis, was assessed by the use of a CYCB1:GUS genetic construction. The results showed significant differences in some of these parameters when comparing the 1 g internal rotational control with the 1 g external control, indicating that the machine by itself was a source of alterations. When the effect of hypergravity was isolated from other environmental factors, by comparing the experimental conditions with the rotational control, cell proliferation appeared depleted, cell growth was increased and there was an enhanced expression of cyclin B1. The functional meaning of these effects is that cell proliferation and cell growth, which are strictly associated functions under normal 1 g ground conditions, are uncoupled under hypergravity. This uncoupling was also described by us in previous experiments as an effect of microgravity, but in an opposite way. Furthermore, root meristems appear thicker in hypergravity-treated than in control samples, which can be related to changes in the cell wall induced by altered gravity.

Rotator cuff repair using cell sheets derived from human rotator cuff in a rat model.

PubMed

Harada, Yoshifumi; Mifune, Yutaka; Inui, Atsuyuki; Sakata, Ryosuke; Muto, Tomoyuki; Takase, Fumiaki; Ueda, Yasuhiro; Kataoka, Takeshi; Kokubu, Takeshi; Kuroda, Ryosuke; Kurosaka, Masahiro

2017-02-01

To achieve biological regeneration of tendon-bone junctions, cell sheets of human rotator-cuff derived cells were used in a rat rotator cuff injury model. Human rotator-cuff derived cells were isolated, and cell sheets were made using temperature-responsive culture plates. Infraspinatus tendons in immunodeficient rats were resected bilaterally at the enthesis. In right shoulders, infraspinatus tendons were repaired by the transosseous method and covered with the cell sheet (sheet group), whereas the left infraspinatus tendons were repaired in the same way without the cell sheet (control group). Histological examinations (safranin-O and fast green staining, isolectin B4, type II collagen, and human-specific CD31) and mRNA expression (vascular endothelial growth factor; VEGF, type II collagen; Col2, and tenomodulin; TeM) were analyzed 4 weeks after surgery. Biomechanical tests were performed at 8 weeks. In the sheet group, proteoglycan at the enthesis with more type II collagen and isolectin B4 positive cells were seen compared with in the control group. Human specific CD31-positive cells were detected only in the sheet group. VEGF and Col2 gene expressions were higher and TeM gene expression was lower in the sheet group than in the control group. In mechanical testing, the sheet group showed a significantly higher ultimate failure load than the control group at 8 weeks. Our results indicated that the rotator-cuff derived cell sheet could promote cartilage regeneration and angiogenesis at the enthesis, with superior mechanical strength compared with the control. Treatment for rotator cuff injury using cell sheets could be a promising strategy for enthesis of tendon tissue engineering. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:289-296, 2017. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Do phosphoinositides regulate membrane water permeability of tobacco protoplasts by enhancing the aquaporin pathway?

PubMed

Ma, Xiaohong; Shatil-Cohen, Arava; Ben-Dor, Shifra; Wigoda, Noa; Perera, Imara Y; Im, Yang Ju; Diminshtein, Sofia; Yu, Ling; Boss, Wendy F; Moshelion, Menachem; Moran, Nava

2015-03-01

Enhancing the membrane content of PtdInsP 2 , the already-recognized protein-regulating lipid, increased the osmotic water permeability of tobacco protoplasts, apparently by increasing the abundance of active aquaporins in their membranes. While phosphoinositides are implicated in cell volume changes and are known to regulate some ion channels, their modulation of aquaporins activity has not yet been reported for any organism. To examine this, we compared the osmotic water permeability (P f) of protoplasts isolated from tobacco (Nicotiana tabacum) cultured cells (NT1) with different (genetically lowered or elevated relative to controls) levels of inositol trisphosphate (InsP3) and phosphatidyl inositol [4,5] bisphosphate (PtdInsP2). To achieve this, the cells were transformed with, respectively, the human InsP3 5-phosphatase ('Ptase cells') or human phosphatidylinositol (4) phosphate 5-kinase ('PIPK cells'). The mean P f of the PIPK cells was several-fold higher relative to that of controls and Ptase cells. Three results favor aquaporins over the membrane matrix as underlying this excessive P f: (1) transient expression of the maize aquaporin ZmPIP2;4 in the PIPK cells increased P f by 12-30 μm s(-1), while in the controls only by 3-4 μm s(-1). (2) Cytosol acidification-known to inhibit aquaporins-lowered the P f in the PIPK cells down to control levels. (3) The transcript of at least one aquaporin was elevated in the PIPK cells. Together, the three results demonstrate the differences between the PIPK cells and their controls, and suggest a hitherto unobserved regulation of aquaporins by phosphoinositides, which could occur through direct interaction or indirect phosphoinositides-dependent cellular effects.

Mobilization Characteristics and Strategies to Improve Hematopoietic Progenitor Cell Mobilization and Collection in Patients with Chronic Granulomatous Disease and Severe Combined Immunodeficiency

PubMed Central

Panch, Sandhya R.; Yau, Yu Ying; Kang, Elizabeth M.; De Ravin, Suk See; Malech, Harry L.; Leitman, Susan F.

2014-01-01

Background G-CSF mobilized autologous hematopoietic progenitor cells (HPC) may be collected by apheresis of patients with chronic granulomatous disease (CGD) and severe combined immunodeficiency (SCID) for use in gene therapy trials. CD34+ cell mobilization has not been well characterized in such patients. Study Design and Methods We retrospectively evaluated CD34+ cell mobilization and collection in 73 consecutive CGD and SCID patients and in 99 age, weight and G-CSF dose-matched healthy allogeneic controls. Results In subjects aged ≤20 years, day 5 pre-apheresis circulating CD34+ counts were significantly lower in CGD and SCID than in controls; mean peak CD34+ cells 58, 64, and 87/uL, respectively, p=0.01. The SCIDs had lower CD34+ collection efficiency than CGDs and controls; mean efficiency 40%, 63% and 57%, respectively, p=0.003. In subjects >20 years, the CGDs had significantly lower CD34+ cell mobilization than controls; mean peak CD34+ cells 41 and 113/uL, respectively, p<0.0001. In a multivariate analysis, lower sedimentation rate (ESR) at mobilization was significantly correlated with better CD34+ cell mobilization, p=0.007. In SCIDs, CD34 collection efficiency was positively correlated with higher red cell indices (MCV: R2=0.77; MCH: R2=0.94; MCHC: R2=0.7, p<0.007) but not hemoglobin. Conclusions CGD and SCID populations are characterized by significantly less robust CD34+ HPC mobilization than healthy controls. The presence of active inflammation/infection as suggested by an elevated ESR may negatively impact mobilization. Among SCIDs, markedly reduced CD34 collection efficiencies were related to iron deficiency, wherein decreased red cell size and density may impair apheresis cell separation mechanics. PMID:25143186

Designing degradable hydrogels for orthogonal control of cell microenvironments

PubMed Central

Kharkar, Prathamesh M.

2013-01-01

Degradable and cell-compatible hydrogels can be designed to mimic the physical and biochemical characteristics of native extracellular matrices and provide tunability of degradation rates and related properties under physiological conditions. Hence, such hydrogels are finding widespread application in many bioengineering fields, including controlled bioactive molecule delivery, cell encapsulation for controlled three-dimensional culture, and tissue engineering. Cellular processes, such as adhesion, proliferation, spreading, migration, and differentiation, can be controlled within degradable, cell-compatible hydrogels with temporal tuning of biochemical or biophysical cues, such as growth factor presentation or hydrogel stiffness. However, thoughtful selection of hydrogel base materials, formation chemistries, and degradable moieties is necessary to achieve the appropriate level of property control and desired cellular response. In this review, hydrogel design considerations and materials for hydrogel preparation, ranging from natural polymers to synthetic polymers, are overviewed. Recent advances in chemical and physical methods to crosslink hydrogels are highlighted, as well as recent developments in controlling hydrogel degradation rates and modes of degradation. Special attention is given to spatial or temporal presentation of various biochemical and biophysical cues to modulate cell response in static (i.e., non-degradable) or dynamic (i.e., degradable) microenvironments. This review provides insight into the design of new cell-compatible, degradable hydrogels to understand and modulate cellular processes for various biomedical applications. PMID:23609001

Fuel cell-gas turbine hybrid system design part II: Dynamics and control

NASA Astrophysics Data System (ADS)

McLarty, Dustin; Brouwer, Jack; Samuelsen, Scott

2014-05-01

Fuel cell gas turbine hybrid systems have achieved ultra-high efficiency and ultra-low emissions at small scales, but have yet to demonstrate effective dynamic responsiveness or base-load cost savings. Fuel cell systems and hybrid prototypes have not utilized controls to address thermal cycling during load following operation, and have thus been relegated to the less valuable base-load and peak shaving power market. Additionally, pressurized hybrid topping cycles have exhibited increased stall/surge characteristics particularly during off-design operation. This paper evaluates additional control actuators with simple control methods capable of mitigating spatial temperature variation and stall/surge risk during load following operation of hybrid fuel cell systems. The novel use of detailed, spatially resolved, physical fuel cell and turbine models in an integrated system simulation enables the development and evaluation of these additional control methods. It is shown that the hybrid system can achieve greater dynamic response over a larger operating envelope than either individual sub-system; the fuel cell or gas turbine. Results indicate that a combined feed-forward, P-I and cascade control strategy is capable of handling moderate perturbations and achieving a 2:1 (MCFC) or 4:1 (SOFC) turndown ratio while retaining >65% fuel-to-electricity efficiency, while maintaining an acceptable stack temperature profile and stall/surge margin.

Controlled regular locomotion of algae cell microrobots.

PubMed

Xie, Shuangxi; Jiao, Niandong; Tung, Steve; Liu, Lianqing

2016-06-01

Algae cells can be considered as microrobots from the perspective of engineering. These organisms not only have a strong reproductive ability but can also sense the environment, harvest energy from the surroundings, and swim very efficiently, accommodating all these functions in a body of size on the order of dozens of micrometers. An interesting topic with respect to random swimming motions of algae cells in a liquid is how to precisely control them as microrobots such that they swim according to manually set routes. This study developed an ingenious method to steer swimming cells based on the phototaxis. The method used a varying light signal to direct the motion of the cells. The swimming trajectory, speed, and force of algae cells were analyzed in detail. Then the algae cell could be controlled to swim back and forth, and traverse a crossroad as a microrobot obeying specific traffic rules. Furthermore, their motions along arbitrarily set trajectories such as zigzag, and triangle were realized successfully under optical control. Robotize algae cells can be used to precisely transport and deliver cargo such as drug particles in microfluidic chip for biomedical treatment and pharmacodynamic analysis. The study findings are expected to bring significant breakthrough in biological drives and new biomedical applications.

TCR Signal Strength Alters T–DC Activation and Interaction Times and Directs the Outcome of Differentiation

PubMed Central

van Panhuys, Nicholas

2016-01-01

The ability of CD4+ T cells to differentiate into effector subsets underpins their ability to shape the immune response and mediate host protection. During T cell receptor-induced activation of CD4+ T cells, both the quality and quantity of specific activatory peptide/MHC ligands have been shown to control the polarization of naive CD4+ T cells in addition to co-stimulatory and cytokine-based signals. Recently, advances in two-­photon microscopy and tetramer-based cell tracking methods have allowed investigators to greatly extend the study of the role of TCR signaling in effector differentiation under in vivo conditions. In this review, we consider data from recent in vivo studies analyzing the role of TCR signal strength in controlling the outcome of CD4+ T cell differentiation and discuss the role of TCR in controlling the critical nature of CD4+ T cell interactions with dendritic cells during activation. We further propose a model whereby TCR signal strength controls the temporal aspects of T–DC interactions and the implications for this in mediating the downstream signaling events, which influence the transcriptional and epigenetic regulation of effector differentiation. PMID:26834747

Influence of simulated microgravity on the longevity of insect-cell culture

NASA Technical Reports Server (NTRS)

Cowger, N. L.; O'Connor, K. C.; Bivins, J. E.

1997-01-01

Simulated microgravity within the NASA High Aspect Rotating-Wall Vessel (HARV) provides a quiescent environment to culture fragile insect cells. In this vessel, the duration of stationary and death phase for cultures of Spodoptera frugiperda cells was greatly extended over that achieved in shaker-flask controls. For both HARV and control cultures, S. frugiperda cells grew to concentrations in excess of 1 x 10(7) viable cells ml-1 with viabilities greater than 90%. In the HARV, stationary phase was maintained 9-15 days in contrast to 4-5 days in the shaker flask. Furthermore, the rate of cell death was reduced in the HARV by a factor of 20-90 relative to the control culture and was characterized with a death rate constant of 0.01-0.02 day-1. Beginning in the stationary phase and continuing in the death phase, there was a significant decrease in population size in the HARV versus an increase in the shaker flask. This phenomenon could represent cell adaptation to simulated microgravity and/or a change in the ratio of apoptotic to necrotic cells. Differences observed in this research between the HARV and its control were attributed to a reduction in hydrodynamic forces in the microgravity vessel.

The UK Stem Cell Bank: a UK government-funded, international resource center for stem cell research.

PubMed

Stacey, Glyn; Hunt, Charles J

2006-01-01

The UK Stem Cell Bank is a UK Research Council-funded initiative that aims to provide ethically sourced and quality controlled stocks of cells for researchers and also establish seed stocks of cell lines for clinical trials. Whilst the Bank is prohibited from carrying out basic stem cell research (to avoid conflicts of interest) it is working to improve stem cell banking procedures including cryopreservation, characterization and quality control. The Bank also supports training activities and has provided the hub for the International Stem Cell Initiative, which includes 17 expert stem cell centers aiming to characterize a large number of human embryonic stem cell lines in a standardized way to improve our understanding of the characteristics of these cells.

YAP expression in normal and neoplastic breast tissue: an immunohistochemical study.

PubMed

Jaramillo-Rodríguez, Yolanda; Cerda-Flores, Ricardo M; Ruiz-Ramos, Ruben; López-Márquez, Francisco C; Calderón-Garcidueñas, Ana Laura

2014-04-01

Yes-associated protein (YAP) is a transcriptional factor involved in normal cell proliferation, apoptosis and carcinogenesis; however, its contribution to breast cancer (BC) is still controversial. We undertook this study to compare the expression of YAP by immunohistochemistry (IHC) in normal breast tissue of women without breast cancer (BC) (controls), non-neoplastic breast tissue in women with cancer (internal controls) and in four different subtypes of invasive ductal carcinoma. There were 17 controls and 105 tumor cases (53 luminal A, 15 luminal B, 20 overexpression of HER2 and 17 triple negative cases) studied by IHC. Statistical analysis included χ(2) for linear trend (Extended Mantel-Haenszel). There were 40% of internal controls that showed expression of YAP in myoepithelial cells, whereas in controls expression was 100%. In controls, 3/17 (17.6%) showed cytoplasmic staining in luminal cells. There was a significant difference in nuclear expression between the ductal BC subtypes. Luminal A had 4% of positive cases with <10% of cells affected in each case; in contrast, there were 17-20% of positive cases in the other groups with 50% or more of stained cells. YAP expression in stromal cells was not observed in controls or in triple-negative cases, and luminal B pattern had the highest YAP nuclear expression (20%). YAP showed decreased expression in tumor cells compared with normal breast tissue. These findings are consistent with a role of YAP as a suppressor gene in BC and show differences in YAP expression in different patterns of ductal BC. Copyright © 2014 IMSS. Published by Elsevier Inc. All rights reserved.

Mammalian phospholipid homeostasis: evidence that membrane curvature elastic stress drives homeoviscous adaptation in vivo

PubMed Central

2016-01-01

Several theories of phospholipid homeostasis have postulated that cells regulate the molecular composition of their bilayer membranes, such that a common biophysical membrane parameter is under homeostatic control. Two commonly cited theories are the intrinsic curvature hypothesis, which states that cells control membrane curvature elastic stress, and the theory of homeoviscous adaptation, which postulates cells control acyl chain packing order (membrane order). In this paper, we present evidence from data-driven modelling studies that these two theories correlate in vivo. We estimate the curvature elastic stress of mammalian cells to be 4–7 × 10−12 N, a value high enough to suggest that in mammalian cells the preservation of membrane order arises through a mechanism where membrane curvature elastic stress is controlled. These results emerge from analysing the molecular contribution of individual phospholipids to both membrane order and curvature elastic stress in nearly 500 cellular compositionally diverse lipidomes. Our model suggests that the de novo synthesis of lipids is the dominant mechanism by which cells control curvature elastic stress and hence membrane order in vivo. These results also suggest that cells can increase membrane curvature elastic stress disproportionately to membrane order by incorporating polyunsaturated fatty acids into lipids. PMID:27534697

Mammalian phospholipid homeostasis: evidence that membrane curvature elastic stress drives homeoviscous adaptation in vivo.

PubMed

Dymond, Marcus K

2016-08-01

Several theories of phospholipid homeostasis have postulated that cells regulate the molecular composition of their bilayer membranes, such that a common biophysical membrane parameter is under homeostatic control. Two commonly cited theories are the intrinsic curvature hypothesis, which states that cells control membrane curvature elastic stress, and the theory of homeoviscous adaptation, which postulates cells control acyl chain packing order (membrane order). In this paper, we present evidence from data-driven modelling studies that these two theories correlate in vivo. We estimate the curvature elastic stress of mammalian cells to be 4-7 × 10(-12) N, a value high enough to suggest that in mammalian cells the preservation of membrane order arises through a mechanism where membrane curvature elastic stress is controlled. These results emerge from analysing the molecular contribution of individual phospholipids to both membrane order and curvature elastic stress in nearly 500 cellular compositionally diverse lipidomes. Our model suggests that the de novo synthesis of lipids is the dominant mechanism by which cells control curvature elastic stress and hence membrane order in vivo These results also suggest that cells can increase membrane curvature elastic stress disproportionately to membrane order by incorporating polyunsaturated fatty acids into lipids. © 2016 The Author(s).

Novel Metrics to Characterize Embryonic Elongation of the Nematode Caenorhabditis elegans.

PubMed

Martin, Emmanuel; Rocheleau-Leclair, Olivier; Jenna, Sarah

2016-03-28

Dissecting the signaling pathways that control the alteration of morphogenic processes during embryonic development requires robust and sensitive metrics. Embryonic elongation of the nematode Caenorhabditis elegans is a late developmental stage consisting of the elongation of the embryo along its longitudinal axis. This developmental stage is controlled by intercellular communication between hypodermal cells and underlying body-wall muscles. These signaling mechanisms control the morphology of hypodermal cells by remodeling the cytoskeleton and the cell-cell junctions. Measurement of embryonic lethality and developmental arrest at larval stages as well as alteration of cytoskeleton and cell-cell adhesion structures in hypodermal and muscle cells are classical phenotypes that have been used for more than 25 years to dissect these signaling pathways. Recent studies required the development of novel metrics specifically targeting either early or late elongation and characterizing morphogenic defects along the antero-posterior axis of the embryo. Here, we provide detailed protocols enabling the accurate measurement of the length and the width of the elongating embryos as well as the length of synchronized larvae. These methods constitute useful tools to identify genes controlling elongation, to assess whether these genes control both early and late phases of this stage and are required evenly along the antero-posterior axis of the embryo.

Touch-plate and statolith formation in graviceptors of ephyrae which developed while weightless in space

NASA Technical Reports Server (NTRS)

Spangenberg, D. B.; Coccaro, E.; Schwarte, R.; Lowe, B.

1996-01-01

Ultrastructural studies of the statocysts and touch-plates of graviceptors (rhopalia) of Aurelia ephyrae revealed that (1) touch-plate hair cells are present; and (2) cytoplasmic strands from the hair cell bases extend from the neurite plexus to touch similar strands from the lithocytes. This close association of hair cell neurites and statocysts may have important implications regarding the transmitting and processing of positional information with respect to the gravity vector. Graviceptors of ephyrae which developed while weightless in microgravity were compared with controls at the ultrastructural level. We found that hair cells of ephyrae which developed in microgravity had fewer lipid droplets in the large spaces near their bases as compared with 1 g controls. In the ephyrae from the first microgravity experiment, hair cells had more large apical vacuoles with filamentous content than were found in hair cells of ephyrae from the second experiment and controls. The neurite plexus and the network of cytoplasmic strands extending to the statocysts were not different in microgravity-developed ephyrae from controls. Behavioral differences in swimming and orienting in ephyrae in microgravity and controls (reported earlier) were not explained by morphological differences in the hair cells of the touch-plates or the statocysts, although functional differences apparently occurred.

Expression of extracellular matrix metalloproteinase inducer (EMMPRIN) in the endometrium of patients with repeated implantation failure after in vitro fertilization.

PubMed

Turgut, A; Goruk, N Y; Tunc, S Y; Agaçayak, E; Alabalik, U; Yalinkaya, A; Gül, T

2014-01-01

To compare the immunohistochemical expression of extracellular matrix metalloproteinase inducer (EMMPRIN) in repeated implantation failure (RIF) patients with normal fertile controls. The study group consisted of primary infertile patients with RIF and normal fertile controls between January 2011 and February 2013. Endometrial samples received at the luteal phase were exposed to immunohistochemical staining for EMMPRIN antibodies. EMMPRIN expression of endometrial glandular epithelial cells, stromal cells and vascular endothelial cells were evaluated. The main outcome measure was defined as immunohistochemical score with regard to the severity and extent of staining. The study group consisted of 26 primary infertile patients, whereas the control group consisted of 40 normal fertile controls. The fertile group was found to have stronger expression of EMMPRIN than the study group when endometrial glandular epithelial cells, stromal cells and vascular endothelial cells were evaluated with regards to the severity of staining (p < 0.001), the extent of staining (p < 0.001) and total staining score (p < 0.001). This is the first study showing low expression of EMMPRIN in the endometrial cells of the patients with RIF compared with fertile healthy controls. We suggest that reduced EMMPRIN expression in the human endometrium may lead to poor endometrial receptivity.

Better Bet-Hedging with coupled positive and negative feedback loops

NASA Astrophysics Data System (ADS)

Narula, Jatin; Igoshin, Oleg

2011-03-01

Bacteria use the phenotypic heterogeneity associated with bistable switches to distribute the risk of activating stress response strategies like sporulation and persistence. However bistable switches offer little control over the timing of phenotype switching and first passage times (FPT) for individual cells are found to be exponentially distributed. We show that a genetic circuit consisting of interlinked positive and negative feedback loops allows cells to control the timing of phenotypic switching. Using a mathematical model we find that in this system a stable high expression state and stable low expression limit cycle coexist and the FPT distribution for stochastic transitions between them shows multiple peaks at regular intervals. A multimodal FPT distribution allows cells to detect the persistence of stress and control the rate of phenotype transition of the population. We further show that extracellular signals from cell-cell communication that change the strength of the feedback loops can modulate the FPT distribution and allow cells even greater control in a bet-hedging strategy.

Planar solid oxide fuel cell with staged indirect-internal air and fuel preheating and reformation

DOEpatents

Geisbrecht, Rodney A; Williams, Mark C

2003-10-21

A solid oxide fuel cell arrangement and method of use that provides internal preheating of both fuel and air in order to maintain the optimum operating temperature for the production of energy. The internal preheat passes are created by the addition of two plates, one on either side of the bipolar plate, such that these plates create additional passes through the fuel cell. This internal preheat fuel cell configuration and method reduce the requirements for external heat exchanger units and air compressors. Air or fuel may be added to the fuel cell as required to maintain the optimum operating temperature through a cathode control valve or an anode control valve, respectively. A control loop comprises a temperature sensing means within the preheat air and fuel passes, a means to compare the measured temperature to a set point temperature and a determination based on the comparison as to whether the control valves should allow additional air or fuel into the preheat or bypass manifolds of the fuel cell.

Topology and Control of the Cell-Cycle-Regulated Transcriptional Circuitry

PubMed Central

Haase, Steven B.; Wittenberg, Curt

2014-01-01

Nearly 20% of the budding yeast genome is transcribed periodically during the cell division cycle. The precise temporal execution of this large transcriptional program is controlled by a large interacting network of transcriptional regulators, kinases, and ubiquitin ligases. Historically, this network has been viewed as a collection of four coregulated gene clusters that are associated with each phase of the cell cycle. Although the broad outlines of these gene clusters were described nearly 20 years ago, new technologies have enabled major advances in our understanding of the genes comprising those clusters, their regulation, and the complex regulatory interplay between clusters. More recently, advances are being made in understanding the roles of chromatin in the control of the transcriptional program. We are also beginning to discover important regulatory interactions between the cell-cycle transcriptional program and other cell-cycle regulatory mechanisms such as checkpoints and metabolic networks. Here we review recent advances and contemporary models of the transcriptional network and consider these models in the context of eukaryotic cell-cycle controls. PMID:24395825

Integument pattern formation involves genetic and epigenetic controls: feather arrays simulated by digital hormone models

PubMed Central

Jiang, Ting-Xin; Widelitz, Randall B.; Shen, Wei-Min; Will, Peter; Wu, Da-Yu; Lin, Chih-Min; Jung, Han-Sung; Chuong, Cheng-Ming

2015-01-01

Pattern formation is a fundamental morphogenetic process. Models based on genetic and epigenetic control have been proposed but remain controversial. Here we use feather morphogenesis for further evaluation. Adhesion molecules and/or signaling molecules were first expressed homogenously in feather tracts (restrictive mode, appear earlier) or directly in bud or inter-bud regions (de novo mode, appear later). They either activate or inhibit bud formation, but paradoxically co-localize in the bud. Using feather bud reconstitution, we showed that completely dissociated cells can reform periodic patterns without reference to previous positional codes. The patterning process has the characteristics of being self-organizing, dynamic and plastic. The final pattern is an equilibrium state reached by competition, and the number and size of buds can be altered based on cell number and activator/inhibitor ratio, respectively. We developed a Digital Hormone Model which consists of (1) competent cells without identity that move randomly in a space, (2) extracellular signaling hormones which diffuse by a reaction-diffusion mechanism and activate or inhibit cell adhesion, and (3) cells which respond with topological stochastic actions manifested as changes in cell adhesion. Based on probability, the results are cell clusters arranged in dots or stripes. Thus genetic control provides combinational molecular information which defines the properties of the cells but not the final pattern. Epigenetic control governs interactions among cells and their environment based on physical-chemical rules (such as those described in the Digital Hormone Model). Complex integument patterning is the sum of these two components of control and that is why integument patterns are usually similar but non-identical. These principles may be shared by other pattern formation processes such as barb ridge formation, fingerprints, pigmentation patterning, etc. The Digital Hormone Model can also be applied to swarming robot navigation, reaching intelligent automata and representing a self-re-configurable type of control rather than a follow-the-instruction type of control. PMID:15272377

[Role of connective tissue growth factor (CTGF) in proliferation and migration of pancreatic cancer cells].

PubMed

Bai, Yu-chun; Kang, Quan; Luo, Qing; Wu, Dao-qi; Ye, Wei-xia; Lin, Xue-mei; Zhao, Yong

2011-10-01

To explore the expression of connective tissue growth factor (CTGF) in pancreatic cancer and its influence on the proliferation and migration of cancer cells. The expression of CTGF in pancreatic cell line PANC-1 cells was analyzed by real-time PCR and in pancreatic carcinoma (50 cases) tissues by immunohistochemistry. The ability of proliferation and migration in vitro of PANC-1 cells was tested by MTT assay, scratch test and Boyden chamber test after the CTGF gene was overexpressed by Ad5-CTGF or silenced with Ad5-siCTGF transfection. CTGF was overexpressed in both pancreatic cancer cells and tissues. Overxpression of CTGF leads to increased proliferation and migration of PANC-1 cells. The CTGF-transfected PANC-1 cells showed apparent stronger proliferation ability and scratch-repair ability than that of empty vector controls. The results of Boyden chamber test showed that there were 34 cells/field (200× magnificantion) of the CTGF-transfected overexpressing cells, much more than the 11 cells/field of the empty vector control cells; and 6 cells/microscopic field of the Ad5-siCTGF-transfected silenced cells, much less than the 15 cells/field of the control cells. CTGF is overexpressed in both pancreatic cancer cells in vitro and in vivo, indicating that it may play an important role in the cell proliferation and migration in pancreatic cancer.

VGF expression by T lymphocytes in patients with Alzheimer's disease

PubMed Central

Glorius, Sarah; Dobrowolny, Henrik; Greiner-Bohl, Sabrina; Mawrin, Christian; Bommhardt, Ursula; Hartig, Roland; Bogerts, Bernhard; Busse, Mandy

2015-01-01

Secretion of VGF is increased in cerebrospinal fluid and blood in neurodegenerative disorders like Alzheimer's disease (AD) and VGF is a potential biomarker for these disorders. We have shown that VGF is expressed in peripheral T cells and is correlated with T cell survival and cytokine secretion. The frequency of VGF+CD3+ T cells increases with normal aging. We found an increased number of VGF-expressing T cells in patients with AD compared to aged healthy controls, which was associated with enhanced HbA1c levels in blood. Upon treatment with rivastigmine, T cell proliferation and VGF expression in AD patients decreased to the level found in controls. Moreover, rapamycin treatment in vitro reduced the number of VGF+CD3+ cells in AD patients to control levels. PMID:26142708

The response of single human cells to zero gravity

NASA Technical Reports Server (NTRS)

Montgomery, P. O., Jr.; Cook, J. E.; Reynolds, R. C.; Paul, J. S.; Hayflick, L.; Schulz, W. W.; Stock, D.; Kinzey, S.; Rogers, T.; Campbell, D.

1975-01-01

Twenty separate cultures of Wistar-38 human embryonic lung cells were exposed to a zero-gravity environment on Skylab for periods of time ranging from one to 59 days. Duplicate cultures were run concurrently as ground controls. Ten cultures were fixed on board the satellite during the first 12 days of flight. Growth curves, DNA microspectrophotometry, phase microscopy, and ultrastructural studies of the fixed cells revealed no effects of a zero-gravity environment on the ten cultures. Two cultures were photographed with phase time lapse cinematography during the first 27 days of flight. No differences were found in mitotic index, cell cycle, and migration between the flight and control cells. Eight cultures were returned to earth in an incubated state. Karyotyping and chromosome banding tests show no differences between the flight and control cells.

Simulation system of arrhythmia using ActiveX control.

PubMed

Takeuchi, Akihiro; Hirose, Minoru; Hamada, Atsushi; Ikeda, Noriaki

2005-07-01

A simulation system for arrhythmias has been developed using Windows-based software technology, ActiveX control. The cardiac module consists of six cells, the sinus, atrium, AV node, ventricle, and ectopic foci. The physiological properties of the cells, the automaticity and conduction delay, were modelled, respectively, by the phase response curve and the excitability recovery curve. Cell functions were implemented in the ActiveX control and incorporated into the cardiac module. The system draws the ECG sequence as a ladder diagram in real time. The system interactively shows diverse arrhythmias for various user settings of the cell function and bidirectional conduction between the cells. Users are able to experiment virtually by setting up a so-called electrophysiological stimulation. This system is useful for learning and for teaching the interaction between the cells and arrhythmias.

Modeling, analysis and control of fuel cell hybrid power systems

NASA Astrophysics Data System (ADS)

Suh, Kyung Won

Transient performance is a key characteristic of fuel cells, that is sometimes more critical than efficiency, due to the importance of accepting unpredictable electric loads. To fulfill the transient requirement in vehicle propulsion and portable fuel cell applications, a fuel cell stack is typically coupled with a battery through a DC/DC converter to form a hybrid power system. Although many power management strategies already exist, they all rely on low level controllers that realize the power split. In this dissertation we design controllers that realize various power split strategies by directly manipulating physical actuators (low level commands). We maintain the causality of the electric dynamics (voltage and current) and investigate how the electric architecture affects the hybridization level and the power management. We first establish the performance limitations associated with a stand-alone and power-autonomous fuel cell system that is not supplemented by an additional energy storage and powers all its auxiliary components by itself. Specifically, we examine the transient performance in fuel cell power delivery as it is limited by the air supplied by a compressor driven by the fuel cell itself. The performance limitations arise from the intrinsic coupling in the fluid and electrical domain between the compressor and the fuel cell stack. Feedforward and feedback control strategies are used to demonstrate these limitations analytically and with simulations. Experimental tests on a small commercial fuel cell auxiliary power unit (APU) confirm the dynamics and the identified limitations. The dynamics associated with the integration of a fuel cell system and a DC/DC converter is then investigated. Decentralized and fully centralized (using linear quadratic techniques) controllers are designed to regulate the power system voltage and to prevent fuel cell oxygen starvation. Regulating these two performance variables is a difficult task and requires a compromise due to the conflicting objectives. The compromise can be mitigated by augmenting the fuel cell power system with an energy buffer such as a battery. We consider two different and popular ways of connecting the battery and the fuel cell to the load and we refer to them as electric architectures. Various controller gains are used to span the fuel cell operation from load-following to load-leveling, and hence, to determine adequate fuel cell-battery sizing (hybridization level) and the associated trends in the system efficiency.

Three-dimensional control of Tetrahymena pyriformis using artificial magnetotaxis

NASA Astrophysics Data System (ADS)

Hyung Kim, Dal; Seung Soo Kim, Paul; Agung Julius, Anak; Jun Kim, Min

2012-01-01

We demonstrate three-dimensional control with the eukaryotic cell Tetrahymena pyriformis (T. pyriformis) using two sets of Helmholtz coils for xy-plane motion and a single electromagnet for z-direction motion. T. pyriformis is modified to have artificial magnetotaxis with internalized magnetite. To track the cell's z-axis position, intensity profiles of non-motile cells at varying distances from the focal plane are used. During vertical motion along the z-axis, the intensity difference is used to determine the position of the cell. The three-dimensional control of the live microorganism T. pyriformis as a cellular robot shows great potential for practical applications in microscale tasks, such as target transport and cell therapy.

Lithium Ion Battery (LIB) Charger: Spacesuit Battery Charger Design with 2-Fault Tolerance to Catastrophic Hazards

NASA Technical Reports Server (NTRS)

Darcy, Eric; Davies, Frank

2009-01-01

Charger design that is 2-fault tolerant to catastrophic has been achieved for the Spacesuit Li-ion Battery with key features. Power supply control circuit and 2 microprocessors independently control against overcharge. 3 microprocessor control against undercharge (false positive: Go for EVA) conditions. 2 independent channels provide functional redundancy. Capable of charge balancing cell banks in series. Cell manufacturing and performance uniformity is excellent with both designs. Once a few outliers are removed, LV cells are slightly more uniform than MoliJ cells. If cell balance feature of charger is ever invoked, it will be an indication of a significant degradation issue, not a nominal condition.

Control of DNA replication: a new facet of Hox proteins?

PubMed

Miotto, Benoit; Graba, Yacine

2010-09-01

Hox proteins are well-known as developmental transcription factors controlling cell and tissue identity, but recent findings suggest that they are also part of the cell replication machinery. Hox-mediated control of transcription and replication may ensure coordinated control of cell growth and differentiation, two processes that need to be tightly and precisely coordinated to allow proper organ formation and patterning. In this review we summarize the available data linking Hox proteins to the replication machinery and discuss the developmental and pathological implications of this new facet of Hox protein function.

A Maximum Power Point Tracking Control Method of a Photovoltaic Power Generator with Consideration of Dynamic Characteristics of Solar Cells

NASA Astrophysics Data System (ADS)

Watanabe, Takashi; Yoshida, Toshiya; Ohniwa, Katsumi

This paper discusses a new control strategy for photovoltaic power generation systems with consideration of dynamic characteristics of the photovoltaic cells. The controller estimates internal currents of an equivalent circuit for the cells. This estimated, or the virtual current and the actual voltage of the cells are fed to a conventional Maximum-Power-Point-Tracking (MPPT) controller. Consequently, this MPPT controller still tracks the optimum point even though it is so designed that the seeking speed of the operating point is extremely high. This system may suit for applications, which are installed in rapidly changeable insolation and temperature-conditions e.g. automobiles, trains, and airplanes. The proposed method is verified by experiment with a combination of this estimating function and the modified Boehringer's MPPT algorithm.

Effect and mechanism of PAR-2 on the proliferation of esophageal cancer cells.

PubMed

Quanjun, D; Qingyu, Z; Qiliang, Z; Liqun, X; Jinmei, C; Ziquan, L; Shike, H

2016-11-01

Esophageal Cancer (EC) is a common malignant tumor occurred in the digestive tract. In this study, we investigated the mechanism of Protease Activated Receptor 2 (PAR-2) on the proliferation of esophageal cancer cell. Transfected esophageal cancer (EC) cell (PAR-2shRNA EC109) was established with low stable PAR-2 expression. EC109 cell was treated with PAR-2 agonist, PAR-2 anti-agonist and MAPK inhibitor respectively; Untreated EC109 cell (blank control) and PAR-2shRNA EC109 cell were used for analysis also. The mRNA expressions of PAR-2, ERK1, Cyclin D1, and c-fos in each group were detected by reverse transcript and polymerase chain reaction. Western blot was used to detect the protein expressions in each group. The cell growth curves were drawn to compare the cell growth. Compared with the blank control, the mRNA and protein expressions of PAR-2, Cyclin D1, and c-fos in PAR-2 agonist group increased significantly (p < 0.05), while decreased significantly in PAR-2shRNA EC109 cell and MAPK inhibitor group (p < 0.05). The mRNA expression of ERK1 and protein expression of p-ERK1 increased in PAR-2 agonist group, decreased in PAR-2shRNA EC109 cell and MAPK inhibitor group when compared with blank control (p < 0.05). The growth of cells was upward in PAR-2 agonist group at cell growth phase when compared with blank control, while decreased in PAR-2 shRNA EC109 cell and MAPK inhibitor group with statistical difference (p < 0.05). PAR-2 regulate cell proliferation through the MAPK pathway in esophageal carcinoma cell, and Cyclin D1, c-fos are involved in this process.

Protease Activated Receptor-2 Expression and Function in Asthmatic Bronchial Smooth Muscle

PubMed Central

Gilbert, Guillaume; Carvalho, Gabrielle; Trian, Thomas; Ozier, Annaig; Gillibert-Duplantier, Jennifer; Ousova, Olga; Maurat, Elise; Thumerel, Matthieu; Quignard, Jean-François; Girodet, Pierre-Olivier; Marthan, Roger; Berger, Patrick

2014-01-01

Asthmatic bronchial smooth muscle (BSM) is characterized by structural remodeling associated with mast cell infiltration displaying features of chronic degranulation. Mast cell-derived tryptase can activate protease activated receptor type-2 (PAR-2) of BSM cells. The aims of the present study were (i) to evaluate the expression of PAR-2 in both asthmatic and non asthmatic BSM cells and, (ii) to analyze the effect of prolonged stimulation of PAR-2 in asthmatic BSM cells on cell signaling and proliferation. BSM cells were obtained from both 33 control subjects and 22 asthmatic patients. PAR-2 expression was assessed by flow cytometry, western blot and quantitative RT-PCR. Calcium response, transduction pathways and proliferation were evaluated before and following PAR-2 stimulation by SLIGKV-NH2 or trypsin for 1 to 3 days. Asthmatic BSM cells expressed higher basal levels of functional PAR-2 compared to controls in terms of mRNA, protein expression and calcium response. When PAR-2 expression was increased by means of lentivirus in control BSM cells to a level similar to that of asthmatic cells, PAR-2-induced calcium response was then similar in both types of cell. However, repeated PAR-2 stimulations increased the proliferation of asthmatic BSM cells but not that of control BSM cells even following lentiviral over-expression of PAR-2. Such an increased proliferation was related to an increased phosphorylation of ERK in asthmatic BSM cells. In conclusion, we have demonstrated that asthmatic BSM cells express increased baseline levels of functional PAR-2. This higher basal level of PAR-2 accounts for the increased calcium response to PAR-2 stimulation, whereas the increased proliferation to repeated PAR-2 stimulation is related to increased ERK phosphorylation. PMID:24551046

Accurate control of oxygen level in cells during culture on silicone rubber membranes with application to stem cell differentiation.

PubMed

Powers, Daryl E; Millman, Jeffrey R; Bonner-Weir, Susan; Rappel, Michael J; Colton, Clark K

2010-01-01

Oxygen level in mammalian cell culture is often controlled by placing culture vessels in humidified incubators with a defined gas phase partial pressure of oxygen (pO(2gas)). Because the cells are consuming oxygen supplied by diffusion, a difference between pO(2gas) and that experienced by the cells (pO(2cell)) arises, which is maximal when cells are cultured in vessels with little or no oxygen permeability. Here, we demonstrate theoretically that highly oxygen-permeable silicone rubber membranes can be used to control pO(2cell) during culture of cells in monolayers and aggregates much more accurately and can achieve more rapid transient response following a disturbance than on polystyrene and fluorinated ethylene-propylene copolymer membranes. Cell attachment on silicone rubber was achieved by physical adsorption of fibronectin or Matrigel. We use these membranes for the differentiation of mouse embryonic stem cells to cardiomyocytes and compare the results with culture on polystyrene or on silicone rubber on top of polystyrene. The fraction of cells that are cardiomyocyte-like increases with decreasing pO(2) only when using oxygen-permeable silicone membrane-based dishs, which contract on silicone rubber but not polystyrene. The high permeability of silicone rubber results in pO(2cell) being equal to pO(2gas) at the tissue-membrane interface. This, together with geometric information from histological sections, facilitates development of a model from which the pO(2) distribution within the resulting aggregates is computed. Silicone rubber membranes have significant advantages over polystyrene in controlling pO(2cell), and these results suggest they are a valuable tool for investigating pO(2) effects in many applications, such as stem cell differentiation. Copyright 2009 American Institute of Chemical Engineers

Impact of modeled microgravity on migration, differentiation, and cell cycle control of primitive human hematopoietic progenitor cells.

PubMed

Plett, P Artur; Abonour, Rafat; Frankovitz, Stacy M; Orschell, Christie M

2004-08-01

Migration, proliferation, and differentiation of bone marrow (BM) hematopoietic stem cells (HSC) are important factors in maintaining hematopoietic homeostasis. Homeostatic control of erythrocytes and lymphocytes is perturbed in humans exposed to microgravity (micro-g), resulting in space flight-induced anemia and immunosuppression. We sought to determine whether any of these anomalies can be explained by micro-g-induced changes in migration, proliferation, and differentiation of human BM CD34+ cells, and whether such changes can begin to explain any of the shifts in hematopoietic homeostasis observed in astronauts. BM CD34+ cells were cultured in modeled micro-g (mmicro-g) using NASA's rotating wall vessels (RWV), or in control cultures at earth gravity for 2 to 18 days. Cells were harvested at different times and CD34+ cells assessed for migration potential, cell-cycle kinetics and regulatory proteins, and maturation status. Culture of BM CD34+ cells in RWV for 2 to 3 days resulted in a significant reduction of stromal cell-derived factor 1 (SDF-1alpha)-directed migration, which correlated with decreased expression of F-actin. Modeled micro-g induced alterations in cell-cycle kinetics that were characterized by prolonged S phase and reduced cyclin A expression. Differentiation of primitive CD34+ cells cultured for 14 to 18 days in RWV favored myeloid cell development at the expense of erythroid development, which was significantly reduced compared to controls. These results illustrate that mmicro-g significantly inhibits the migration potential, cell-cycle progression, and differentiation patterns of primitive BM CD34+ cells, which may contribute to some of the hematologic abnormalities observed in humans during space flight.

Accelerated stress testing of amorphous silicon solar cells

NASA Technical Reports Server (NTRS)

Stoddard, W. G.; Davis, C. W.; Lathrop, J. W.

1985-01-01

A technique for performing accelerated stress tests of large-area thin a-Si solar cells is presented. A computer-controlled short-interval test system employing low-cost ac-powered ELH illumination and a simulated a-Si reference cell (seven individually bandpass-filtered zero-biased crystalline PIN photodiodes) calibrated to the response of an a-Si control cell is described and illustrated with flow diagrams, drawings, and graphs. Preliminary results indicate that while most tests of a program developed for c-Si cells are applicable to a-Si cells, spurious degradation may appear in a-Si cells tested at temperatures above 130 C.

TCR tuning of T cell subsets.

PubMed

Cho, Jae-Ho; Sprent, Jonathan

2018-05-01

After selection in the thymus, the post-thymic T cell compartments comprise heterogenous subsets of naive and memory T cells that make continuous T cell receptor (TCR) contact with self-ligands bound to major histocompatibility complex (MHC) molecules. T cell recognition of self-MHC ligands elicits covert TCR signaling and is particularly important for controlling survival of naive T cells. Such tonic TCR signaling is tightly controlled and maintains the cells in a quiescent state to avoid autoimmunity. Here, we review how naive and memory T cells are differentially tuned and wired for TCR sensitivity to self and foreign ligands. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Generation of glucose-sensitive insulin-secreting beta-like cells from human embryonic stem cells by incorporating a synthetic lineage-control network.

PubMed

Saxena, Pratik; Bojar, Daniel; Zulewski, Henryk; Fussenegger, Martin

2017-10-10

We previously reported novel technology to differentiate induced pluripotent stem cells (IPSCs) into glucose-sensitive insulin-secreting beta-like cells by engineering a synthetic lineage-control network regulated by the licensed food additive vanillic acid. This genetic network was able to program intricate expression dynamics of the key transcription factors Ngn3 (neurogenin 3, OFF-ON-OFF), Pdx1 (pancreatic and duodenal homeobox 1, ON-OFF-ON) and MafA (V-maf musculoaponeurotic fibrosarcoma oncogene homologue A, OFF-ON) to guide the differentiation of IPSC-derived pancreatic progenitor cells to beta-like cells. In the present study, we show for the first time that this network can also program the expression dynamics of Ngn3, Pdx1 and MafA in human embryonic stem cell (hESC)-derived pancreatic progenitor cells and drive differentiation of these cells into glucose-sensitive insulin-secreting beta-like cells. Therefore, synthetic lineage-control networks appear to be a robust methodology for differentiating pluripotent stem cells into somatic cell types for basic research and regenerative medicine. Copyright © 2017 Elsevier B.V. All rights reserved.

Lamellipodin and the Scar/WAVE complex cooperate to promote cell migration in vivo

PubMed Central

Law, Ah-Lai; Vehlow, Anne; Kotini, Maria; Dodgson, Lauren; Soong, Daniel; Theveneau, Eric; Bodo, Cristian; Taylor, Eleanor; Navarro, Christel; Perera, Upamali; Michael, Magdalene; Dunn, Graham A.; Bennett, Daimark; Mayor, Roberto

2013-01-01

Cell migration is essential for development, but its deregulation causes metastasis. The Scar/WAVE complex is absolutely required for lamellipodia and is a key effector in cell migration, but its regulation in vivo is enigmatic. Lamellipodin (Lpd) controls lamellipodium formation through an unknown mechanism. Here, we report that Lpd directly binds active Rac, which regulates a direct interaction between Lpd and the Scar/WAVE complex via Abi. Consequently, Lpd controls lamellipodium size, cell migration speed, and persistence via Scar/WAVE in vitro. Moreover, Lpd knockout mice display defective pigmentation because fewer migrating neural crest-derived melanoblasts reach their target during development. Consistently, Lpd regulates mesenchymal neural crest cell migration cell autonomously in Xenopus laevis via the Scar/WAVE complex. Further, Lpd’s Drosophila melanogaster orthologue Pico binds Scar, and both regulate collective epithelial border cell migration. Pico also controls directed cell protrusions of border cell clusters in a Scar-dependent manner. Taken together, Lpd is an essential, evolutionary conserved regulator of the Scar/WAVE complex during cell migration in vivo. PMID:24247431

Sulfonated polyaniline-based organic electrodes for controlled electrical stimulation of human osteosarcoma cells.

PubMed

Min, Yong; Yang, Yanyin; Poojari, Yadagiri; Liu, Yidong; Wu, Jen-Chieh; Hansford, Derek J; Epstein, Arthur J

2013-06-10

Electrically conducting polymers (CPs) were found to stimulate various cell types such as neurons, osteoblasts, and fibroblasts in both in vitro and in vivo studies. However, to our knowledge, no studies have been reported on the utility of CPs in stimulation of cancer or tumor cells in the literature. Here we report a facile fabrication method of self-doped sulfonated polyaniline (SPAN)-based interdigitated electrodes (IDEs) for controlled electrical stimulation of human osteosarcoma (HOS) cells. Increased degree of sulfonation was found to increase the SPAN conductivity, which in turn improved the cell attachment and cell growth without electrical stimulation. However, an enhanced cell growth was observed under controlled electrical (AC) stimulation at low applied voltage and frequency (≤800 mV and ≤1 kHz). The cell growth reached a maximum threshold at an applied voltage or frequency and beyond which pronounced cell death was observed. We believe that these organic electrodes may find utility in electrical stimulation of cancer or tumor cells for therapy and research and may also provide an alternative to the conventional metal-based electrodes.

Clinical significance of Tim3-positive T cell subsets in patients with multiple sclerosis.

PubMed

Feng, Xuemei; Feng, Juan

2016-12-01

The present study evaluated associations between the percentages of T cell immunoglobulin and mucin domain 3 (Tim3)-positive T cells and related cytokines and multiple sclerosis (MS). We collected peripheral blood samples from 30 MS patients and 30 healthy controls. Flow cytometry was used to determine the proportions of CD3 + Tim3 + , CD4 + Tim3 + , and CD4 + CD25 + Tim3 + in peripheral blood mononuclear cells (PBMCs) and related cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ also were determined using enzyme-linked immunosorbent assays (ELISA). The percentages of Tim3-positive T cells in CD4 + and CD4 + CD25 + T cell subsets were significantly lower among MS patients than among controls. This difference was particularly evident in the CD4 + CD25(high) T cell subset. The proportions of CD4 + Tim3 + and CD4 + CD25 + Tim3 + cells in PBMCs were significantly lower in the MS group than in the control group, whereas no significant differences were detected regarding the percentages of CD3 + Tim3 + in PBMCs and T cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ all were increased in MS patients compared with healthy controls. Our results support that Tim3 and related cytokines may be involved in the onset of MS. Copyright © 2016 Elsevier Ltd. All rights reserved.

Chemical synthesis, characterisation, and biocompatibility of nanometre scale porous anodic aluminium oxide membranes for use as a cell culture substrate for the vero cell line: a preliminary study.

PubMed

Poinern, Gérrard Eddy Jai; Le, Xuan Thi; O'Dea, Mark; Becker, Thomas; Fawcett, Derek

2014-01-01

In this preliminary study we investigate for the first time the biomedical potential of using porous anodic aluminium oxide (AAO) membranes as a cell substrate for culturing the Cercopithecus aethiops (African green monkey) Kidney (Vero) epithelial cell line. One advantage of using the inorganic AAO membrane is the presence of nanometre scale pore channels that allow the exchange of molecules and nutrients across the membrane. The size of the pore channels can be preselected by adjusting the controlling parameters of a temperature controlled two-step anodization process. The cellular interaction and response of the Vero cell line with an in-house synthesised AAO membrane, a commercially available membrane, and a glass control were assessed by investigating cell adhesion, morphology, and proliferation over a 72 h period. The number of viable cells proliferating over the respective membrane surfaces revealed that the locally produced in-house AAO membrane had cells numbers similar to the glass control. The study revealed evidence of focal adhesion sites over the surface of the nanoporous membranes and the penetration of cellular extensions into the pore structure as well. The outcome of the study has revealed that nanometre scale porous AAO membranes have the potential to become practical cell culture scaffold substrates with the capability to enhance adhesion and proliferation of Vero cells.

Chemical Synthesis, Characterisation, and Biocompatibility of Nanometre Scale Porous Anodic Aluminium Oxide Membranes for Use as a Cell Culture Substrate for the Vero Cell Line: A Preliminary Study

PubMed Central

Poinern, Gérrard Eddy Jai; Le, Xuan Thi; Becker, Thomas; Fawcett, Derek

2014-01-01

In this preliminary study we investigate for the first time the biomedical potential of using porous anodic aluminium oxide (AAO) membranes as a cell substrate for culturing the Cercopithecus aethiops (African green monkey) Kidney (Vero) epithelial cell line. One advantage of using the inorganic AAO membrane is the presence of nanometre scale pore channels that allow the exchange of molecules and nutrients across the membrane. The size of the pore channels can be preselected by adjusting the controlling parameters of a temperature controlled two-step anodization process. The cellular interaction and response of the Vero cell line with an in-house synthesised AAO membrane, a commercially available membrane, and a glass control were assessed by investigating cell adhesion, morphology, and proliferation over a 72 h period. The number of viable cells proliferating over the respective membrane surfaces revealed that the locally produced in-house AAO membrane had cells numbers similar to the glass control. The study revealed evidence of focal adhesion sites over the surface of the nanoporous membranes and the penetration of cellular extensions into the pore structure as well. The outcome of the study has revealed that nanometre scale porous AAO membranes have the potential to become practical cell culture scaffold substrates with the capability to enhance adhesion and proliferation of Vero cells. PMID:24579077

Involvement of DNA methylation in the control of cell growth during heat stress in tobacco BY-2 cells.

PubMed

Centomani, Isabella; Sgobba, Alessandra; D'Addabbo, Pietro; Dipierro, Nunzio; Paradiso, Annalisa; De Gara, Laura; Dipierro, Silvio; Viggiano, Luigi; de Pinto, Maria Concetta

2015-11-01

The alteration of growth patterns, through the adjustment of cell division and expansion, is a characteristic response of plants to environmental stress. In order to study this response in more depth, the effect of heat stress on growth was investigated in tobacco BY-2 cells. The results indicate that heat stress inhibited cell division, by slowing cell cycle progression. Cells were stopped in the pre-mitotic phases, as shown by the increased expression of CycD3-1 and by the decrease in the NtCycA13, NtCyc29 and CDKB1-1 transcripts. The decrease in cell length and the reduced expression of Nt-EXPA5 indicated that cell expansion was also inhibited. Since DNA methylation plays a key role in controlling gene expression, the possibility that the altered expression of genes involved in the control of cell growth, observed during heat stress, could be due to changes in the methylation state of their promoters was investigated. The results show that the altered expression of CycD3-1 and Nt-EXPA5 was consistent with changes in the methylation state of the upstream region of these genes. These results suggest that DNA methylation, controlling the expression of genes involved in plant development, contributes to growth alteration occurring in response to environmental changes.

Early Gag Immunodominance of the HIV-Specific T-Cell Response during Acute/Early Infection Is Associated with Higher CD8+ T-Cell Antiviral Activity and Correlates with Preservation of the CD4+ T-Cell Compartment

PubMed Central

Ghiglione, Yanina; Falivene, Juliana; Socias, María Eugenia; Laufer, Natalia; Coloccini, Romina Soledad; Rodriguez, Ana María; Ruiz, María Julia; Pando, María Ángeles; Giavedoni, Luis David; Cahn, Pedro; Sued, Omar; Salomon, Horacio; Gherardi, María Magdalena

2013-01-01

The important role of the CD8+ T-cell response on HIV control is well established. Moreover, the acute phase of infection represents a proper scenario to delineate the antiviral cellular functions that best correlate with control. Here, multiple functional aspects (specificity, ex vivo viral inhibitory activity [VIA] and polyfunctionality) of the HIV-specific CD8+ T-cell subset arising early after infection, and their association with disease progression markers, were examined. Blood samples from 44 subjects recruited within 6 months from infection (primary HIV infection [PHI] group), 16 chronically infected subjects, 11 elite controllers (EC), and 10 healthy donors were obtained. Results indicated that, although Nef dominated the anti-HIV response during acute/early infection, a higher proportion of early anti-Gag T cells correlated with delayed progression. Polyfunctional HIV-specific CD8+ T cells were detected at early time points but did not associate with virus control. Conversely, higher CD4+ T-cell set points were observed in PHI subjects with higher HIV-specific CD8+ T-cell VIA at baseline. Importantly, VIA levels correlated with the magnitude of the anti-Gag cellular response. The advantage of Gag-specific cells may result from their enhanced ability to mediate lysis of infected cells (evidenced by a higher capacity to degranulate and to mediate VIA) and to simultaneously produce IFN-γ. Finally, Gag immunodominance was associated with elevated plasma levels of interleukin 2 (IL-2) and macrophage inflammatory protein 1β (MIP-1β). All together, this study underscores the importance of CD8+ T-cell specificity in the improved control of disease progression, which was related to the capacity of Gag-specific cells to mediate both lytic and nonlytic antiviral mechanisms at early time points postinfection. PMID:23616666

Interactions between insulin-like growth factor-I, estrogen receptor-α (ERα) and ERβ in regulating growth/apoptosis of MCF-7 human breast cancer cells

PubMed Central

Mendoza, Rhone A.; Enriquez, Marlene I; Mejia, Sylvia M; Moody, Emily E; Thordarson, Gudmundur

2011-01-01

Understanding of the interactions between estradiol (E2) and insulin-like growth factor-I (IGF-I) is still incomplete. Cell lines derived from the MCF-7 breast cancer cells were generated with suppressed expression of the IGF-I receptor (IGF-IR), termed IGF-IR.low cells, by stable transfection using small interfering RNA (siRNA) expression vector. Vector for control cells carried sequence generating non-interfering RNA. Concomitant with reduction in the IGF-IR levels, the IGF-IR.low cells also showed a reduction in estrogen receptor α (ERα) and progesterone receptor expressions and an elevation in the expression of ERβ. The number of the IGF-IR.low cells was reduced in response to IGF-I and human growth hormone plus epidermal growth factor, but E2 did not cause increase in the number of the IGF-IR.low cells compared to controls. Proliferation rate of IGF-IR.low cells was only reduced in response to E2 compared to controls, whereas their basal and hormone stimulated apoptosis rate was increased. Phosphorylation of p38 mitogen activated protein kinase (p38 MAPK) was increased in the IGF-IR.low cells after treatment with E2, without affecting control cells. Further, phosphorylation of the tumor suppressor protein p53 was elevated in the IGF-IR.low cells compared to the controls. Summary, suppressing the IGF-IR expression decreased the level of ERα but increased the level of ERβ. Overall growth rate of the IGF-IR.low cells was reduced mostly through an increase in apoptosis without affecting proliferation substantially. We hypothesize that a decreased ERα:ERβ ratio triggered a rapid phosphorylation of p38 MAPK which in turn phosphorylated the p53 tumor suppressor and accelerated apoptosis rate. PMID:20974640

Oxalomalate, a competitive inhibitor of NADP+ -dependent isocitrate dehydrogenase, regulates lipid peroxidation-mediated apoptosis in U937 cells.

PubMed

Yang, Eun Sun; Yang, Joon-Hyuck; Park, Ji Eun; Park, Jeen-Woo

2005-01-01

Membrane lipid peroxidation processes yield products that may react with DNA and proteins to cause oxidative modifications. Recently, we demonstrated that the control of cytosolic redox balance and the cellular defense against oxidative damage is one of the primary functions of cytosolic NADP+ -dependent isocitrate dehydrogenase (IDPc) through to supply NADPH for antioxidant systems. The protective role of IDPc against lipid peroxidation-mediated apoptosis in U937 cells was investigated in control and cells pre-treated with oxlalomalate, a competitive inhibitor of IDPc. Upon exposure to 2,2'-azobis (2-amidinopropane) hydrochloride (AAPH) to U937 cells, which induces lipid peroxidation in membranes, the susceptibility to apoptosis was higher in oxalomalate-treated cells as compared to control cells. The results suggest that IDPc plays an important protective role in apoptosis of U937 cells induced by lipid peroxidation-mediated oxidative stress.

Effect of Thermodiffusion Nitriding on Cytocompatibility of Ti-6Al-4V Titanium Alloy

NASA Astrophysics Data System (ADS)

Pohrelyuk, I. M.; Tkachuk, O. V.; Proskurnyak, R. V.; Boiko, N. M.; Kluchivska, O. Yu.; Stoika, R. S.

2016-04-01

The nitrided layer was formed on the surface of Ti-6Al-4V titanium alloy by the thermodiffusion saturation in nitrogen at the atmospheric pressure. The study of the vitality of pseudonormal human embryo kidney cells of the HEK293T line showed that their cultivation in the presence of the untreated alloy sample is accompanied by a statistically significant reduction in the number of living cells compared with the control sample (untreated cells), whereas their cultivation in the presence of the nitrided alloy sample does not change the cell number considerably. In addition, it was shown that cell behavior in the presence of the nitrided sample differs only slightly from the control sample, whereas the growth of cells in the presence of the untreated alloy differed significantly from that in the control sample, demonstrating small groups of cells instead of their big clusters.

Toxicity and Radioprotective Effects of DF-1 and Carbon Nanotubes in Human Lung and Liver Cell Lines

NASA Technical Reports Server (NTRS)

Burgoyne, Madeline; Holtorf, Heidi; Huff, Janice; Moore, Valerie; Jeevarajan, Antony

2007-01-01

The DF-1 compound, a sixty carbon fullerene derivative, has been shown to have antioxidant effects and is thought to possibly help mediate the effects of radiation on cells. While this is potentially useful, it is important to first understand the effect that the DF-1 has on the cells and the growth rate of the cells to determine if the material itself has any innate toxicity. A growth curve was established for both HF-19 cells, human fibroblasts, and HepG2 cells, liver tissue cells in the presence of two different concentrations of DF-1 and for untreated controls. The cells were plated in triplicate in 60mm dishes and were lifted and counted with a hemocytometer daily for one week. The growth curve data for the HF-19 cells show that while the low concentration of DF-1 had no apparent effect on the growth rate, the high concentration of DF-1 appeared to severely inhibit the growth of the HF-19 cells. The growth curve data for the HepG2 cells shows that the DF-1 compound had no significant effect on the rate at which the cells grew. A second growth curve study was performed plain carbon nanotubes, but with only 24 hour exposure to a high and low concentration of material. The carbon nanotubes are another carbon compound similar to DF-1, but in the shape of a tube, rather than a ball. We hypothesize that nanotubes may also mediate the effect of radiation on cells. This time, nanotubes did not showed any significant effect on the growth rate HF-19 or HepG2 cells. A third growth curve study is underway to further determine the effect of DF-1, nanotubes, and a derivatized nanotube (BHT-nanotubes). This derivatized nanotube has been modified with a compound that is known to be very effective at neutralizing free radicals. We expect that the high concentration of DF-1 and possibly the nanotubes and BHT-nanotubes may inhibit the growth of the HF-19 cells while the low concentration will resemble the growth of the control. We also hypothesize that there will be no significant effect on the growth of the HepG2 cells by the nanotubes, and BHT-nanotubes. In order to examine the usefulness of the DF-1, nanotubes, and BHT-nanotubes in mediating the effects of radiation a clonogenic assay is being performed. The HF-19 cells were plated in different concentrations of the various compounds and exposed to varying amounts of radiation. The cells are being allowed to grow in a small enough concentration so that the ability of each cell to divide can be seen by the development of cell clusters. By comparing the irradiated control to the un-irradiated control the effects of radiation alone can be seen. By comparing the compound treated irradiated cells to the irradiated control the usefulness of each compound can be seen. It is thought that Amifostine, the positive control, will have more regularly dividing cells then the irradiated control, as will DF-1 and hopefully both nanotube materials as well.

[A skin cell segregating control system based on PC].

PubMed

Liu, Wen-zhong; Zhou, Ming; Zhang, Hong-bing

2005-11-01

A skin cell segregating control system based on PC (personal computer) is presented in this paper. Its front controller is a single-chip microcomputer which enables the manipulation for 6 patients simultaneously, and thus provides a great convenience for clinical treatments for vitiligo. With the use of serial port communication technology, it's possible to monitor and control the front controller in a PC terminal. And the application of computer image acquisition technology realizes the synchronous acquisition of pathologic shin cell images pre/after the operation and a case history. Clinical tests prove its conformity with national standards and the pre-set technological requirements.

Some Kinds of Cancer Kids Get

MedlinePlus

... normal grow too fast and spread out of control. A group or mass of growing cells is called a ... person, these white blood cells multiply out of control. They fill up the bone ... Cancer A brain tumor is a group or clump of fast-growing cells that can ...

Suppression of BRCA2 by Mutant Mitochondrial DNA in Prostate Cancer

DTIC Science & Technology

2014-07-01

growth of prostatic epithelia both in vitro and in vivo To evaluate the impact of interaction between DAB2IP and Skp2 on cell growth , MTT assay and soft...determined using western blot and actin was used as a loading control. One thousand cells /well were seeded using 96-well plate. In vitro cell growth ...SEM. (E) 1 × 103 cells of C4-2 shSkp2 cells and its control were seeded at 96-well plate. In vitro cell growth was determined using

A co-culture device with a tunable stiffness to understand combinatorial cell-cell and cell-matrix interactions.

PubMed

Rao, Nikhil; Grover, Gregory N; Vincent, Ludovic G; Evans, Samantha C; Choi, Yu Suk; Spencer, Katrina H; Hui, Elliot E; Engler, Adam J; Christman, Karen L

2013-11-01

Cell behavior on 2-D in vitro cultures is continually being improved to better mimic in vivo physiological conditions by combining niche cues including multiple cell types and substrate stiffness, which are well known to impact cell phenotype. However, no system exists in which a user can systematically examine cell behavior on a substrate with a specific stiffness (elastic modulus) in culture with a different cell type, while maintaining distinct cell populations. We demonstrate the modification of a silicon reconfigurable co-culture system with a covalently linked hydrogel of user-defined stiffness. This device allows the user to control whether two separate cell populations are in contact with each other or only experience paracrine interactions on substrates of controllable stiffness. To illustrate the utility of this device, we examined the role of substrate stiffness combined with myoblast co-culture on adipose derived stem cell (ASC) differentiation and found that the presence of myoblasts and a 10 kPa substrate stiffness increased ASC myogenesis versus co-culture on stiff substrates. As this example highlights, this technology better controls the in vitro microenvironment, allowing the user to develop a more thorough understanding of the combined effects of cell-cell and cell-matrix interactions.

Surface receptor Toso controls B cell-mediated regulation of T cell immunity.

PubMed

Yu, Jinbo; Duong, Vu Huy Hoang; Westphal, Katrin; Westphal, Andreas; Suwandi, Abdulhadi; Grassl, Guntram A; Brand, Korbinian; Chan, Andrew C; Föger, Niko; Lee, Kyeong-Hee

2018-05-01

The immune system is tightly controlled by regulatory processes that allow for the elimination of invading pathogens, while limiting immunopathological damage to the host. In the present study, we found that conditional deletion of the cell surface receptor Toso on B cells unexpectedly resulted in impaired proinflammatory T cell responses, which led to impaired immune protection in an acute viral infection model and was associated with reduced immunopathological tissue damage in a chronic inflammatory context. Toso exhibited its B cell-inherent immunoregulatory function by negatively controlling the pool of IL-10-competent B1 and B2 B cells, which were characterized by a high degree of self-reactivity and were shown to mediate immunosuppressive activity on inflammatory T cell responses in vivo. Our results indicate that Toso is involved in the differentiation/maintenance of regulatory B cells by fine-tuning B cell receptor activation thresholds. Furthermore, we showed that during influenza A-induced pulmonary inflammation, the application of Toso-specific antibodies selectively induced IL-10-competent B cells at the site of inflammation and resulted in decreased proinflammatory cytokine production by lung T cells. These findings suggest that Toso may serve as a novel therapeutic target to dampen pathogenic T cell responses via the modulation of IL-10-competent regulatory B cells.

Peptidases released by necrotic cells control CD8+ T cell cross-priming

PubMed Central

Gamrekelashvili, Jaba; Kapanadze, Tamar; Han, Miaojun; Wissing, Josef; Ma, Chi; Jaensch, Lothar; Manns, Michael P.; Armstrong, Todd; Jaffee, Elizabeth; White, Ayla O.; Citrin, Deborah E.; Korangy, Firouzeh; Greten, Tim F.

2013-01-01

Cross-priming of CD8+ T cells and generation of effector immune responses is pivotal for tumor immunity as well as for successful anticancer vaccination and therapy. Dead and dying cells produce signals that can influence Ag processing and presentation; however, there is conflicting evidence regarding the immunogenicity of necrotic cell death. We used a mouse model of sterile necrosis, in which mice were injected with sterile primary necrotic cells, to investigate a role of these cells in priming of CD8+ T cells. We discovered a molecular mechanism operating in Ag donor cells that regulates cross-priming of CD8+ T cells during primary sterile necrosis and thereby controls adaptive immune responses. We found that the cellular peptidases dipeptidyl peptidase 3 (DPP-3) and thimet oligopeptidase 1 (TOP-1), both of which are present in nonimmunogenic necrotic cells, eliminated proteasomal degradation products and blocked Ag cross-presentation. While sterile necrotic tumor cells failed to induce CD8+ T cell responses, their nonimmunogenicity could be reversed in vitro and in vivo by inactivation of DPP-3 and TOP-1. These results indicate that control of cross-priming and thereby immunogenicity of primary sterile necrosis relies on proteasome-dependent oligopeptide generation and functional status of peptidases in Ag donor cells. PMID:24216478

Peptidases released by necrotic cells control CD8+ T cell cross-priming.

PubMed

Gamrekelashvili, Jaba; Kapanadze, Tamar; Han, Miaojun; Wissing, Josef; Ma, Chi; Jaensch, Lothar; Manns, Michael P; Armstrong, Todd; Jaffee, Elizabeth; White, Ayla O; Citrin, Deborah E; Korangy, Firouzeh; Greten, Tim F

2013-11-01

Cross-priming of CD8+ T cells and generation of effector immune responses is pivotal for tumor immunity as well as for successful anticancer vaccination and therapy. Dead and dying cells produce signals that can influence Ag processing and presentation; however, there is conflicting evidence regarding the immunogenicity of necrotic cell death. We used a mouse model of sterile necrosis, in which mice were injected with sterile primary necrotic cells, to investigate a role of these cells in priming of CD8+ T cells. We discovered a molecular mechanism operating in Ag donor cells that regulates cross-priming of CD8+ T cells during primary sterile necrosis and thereby controls adaptive immune responses. We found that the cellular peptidases dipeptidyl peptidase 3 (DPP-3) and thimet oligopeptidase 1 (TOP-1), both of which are present in nonimmunogenic necrotic cells, eliminated proteasomal degradation products and blocked Ag cross-presentation. While sterile necrotic tumor cells failed to induce CD8+ T cell responses, their nonimmunogenicity could be reversed in vitro and in vivo by inactivation of DPP-3 and TOP-1. These results indicate that control of cross-priming and thereby immunogenicity of primary sterile necrosis relies on proteasome-dependent oligopeptide generation and functional status of peptidases in Ag donor cells.

Drug-loaded nanoparticles induce gene expression in human pluripotent stem cell derivatives

NASA Astrophysics Data System (ADS)

Gajbhiye, Virendra; Escalante, Leah; Chen, Guojun; Laperle, Alex; Zheng, Qifeng; Steyer, Benjamin; Gong, Shaoqin; Saha, Krishanu

2013-12-01

Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives.Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives. Electronic supplementary information (ESI) available: ESI containing 1H NMR spectra and additional fibroblast characterization data. See DOI: 10.1039/c3nr04794f

Expression of natural killer cell activity with CD107a on ectopic endometrium in woman with endometriosis compared with non-endometriosis

NASA Astrophysics Data System (ADS)

Lubis, H. P.; Aldiansyah, D.; Siregar, H. S.; Rivany, R.; Hariadi, T. S.

2018-03-01

Some factors have an important role in endometriosis pathogenesis; there is an immune cell that plays an important role in endometrial cells that have reflux. Woman with endometriosis experienced the cellular immune disorder. It is suspected that decrease of NK cell in the peritoneal fluid caused by its qualitative defect with CD107a expression as the best marker. The aim of this study was to compare expression of NK Cell activity with CD107a between awoman with endometriosis and non-endometriosis. A case-control study from March until July 2015 in Haji Adam Malik General Hospital. The case group was ectopic endometrial tissue block paraffin and control group was normal endometrial tissue block paraffin. This study included 23 patients in endometriosis group and control group respectively. A majority proportion of CD107a expression in endometriosis group was +1 (16 patients (69.6%)), while the control group was +3 (9 patients (39.1%)). Expression of NK cell activity with CD107a in patients with endometriosis was lower than the control group (p<0.05). It suggested that cellular immune factors may play a role in the pathogenesis of endometriosis.

[Microtubules suppress blebbing and stimulate lamellae extension in spreading fibroblasts].

PubMed

Tvorogova, A V; Vorob'ev, I A

2012-01-01

We compared spreading of Vero fibroblasts when microtubules were depolymerized or stabilized. After initial attachment cells start blebbing that continues for different time and abruptly transfers into spreading. After spreading initiation, most cells spread in an anisotropic manner through stochastic formation of lamellipodia. A second mode was rapid, isotropic spreading via formation of circular lamellum that occurs in 15% of cells. The rate of spreading was maximal at the beginning and decreased during the first hour according to logarithmic law. After 60 min many cells formed stable efges and started migrating on the substrate. However, cell area slowly continued to increase. Actin bundles are formed 20 min after cell attachment and they first run along cell boundary. This system disassembles within 20-40 min and is substituted with stress fibers crossing the cell. In the isotropically spread cells no actin bunbles are seen. Microtubules in the spreading cells enter into large blebs and all nascent lamella and later form radial array. When MTs has been depolymerized or stabilized blebbing started before cells attached to the substrate and continue much longer than in control cells. In both cases the initial rate of spreading decrease several fold, and remains constant for many hours. After 24 h the mean area occupied by cells with altered MT system was the same as in control. Alteration of MT system had moderate effect on actin system--formation of actin cables started at the same time as in control (within 20 min upon cell attachment), however, they grew even in cells undergoing prolonged blebbing. Actin cables running along cell margin were similar to tat in control cells, but they did not disappear up to 1 h. When stabilized, microtubules form chaotic array: they do not enter blebs and in spread cells run parallel to the cell margin at a distance of 3-5 microm. We conclude that dynamic microtubules speed up completion of blebbing and promote early stages of fibroblasts spreading.

The effect of well-characterized, very low-dose x-ray radiation on fibroblasts

PubMed Central

Truong, Katelyn; Bradley, Suzanne; Baginski, Bryana; Wilson, Joseph R.; Medlin, Donald; Zheng, Leon; Wilson, R. Kevin; Rusin, Matthew; Takacs, Endre

2018-01-01

The purpose of this study is to determine the effects of low-dose radiation on fibroblast cells irradiated by spectrally and dosimetrically well-characterized soft x-rays. To achieve this, a new cell culture x-ray irradiation system was designed. This system generates characteristic fluorescent x-rays to irradiate the cell culture with x-rays of well-defined energies and doses. 3T3 fibroblast cells were cultured in cups with Mylar® surfaces and were irradiated for one hour with characteristic iron (Fe) K x-ray radiation at a dose rate of approximately 550 μGy/hr. Cell proliferation, total protein analysis, flow cytometry, and cell staining were performed on fibroblast cells to determine the various effects caused by the radiation. Irradiated cells demonstrated increased proliferation and protein production compared to control samples. Flow cytometry revealed that a higher percentage of irradiated cells were in the G0/G1 phase of the cell cycle compared to control counterparts, which is consistent with other low-dose studies. Cell staining results suggest that irradiated cells maintained normal cell functions after radiation exposure, as there were no qualitative differences between the images of the control and irradiated samples. The result of this study suggest that low-dose soft x-ray radiation might cause an initial pause, followed by a significant increase, in proliferation. An initial “pause” in cell proliferation could be a protective mechanism of the cells to minimize DNA damage caused by radiation exposure. The new cell irradiation system developed here allows for unprecedented control over the properties of the x-rays given to the cell cultures. This will allow for further studies on various cell types with known spectral distribution and carefully measured doses of radiation, which may help to elucidate the mechanisms behind varied cell responses to low-dose x-rays reported in the literature. PMID:29300773

Miniature Bioreactor System for Long-Term Cell Culture

NASA Technical Reports Server (NTRS)

Gonda, Steve R.; Kleis, Stanley J.; Geffert, Sandara K.

2010-01-01

A prototype miniature bioreactor system is designed to serve as a laboratory benchtop cell-culturing system that minimizes the need for relatively expensive equipment and reagents and can be operated under computer control, thereby reducing the time and effort required of human investigators and reducing uncertainty in results. The system includes a bioreactor, a fluid-handling subsystem, a chamber wherein the bioreactor is maintained in a controlled atmosphere at a controlled temperature, and associated control subsystems. The system can be used to culture both anchorage-dependent and suspension cells, which can be either prokaryotic or eukaryotic. Cells can be cultured for extended periods of time in this system, and samples of cells can be extracted and analyzed at specified intervals. By integrating this system with one or more microanalytical instrument(s), one can construct a complete automated analytical system that can be tailored to perform one or more of a large variety of assays.

CacyBP/SIP nuclear translocation induced by gastrin promotes gastric cancer cell proliferation

PubMed Central

Zhai, Hui-Hong; Meng, Juan; Wang, Jing-Bo; Liu, Zhen-Xiong; Li, Yuan-Fei; Feng, Shan-Shan

2014-01-01

AIM: To investigate the role of nuclear translocation of calcyclin binding protein, also called Siah-1 interacting protein (CacyBP/SIP), in gastric carcinogenesis. METHODS: The expression of CacyBP/SIP protein in gastric cancer cell lines was detected by Western blot. Immunofluorescence experiments were performed on gastric cancer cell lines that had been either unstimulated or stimulated with gastrin. To confirm the immunofluorescence findings, the relative abundance of CacyBP/SIP in nuclear and cytoplasmic compartments was assessed by Western blot. The effect of nuclear translocation of CacyBP/SIP on cell proliferation was examined using MTT assay. The colony formation assay was used to measure clonogenic cell survival. The effect of CacyBP/SIP nuclear translocation on cell cycle progression was investigated. Two CacyBP/SIP-specific siRNA vectors were designed and constructed to inhibit CacyBP/SIP expression in order to reduce the nuclear translocation of CacyBP/SIP, and the expression of CacyBP/SIP in stably transfected cells was determined by Western blot. The effect of inhibiting CacyBP/SIP nuclear translocation on cell proliferation was then assessed. RESULTS: CacyBP/SIP protein was present in most of gastric cancer cell lines. In unstimulated cells, CacyBP/SIP was distributed throughout the cytoplasm; while in stimulated cells, CacyBP/SIP was found mainly in the perinuclear region. CacyBP/SIP nuclear translocation generated a growth-stimulatory effect on cells. The number of colonies in the CacyBP/SIP nuclear translocation group was significantly higher than that in the control group. The percentage of stimulated cells in G1 phase was significantly lower than that of control cells (69.70% ± 0.46% and 65.80% ± 0.60%, control cells and gastrin-treated SGC7901 cells, P = 0.008; 72.99% ± 0.46% and 69.36% ± 0.51%, control cells and gastrin-treated MKN45 cells, P = 0.022). CacyBP/SIPsi1 effectively down-regulated the expression of CacyBP/SIP, and cells stably transfected by CacyBP/SIPsi1 were then chosen for further cellular assays. In CacyBP/SIPsi1 stably transfected cells, CacyBP/SIP was shown to be distributed throughout the cytoplasm, irregardless of whether they were stimulated or not. After CacyBP/SIP nuclear translocation was reduced, there had no major effect on cell proliferation, as shown by MTT assay. There had no enhanced anchorage-dependent growth upon stimulation, as indicated by colony formation in flat plates. No changes appeared in the percentage of cells in G0-G1 phase in either cell line (71.09% ± 0.16% and 70.86% ± 0.25%, control cells and gastrin-treated SGC7901-CacyBP/SIPsi1 cells, P = 0.101; 74.17% ± 1.04% and 73.07% ± 1.00%, control cells and gastrin-treated MKN45-CacyBP/SIPsi1 cells, P = 0.225). CONCLUSION: CacyBP/SIP nuclear translocation promotes the proliferation and cell cycle progression of gastric cancer cells. PMID:25110433

CacyBP/SIP nuclear translocation induced by gastrin promotes gastric cancer cell proliferation.

PubMed

Zhai, Hui-Hong; Meng, Juan; Wang, Jing-Bo; Liu, Zhen-Xiong; Li, Yuan-Fei; Feng, Shan-Shan

2014-08-07

To investigate the role of nuclear translocation of calcyclin binding protein, also called Siah-1 interacting protein (CacyBP/SIP), in gastric carcinogenesis. The expression of CacyBP/SIP protein in gastric cancer cell lines was detected by Western blot. Immunofluorescence experiments were performed on gastric cancer cell lines that had been either unstimulated or stimulated with gastrin. To confirm the immunofluorescence findings, the relative abundance of CacyBP/SIP in nuclear and cytoplasmic compartments was assessed by Western blot. The effect of nuclear translocation of CacyBP/SIP on cell proliferation was examined using MTT assay. The colony formation assay was used to measure clonogenic cell survival. The effect of CacyBP/SIP nuclear translocation on cell cycle progression was investigated. Two CacyBP/SIP-specific siRNA vectors were designed and constructed to inhibit CacyBP/SIP expression in order to reduce the nuclear translocation of CacyBP/SIP, and the expression of CacyBP/SIP in stably transfected cells was determined by Western blot. The effect of inhibiting CacyBP/SIP nuclear translocation on cell proliferation was then assessed. CacyBP/SIP protein was present in most of gastric cancer cell lines. In unstimulated cells, CacyBP/SIP was distributed throughout the cytoplasm; while in stimulated cells, CacyBP/SIP was found mainly in the perinuclear region. CacyBP/SIP nuclear translocation generated a growth-stimulatory effect on cells. The number of colonies in the CacyBP/SIP nuclear translocation group was significantly higher than that in the control group. The percentage of stimulated cells in G1 phase was significantly lower than that of control cells (69.70% ± 0.46% and 65.80% ± 0.60%, control cells and gastrin-treated SGC7901 cells, P = 0.008; 72.99% ± 0.46% and 69.36% ± 0.51%, control cells and gastrin-treated MKN45 cells, P = 0.022). CacyBP/SIPsi1 effectively down-regulated the expression of CacyBP/SIP, and cells stably transfected by CacyBP/SIPsi1 were then chosen for further cellular assays. In CacyBP/SIPsi1 stably transfected cells, CacyBP/SIP was shown to be distributed throughout the cytoplasm, irregardless of whether they were stimulated or not. After CacyBP/SIP nuclear translocation was reduced, there had no major effect on cell proliferation, as shown by MTT assay. There had no enhanced anchorage-dependent growth upon stimulation, as indicated by colony formation in flat plates. No changes appeared in the percentage of cells in G0-G1 phase in either cell line (71.09% ± 0.16% and 70.86% ± 0.25%, control cells and gastrin-treated SGC7901-CacyBP/SIPsi1 cells, P = 0.101; 74.17% ± 1.04% and 73.07% ± 1.00%, control cells and gastrin-treated MKN45-CacyBP/SIPsi1 cells, P = 0.225). CacyBP/SIP nuclear translocation promotes the proliferation and cell cycle progression of gastric cancer cells.

Innate control of adaptive immunity: Beyond the three-signal paradigm

PubMed Central

Jain, Aakanksha; Pasare, Chandrashekhar

2017-01-01

Activation of cells in the adaptive immune system is a highly orchestrated process dictated by multiples cues from the innate immune system. Although the fundamental principles of innate control of adaptive immunity are well established, it is not fully understood how innate cells integrate qualitative pathogenic information in order to generate tailored protective adaptive immune responses. In this review, we discuss complexities involved in the innate control of adaptive immunity that extend beyond T cell receptor engagement, co-stimulation and priming cytokine production but are critical for generation of protective T cell immunity. PMID:28483987

Morphological properties of vestibulospinal neurons in primates

NASA Technical Reports Server (NTRS)

Boyle, Richard; Johanson, Curt

2003-01-01

The lateral and medial vestibulospinal tracts constitute the major descending pathways controlling extensor musculature of the body. We examined the axon morphology and synaptic input patterns and targets in the cervical spinal segments from these tract cells using intracellular recording and biocytin labeling in the squirrel monkey. Lumbosacral projecting cells represent a private, and mostly rapid, communication pathway between the dorsal Deiters' nucleus and the motor circuits controlling the lower limbs and tail. The cervical projecting cells provide both redundant and variable synaptic input to spinal cell groups, suggesting both general and specific control of the head and neck reflexes.

Overexpression of SASH1 related to the decreased invasion ability of human glioma U251 cells.

PubMed

Yang, Liu; Liu, Mei; Gu, Zhikai; Chen, Jianguo; Yan, Yaohua; Li, Jian

2012-12-01

The purpose of this study was to investigate the impact of SAM- and SH3-domain containing 1 (SASH1) on the biological behavior of glioma cells, including its effects on cellular growth, proliferation, apoptosis, invasion, and metastasis, and thereby to provide an experimental basis for future therapeutic treatments. A pcDNA3.1-SASH1 eukaryotic expression vector was constructed and transfected into the U251 human glioma cell line. Using the tetrazolium-based colorimetric (MTT) assay, flow cytometry analyses, transwell invasion chamber experiments, and other methods, we examined the impact of SASH1 on the biological behaviors of U251 cells, including effects on viability, cell cycle, apoptosis, and invasion. Furthermore, the effect of SASH1 on the expression of cyclin D1, caspase-3, matrix metalloproteinase (MMP)-2, MMP-9, and other proteins was observed. Compared to the empty vector and blank control groups, the pcDNA3.1-SASH1 group of U251 cells exhibited significantly reduced cell viability, proliferation, and invasion (p < 0.05), although there was no difference between the empty vector and blank control groups. The pcDNA3.1-SASH1 group demonstrated a significantly higher apoptotic index than did the empty vector and blank control groups (p < 0.05), and the percentage of apoptotic cells was similar between the empty vector and blank control groups. In addition, the pcDNA3.1-SASH1 group expressed significantly lower protein levels of cyclin D1 and MMP-2/9 compared to the control and empty vector groups (p < 0.05) and significantly higher protein levels of caspase-3 than the other two groups (p < 0.05). Cyclin D1, caspase-3, and MMP-2/9 expression was unchanged between the empty vector and blank control groups. SASH1 gene expression might be related to the inhibition of the growth, proliferation, and invasion of U251 cells and the promotion of U251 cells apoptosis.

Supraphysiologic control over HIV-1 replication mediated by CD8 T cells expressing a re-engineered CD4-based chimeric antigen receptor

PubMed Central

Richardson, Max W.; Ellebrecht, Christoph T.; Glover, Joshua A.; Secreto, Anthony J.; Kulikovskaya, Irina; Yi, Yanjie; Wang, Jianbin; Dufendach, Keith A.; Holmes, Michael C.; Collman, Ronald G.

2017-01-01

HIV is adept at avoiding naturally generated T cell responses; therefore, there is a need to develop HIV-specific T cells with greater potency for use in HIV cure strategies. Starting with a CD4-based chimeric antigen receptor (CAR) that was previously used without toxicity in clinical trials, we optimized the vector backbone, promoter, HIV targeting moiety, and transmembrane and signaling domains to determine which components augmented the ability of T cells to control HIV replication. This re-engineered CAR was at least 50-fold more potent in vitro at controlling HIV replication than the original CD4 CAR, or a TCR-based approach, and substantially better than broadly neutralizing antibody-based CARs. A humanized mouse model of HIV infection demonstrated that T cells expressing optimized CARs were superior at expanding in response to antigen, protecting CD4 T cells from infection, and reducing viral loads compared to T cells expressing the original, clinical trial CAR. Moreover, in a humanized mouse model of HIV treatment, CD4 CAR T cells containing the 4-1BB costimulatory domain controlled HIV spread after ART removal better than analogous CAR T cells containing the CD28 costimulatory domain. Together, these data indicate that potent HIV-specific T cells can be generated using improved CAR design and that CAR T cells could be important components of an HIV cure strategy. PMID:29023549

[Protective effect of hydrogen against hyperoxia-induced type II alveolar epithelial cell injury].

PubMed

Yao, Lan; Xu, Feng; Luo, Chong; Yu, Pan; Dong, Xinxin; Sun, Xuejun; Liu, Chengjun

2013-02-01

To investigate the protective effect of hydrogen against hyperoxia-induced oxidative stress injury in premature rat type II alveolar epithelial cells (AECs). The type II AECs isolated from premature rats were randomly divided into air (21% oxygen) control group, hyperoxia (95% oxygen) control group, air + hydrogen group, and hyperoxia+ hydrogen group. The cells with hydrogen treatment were cultured in the presence of rich hydrogen. After the corresponding exposure for 24 h, the cell morphology was observed microscopically. MTT assay was used to evaluated the cell proliferation ability, and JC-1 fluorescence probe was used to detect the mitochondrial membrane potential (δφ) changes of the type II AECs. The concentration of maleic dialdehyde (MDA) and superoxide dismutase (SOD) activity in the cell supernatant were detected using colorimetric method. No significant differences were found in cell growth or measurements between air control and air + hydrogen groups. Compared with air control group, the cells exposed to hyperoxia showed significantly suppressed proliferation, reduced mitochondrial membrane potential, increased MDA content, and decreased SOD activity. Intervention with hydrogen resulted in significantly increased cell proliferation and SOD activity and lowered MDA content, and restored the mitochondrial membrane potential in the cells with hyperoxia exposure (P<0.05). Hydrogen can significantly reduce hyperoxia-induced oxidative stress injury in premature rat type II AECs, improve the cellular antioxidant capacity, stabilize the mitochondrial membrane potential, and reduce the inhibitory effect of hyperoxia on cell proliferation.

[Basic studies on oral administration of lentinan (I)--influence on lymphocyte subsets in peripheral venous blood].

PubMed

Hanaue, H; Tokuda, Y; Machimura, T; Tsukui, M; Mizutani, K; Huang, C M; Kamijoh, A; Kondo, Y; Ogoshi, K; Makuuchi, H

1989-08-20

The effect of oral administration of lentinan (LTN), a biological response modifier, in the control of systemic immune function was studied in 6-week old male Wistar-Imamichi SPF rats. In the LTN group, 1 mg LTN dissolved in 1 ml physiological saline was administration forcibly into the stomach twice weekly. Physiological saline alone was administered in a similar fashion to the control group. Blood samples were obtained prior to and after four and eight weeks of administration. White blood cells and lymphocyte counts were obtained and lymphocyte subsets were measured using monoclonal antibodies W3/13, W3/25 and 0 X 8 (Sera-Lab), and a laser flow cytometry system (Orthospectrum III, Orthodiagnostic System). The T cell ratio, helper/inducer T (Th) cell ratio, and suppressor/cytotoxic T (Ts) cell ratio were measured. The peripheral white blood cell count and lymphocyte count were not significantly different between the control and LTN groups. After four weeks of LTN administration, however, the LTN group showed a significantly higher T cell ratio, Th cell ratio and Th/Ts cell ratio than did the control group, and the Ts cell ratio was significantly lower. In the groups undergoing administration for eight weeks, no difference was noted in the lymphocyte subsets between the two groups. Oral administration of LTN apparently modulates the systemic immune function through T cell stimulation, especially Th cells, but continued administration may induce a tolerance to the effect of LTN.

Prevention of Autoimmune Diabetes and Induction of β-Cell Proliferation in NOD Mice by Hyperbaric Oxygen Therapy

PubMed Central

Faleo, Gaetano; Fotino, Carmen; Bocca, Nicola; Molano, R. Damaris; Zahr-Akrawi, Elsie; Molina, Judith; Villate, Susana; Umland, Oliver; Skyler, Jay S.; Bayer, Allison L.; Ricordi, Camillo; Pileggi, Antonello

2012-01-01

We evaluated the effects of hyperbaric oxygen therapy (HOT) on autoimmune diabetes development in nonobese diabetic (NOD) mice. Animals received no treatment or daily 60-min HOT 100% oxygen (HOT-100%) at 2.0 atmospheres absolute and were monitored for diabetes onset, insulitis, infiltrating cells, immune cell function, and β-cell apoptosis and proliferation. Cyclophosphamide-induced diabetes onset was reduced from 85.3% in controls to 48% after HOT-100% (P < 0.005) and paralleled by lower insulitis. Spontaneous diabetes incidence reduced from 85% in controls to 65% in HOT-100% (P = 0.01). Prediabetic mice receiving HOT-100% showed lower insulitis scores, reduced T-cell proliferation upon stimulation in vitro (P < 0.03), increased CD62L expression in T cells (P < 0.04), reduced costimulation markers (CD40, DC80, and CD86), and reduced major histocompatibility complex class II expression in dendritic cells (DCs) (P < 0.025), compared with controls. After autoimmunity was established, HOT was less effective. HOT-100% yielded reduced apoptosis (transferase-mediated dUTP nick-end labeling-positive insulin-positive cells; P < 0.01) and increased proliferation (bromodeoxyuridine incorporation; P < 0.001) of insulin-positive cells compared with controls. HOT reduces autoimmune diabetes incidence in NOD mice via increased resting T cells and reduced activation of DCs with preservation of β-cell mass resulting from decreased apoptosis and increased proliferation. The safety profile and noninvasiveness makes HOT an appealing adjuvant therapy for diabetes prevention and intervention trials. PMID:22566533

Smooth muscle-like tissue constructs with circumferentially oriented cells formed by the cell fiber technology.

PubMed

Hsiao, Amy Y; Okitsu, Teru; Onoe, Hiroaki; Kiyosawa, Mahiro; Teramae, Hiroki; Iwanaga, Shintaroh; Kazama, Tomohiko; Matsumoto, Taro; Takeuchi, Shoji

2015-01-01

The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments.

Smooth Muscle-Like Tissue Constructs with Circumferentially Oriented Cells Formed by the Cell Fiber Technology

PubMed Central

Hsiao, Amy Y.; Okitsu, Teru; Onoe, Hiroaki; Kiyosawa, Mahiro; Teramae, Hiroki; Iwanaga, Shintaroh; Kazama, Tomohiko; Matsumoto, Taro; Takeuchi, Shoji

2015-01-01

The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments. PMID:25734774

Open-access and multi-directional electroosmotic flow chip for positioning heterotypic cells.

PubMed

Terao, Kyohei; Kitazawa, Yuko; Yokokawa, Ryuji; Okonogi, Atsuhito; Kotera, Hidetoshi

2011-04-21

We propose a novel method of cell positioning using electroosmotic flow (EOF) to analyze cell-cell interactions. The EOF chip has an open-to-air configuration, is equipped with four electrodes to induce multi-directional EOF, and allows access of tools for liquid handling and of physical probes for cell measurements. Evaluation of the flow within this chip indicated that it controlled hydrodynamic transport of cells, in terms of both speed and direction. We also evaluated cell viability after EOF application and determined appropriate conditions for cell positioning. Two cells were successively positioned in pocket-like microstructures, one in each micropocket, by controlling the EOF direction. As an experimental demonstration, we observed contact interactions between two individual cells through gap junction channels. The EOF chip should provide ways to elucidate various cell-cell interactions between heterotypic cells.

CD4 T lymphocytes from patients with chronic fatigue syndrome have decreased interferon-gamma production and increased sensitivity to dexamethasone.

PubMed

Visser, J; Blauw, B; Hinloopen, B; Brommer, E; de Kloet, E R; Kluft, C; Nagelkerken, L

1998-02-01

A disturbed hypothalamus-pituitary-adrenal gland axis and alterations at the immune system level have been observed in patients with chronic fatigue syndrome (CFS). Glucocorticoids are known to modulate T cell responses; therefore, purified CD4 T cells from CFS patients were studied to determine whether they have an altered sensitivity to dexamethasone (DEX). CD4 T cells from CFS patients produced less interferon-gamma than did cells from controls; by contrast, interleukin-4 production and cell proliferation were comparable. With CD4 T cells from CFS patients (compared with cells from controls), a 10- to 20-fold lower DEX concentration was needed to achieve 50% inhibition of interleukin-4 production and proliferation, indicating an increased sensitivity to DEX in CFS patients. Surprisingly, interferon-gamma production in patients and controls was equally sensitive to DEX. A differential sensitivity of cytokines or CD4 T cell subsets to glucocorticoids might explain an altered immunologic function in CFS patients.

Independent controls for neocortical neuron production and histogenetic cell death

NASA Technical Reports Server (NTRS)

Verney, C.; Takahashi, T.; Bhide, P. G.; Nowakowski, R. S.; Caviness, V. S. Jr

2000-01-01

We estimated the proportion of cells eliminated by histogenetic cell death during the first 2 postnatal weeks in areas 1, 3 and 40 of the mouse parietal neocortex. For each layer and for the subcortical white matter in each neocortical area, the number of dying cells per mm(2) was calculated and the proportionate cell death for each day of the 2-week interval was estimated. The data show that cell death proceeds essentially uniformly across the neocortical areas and layers and that it does not follow either the spatiotemporal gradient of cell cycle progression in the pseudostratified ventricular epithelium of the cerebral wall, the source of neocortical neurons, or the 'inside-out' neocortical neuronogenetic sequence. Therefore, we infer that the control mechanisms of neocortical histogenetic cell death are independent of mechanisms controlling neuronogenesis or neuronal migration but may be associated with the ingrowth, expansion and a system-wide matching of neuronal connectivity. Copyright 2000 S. Karger AG, Basel.

Elevated IL-8 levels during sickle cell crisis.

PubMed

Duits, A J; Schnog, J B; Lard, L R; Saleh, A W; Rojer, R A

1998-11-01

The vaso-occlusive process (VOC) in sickle cell disease is of a complex nature. It involves intricate interactions between sickle red blood cells, endothelium and probably also leukocytes. As these interactions are regulated by cytokines, we analyzed the role of the potent neutrophil chemokine IL-8 by measuring serum levels in sickle cell patients during sickle cell crisis. These results were compared to nonsymptomatics and healthy controls. In patients having a vaso-occlusive crisis both HbSS and HbSC patients showed significantly enhanced serum IL-8 levels compared to healthy controls. Several of these patients showed extremely elevated serum IL-8 levels which were independent of the crisis inducing factor. Furthermore, a sickle cell patient with VOC as a complication of rhGM-CSF treatment similarly showed high IL-8 serum levels at crisis onset. Nonsymptomatic sickle cell patients serum IL-8 levels were comparable to healthy controls. These results implicate a role for IL-8 at or during (the initiation of) sickle cell crisis.

Submergible barge retrievable storage and permanent disposal system for radioactive waste

DOEpatents

Goldsberry, Fred L.; Cawley, William E.

1981-01-01

A submergible barge and process for submerging and storing radioactive waste material along a seabed. A submergible barge receives individual packages of radwaste within segregated cells. The cells are formed integrally within the barge, preferably surrounded by reinforced concrete. The cells are individually sealed by a concrete decking and by concrete hatch covers. Seawater may be vented into the cells for cooling, through an integral vent arrangement. The vent ducts may be attached to pumps when the barge is bouyant. The ducts are also arranged to promote passive ventilation of the cells when the barge is submerged. Packages of the radwaste are loaded into individual cells within the barge. The cells are then sealed and the barge is towed to the designated disposal-storage site. There, the individual cells are flooded and the barge will begin descent controlled by a powered submarine control device to the seabed storage site. The submerged barge will rest on the seabed permanently or until recovered by a submarine control device.

Subcellular and supracellular mechanical stress prescribes cytoskeleton behavior in Arabidopsis cotyledon pavement cells

PubMed Central

Sampathkumar, Arun; Krupinski, Pawel; Wightman, Raymond; Milani, Pascale; Berquand, Alexandre; Boudaoud, Arezki; Hamant, Olivier; Jönsson, Henrik; Meyerowitz, Elliot M

2014-01-01

Although it is a central question in biology, how cell shape controls intracellular dynamics largely remains an open question. Here, we show that the shape of Arabidopsis pavement cells creates a stress pattern that controls microtubule orientation, which then guides cell wall reinforcement. Live-imaging, combined with modeling of cell mechanics, shows that microtubules align along the maximal tensile stress direction within the cells, and atomic force microscopy demonstrates that this leads to reinforcement of the cell wall parallel to the microtubules. This feedback loop is regulated: cell-shape derived stresses could be overridden by imposed tissue level stresses, showing how competition between subcellular and supracellular cues control microtubule behavior. Furthermore, at the microtubule level, we identified an amplification mechanism in which mechanical stress promotes the microtubule response to stress by increasing severing activity. These multiscale feedbacks likely contribute to the robustness of microtubule behavior in plant epidermis. DOI: http://dx.doi.org/10.7554/eLife.01967.001 PMID:24740969

Division of labour between Myc and G1 cyclins in cell cycle commitment and pace control.

PubMed

Dong, Peng; Maddali, Manoj V; Srimani, Jaydeep K; Thélot, François; Nevins, Joseph R; Mathey-Prevot, Bernard; You, Lingchong

2014-09-01

A body of evidence has shown that the control of E2F transcription factor activity is critical for determining cell cycle entry and cell proliferation. However, an understanding of the precise determinants of this control, including the role of other cell-cycle regulatory activities, has not been clearly defined. Here, recognizing that the contributions of individual regulatory components could be masked by heterogeneity in populations of cells, we model the potential roles of individual components together with the use of an integrated system to follow E2F dynamics at the single-cell level and in real time. These analyses reveal that crossing a threshold amplitude of E2F accumulation determines cell cycle commitment. Importantly, we find that Myc is critical in modulating the amplitude, whereas cyclin D/E activities have little effect on amplitude but do contribute to the modulation of duration of E2F activation, thereby affecting the pace of cell cycle progression.

A new insight in chimeric antigen receptor-engineered T cells for cancer immunotherapy.

PubMed

Zhang, Erhao; Xu, Hanmei

2017-01-03

Adoptive cell therapy using chimeric antigen receptor (CAR)-engineered T cells has emerged as a very promising approach to combating cancer. Despite its ability to eliminate tumors shown in some clinical trials, CAR-T cell therapy involves some significant safety challenges, such as cytokine release syndrome (CRS) and "on-target, off-tumor" toxicity, which is related to poor control of the dose, location, and timing of T cell activity. In the past few years, some strategies to avoid the side effects of CAR-T cell therapy have been reported, including suicide gene, inhibitory CAR, dual-antigen receptor, and the use of exogenous molecules as switches to control the CAR-T cell functions. Because of the advances of the CAR paradigm and other forms of cancer immunotherapy, the most effective means of defeating the cancer has become the integration therapy with the combinatorial control system of switchable dual-receptor CAR-T cell and immune checkpoint blockade.

Behavior-dependent specialization of identified hippocampal interneurons

PubMed Central

Lapray, Damien; Lasztoczi, Balint; Lagler, Michael; Viney, Tim James; Katona, Linda; Valenti, Ornella; Hartwich, Katja; Borhegyi, Zsolt; Somogyi, Peter; Klausberger, Thomas

2012-01-01

A large variety of GABAergic interneurons control information processing in hippocampal circuits governing the formation of neuronal representations. Whether distinct hippocampal interneuron types contribute differentially to information-processing during behavior is not known. We employed a novel technique for recording and labeling interneurons and pyramidal cells in drug-free, freely-moving rats. Recorded parvalbumin-expressing basket interneurons innervate somata and proximal pyramidal cell dendrites, whereas nitric-oxide-synthase- and neuropeptide-Y-expressing ivy cells provide synaptic and extrasynaptic dendritic modulation. Basket and ivy cells showed distinct spike timing dynamics, firing at different rates and times during theta and ripple oscillations. Basket but not ivy cells changed their firing rates during movement, sleep and quiet wakefulness, suggesting that basket cells coordinate cell assemblies in a behavioral state-contingent manner, whereas persistently-firing ivy cells might control network excitability and homeostasis. Different interneuron types provide GABA to specific subcellular domains at defined times and rates, thus differentially controlling network activity during behavior. PMID:22864613

The C. elegans engrailed homolog ceh-16 regulates the self-renewal expansion division of stem cell-like seam cells.

PubMed

Huang, Xinxin; Tian, E; Xu, Yanhua; Zhang, Hong

2009-09-15

Stem cells undergo symmetric and asymmetric division to maintain the dynamic equilibrium of the stem cell pool and also to generate a variety of differentiated cells. The homeostatic mechanism controlling the choice between self-renewal and differentiation of stem cells is poorly understood. We show here that ceh-16, encoding the C. elegans ortholog of the transcription factor Engrailed, controls symmetric and asymmetric division of stem cell-like seam cells. Loss of function of ceh-16 causes certain seam cells, which normally undergo symmetric self-renewal expansion division with both daughters adopting the seam cell fate, to divide asymmetrically with only one daughter retaining the seam cell fate. The human engrailed homolog En2 functionally substitutes the role of ceh-16 in promoting self-renewal expansion division of seam cells. Loss of function of apr-1, encoding the C. elegans homolog of the Wnt signaling component APC, results in transformation of self-renewal maintenance seam cell division to self-renewal expansion division, leading to seam cell hyperplasia. The apr-1 mutation suppresses the seam cell division defect in ceh-16 mutants. Our study reveals that ceh-16 interacts with the Wnt signaling pathway to control the choice between self-renewal expansion and maintenance division and also demonstrates an evolutionarily conserved function of engrailed in promoting cell proliferation.

Neonatal rat heart cells cultured in simulated microgravity

NASA Technical Reports Server (NTRS)

Akins, Robert E.; Schroedl, Nancy A.; Gonda, Steve R.; Hartzell, Charles R.

1994-01-01

In vitro characteristics of cardiac cells cultured in simulated microgravity are reported. Tissue culture methods performed at unit gravity constrain cells to propagate, differentiate, and interact in a two dimensional (2D) plane. Neonatal rat cardiac cells in 2D culture organize predominantly as bundles of cardiomyocytes with the intervening areas filled by non-myocyte cell types. Such cardiac cell cultures respond predictably to the addition of exogenous compounds, and in many ways they represent an excellent in vitro model system. The gravity-induced 2D organization of the cells, however, does not accurately reflect the distribution of cells in the intact tissue. We have begun characterizations of a three-dimensional (3D) culturing system designed to mimic microgravity. The NASA designed High-Aspect-Ratio-Vessel (HARV) bioreactors provide a low shear environment which allows cells to be cultured in static suspension. HARV-3D cultures were prepared on microcarrier beads and compared to control-2D cultures using a combination of microscopic and biochemical techniques. Both systems were uniformly inoculated and medium exchanged at standard intervals. Cells in control cultures adhered to the polystyrene surface of the tissue culture dishes and exhibited typical 2D organization. Cells in cultured in HARV's adhered to microcarrier beads, the beads aggregated into defined clusters containing 8 to 15 beads per cluster, and the clusters exhibited distinct 3D layers: myocytes and fibroblasts appeared attached to the surfaces of beads and were overlaid by an outer cell type. In addition, cultures prepared in HARV's using alternative support matrices also displayed morphological formations not seen in control cultures. Generally, the cells prepared in HARV and control cultures were similar, however, the dramatic alterations in 3D organization recommend the HARV as an ideal vessel for the generation of tissue-like organizations of cardiac cells in simulated microgravity.

Innate (inherent) control of brain infection, brain inflammation and brain repair: the role of microglia, astrocytes, "protective" glial stem cells and stromal ependymal cells.

PubMed

Hauwel, Mathieu; Furon, Emeline; Canova, Cecile; Griffiths, Mark; Neal, Jim; Gasque, Philippe

2005-04-01

In invertebrates and primitive vertebrates, the brain contains large numbers of "professional" macrophages associated with neurones, ependymal tanycytes and radial glia to promote robust regenerative capacity. In higher vertebrates, hematogenous cells are largely excluded from the brain, and innate immune molecules and receptors produced by the resident "amateur" macrophages (microglia, astrocytes and ependymal cells) control pathogen infiltration and clearance of toxic cell debris. However, there is minimal capacity for regeneration. The transfer of function from hematogenous cells to macroglia and microglia is associated with the sophistication of a yet poorly-characterized neurone-glia network. This evolutionary pattern may have been necessary to reduce the risk of autoimmune attack while preserving the neuronal web but the ability to repair central nervous system damage may have been sacrificed in the process. We herein argue that it may be possible to re-educate and stimulate the resident phagocytes to promote clearance of pathogens (e.g., Prion), toxic cell debris (e.g., amyloid fibrils and myelin) and apoptotic cells. Moreover, as part of this greater division of labour between cell types in vertebrate brains, it may be possible to harness the newly described properties of glial stem cells in neuronal protection (revitalization) rather than replacement, and to control brain inflammation. We will also highlight the emerging roles of stromal ependymal cells in controlling stem cell production and migration into areas of brain damage. Understanding the mechanisms involved in the nurturing of damaged neurons by protective glial stem cells with the safe clearance of cell debris could lead to remedial strategies for chronic brain diseases.

Neonatal rat heart cells cultured in simulated microgravity

NASA Technical Reports Server (NTRS)

Akins, R. E.; Schroedl, N. A.; Gonda, S. R.; Hartzell, C. R.

1997-01-01

In vitro characteristics of cardiac cells cultured in simulated microgravity are reported. Tissue culture methods performed at unit gravity constrain cells to propagate, differentiate, and interact in a two-dimensional (2D) plane. Neonatal rat cardiac cells in 2D culture organize predominantly as bundles of cardiomyocytes with the intervening areas filled by nonmyocyte cell types. Such cardiac cell cultures respond predictably to the addition of exogenous compounds, and in many ways they represent an excellent in vitro model system. The gravity-induced 2D organization of the cells, however, does not accurately reflect the distribution of cells in the intact tissue. We have begun characterizations of a three-dimensional (3D) culturing system designed to mimic microgravity. The NASA-designed High-Aspect Ratio Vessel (HARV) bioreactors provide a low shear environment that allows cells to be cultured in static suspension. HARV-3D cultures were prepared on microcarrier beads and compared to control-2D cultures using a combination of microscopic and biochemical techniques. Both systems were uniformly inoculated and medium exchanged at standard intervals. Cells in control cultures adhered to the polystyrene surface of the tissue culture dishes and exhibited typical 2D organization. Cells cultured in HARVs adhered to microcarrier beads, the beads aggregated into defined clusters containing 8 to 15 beads per cluster, and the clusters exhibited distinct 3D layers: myocytes and fibroblasts appeared attached to the surfaces of beads and were overlaid by an outer cell type. In addition, cultures prepared in HARVs using alternative support matrices also displayed morphological formations not seen in control cultures. Generally, the cells prepared in HARV and control cultures were similar; however, the dramatic alterations in 3D organization recommend the HARV as an ideal vessel for the generation of tissuelike organization of cardiac cells in vitro.

Amyloid precursor protein regulates migration and metalloproteinase gene expression in prostate cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyazaki, Toshiaki; Ikeda, Kazuhiro; Horie-Inoue, Kuniko

Highlights: • APP knockdown reduced proliferation and migration of prostate cancer cells. • APP knockdown reduced expression of metalloproteinase and EMT-related genes. • APP overexpression promoted LNCaP cell migration. • APP overexpression increased expression of metalloproteinase and EMT-related genes. - Abstract: Amyloid precursor protein (APP) is a type I transmembrane protein, and one of its processed forms, β-amyloid, is considered to play a central role in the development of Alzheimer’s disease. We previously showed that APP is a primary androgen-responsive gene in prostate cancer and that its increased expression is correlated with poor prognosis for patients with prostate cancer. APPmore » has also been implicated in several human malignancies. Nevertheless, the mechanism underlying the pro-proliferative effects of APP on cancers is still not well-understood. In the present study, we explored a pathophysiological role for APP in prostate cancer cells using siRNA targeting APP (siAPP). The proliferation and migration of LNCaP and DU145 prostate cancer cells were significantly suppressed by siAPP. Differentially expressed genes in siAPP-treated cells compared to control siRNA-treated cells were identified by microarray analysis. Notably, several metalloproteinase genes, such as ADAM10 and ADAM17, and epithelial–mesenchymal transition (EMT)-related genes, such as VIM, and SNAI2, were downregulated in siAPP-treated cells as compared to control cells. The expression of these genes was upregulated in LNCaP cells stably expressing APP when compared with control cells. APP-overexpressing LNCaP cells exhibited enhanced migration in comparison to control cells. These results suggest that APP may contribute to the proliferation and migration of prostate cancer cells by modulating the expression of metalloproteinase and EMT-related genes.« less

Proof-of-Principle for Immune Control of Global HIV-1 Reactivation In Vivo

PubMed Central

Smith, Nicola M. G.; Mlcochova, Petra; Watters, Sarah A.; Aasa-Chapman, Marlene M. I.; Rabin, Neil; Moore, Sally; Edwards, Simon G.; Garson, Jeremy A.; Grant, Paul R.; Ferns, R. Bridget; Kashuba, Angela; Mayor, Neema P.; Schellekens, Jennifer; Marsh, Steven G. E.; McMichael, Andrew J.; Perelson, Alan S.; Pillay, Deenan; Goonetilleke, Nilu; Gupta, Ravindra K.

2015-01-01

Background. Emerging data relating to human immunodeficiency virus type 1 (HIV-1) cure suggest that vaccination to stimulate the host immune response, particularly cytotoxic cells, may be critical to clearing of reactivated HIV-1–infected cells. However, evidence for this approach in humans is lacking, and parameters required for a vaccine are unknown because opportunities to study HIV-1 reactivation are rare. Methods. We present observations from a HIV-1 elite controller, not treated with combination antiretroviral therapy, who experienced viral reactivation following treatment for myeloma with melphalan and autologous stem cell transplantation. Mathematical modeling was performed using a standard viral dynamic model. Enzyme-linked immunospot, intracellular cytokine staining, and tetramer staining were performed on peripheral blood mononuclear cells; in vitro CD8 T-cell–mediated control of virion production by autologous CD4 T cells was quantified; and neutralizing antibody titers were measured. Results. Viral rebound was measured at 28 000 copies/mL on day 13 post-transplant before rapid decay to <50 copies/mL in 2 distinct phases with t1/2 of 0.71 days and 4.1 days. These kinetics were consistent with an expansion of cytotoxic effector cells and killing of productively infected CD4 T cells. Following transplantation, innate immune cells, including natural killer cells, recovered with virus rebound. However, most striking was the expansion of highly functional HIV-1–specific cytotoxic CD8 T cells, at numbers consistent with those applied in modeling, as virus control was regained. Conclusions. These observations provide evidence that the human immune response is capable of controlling coordinated global HIV-1 reactivation, remarkably with potency equivalent to combination antiretroviral therapy. These data will inform design of vaccines for use in HIV-1 curative interventions. PMID:25778749

Feasibility of controlling CD38-CAR T cell activity with a Tet-on inducible CAR design

PubMed Central

Poels, Renée; Mulders, Manon J.; van de Donk, Niels W. C. J.; Themeli, Maria; Lokhorst, Henk M.; Mutis, Tuna

2018-01-01

Recent clinical advances with chimeric antigen receptor (CAR) T cells have led to the accelerated clinical approval of CD19-CARs to treat acute lymphoblastic leukemia. The CAR T cell therapy is nevertheless associated with toxicities, especially if the CARs are not entirely tumor-specific. Therefore, strategies for controlling the CAR T cell activity are required to improve their safety profile. Here, by using the multiple myeloma (MM)-associated CD38 molecule as target molecule, we tested the feasibility and utility of a doxycycline (DOX) inducible Tet-on CD38-CAR design to control the off-target toxicities of CAR T cells. Using CARs with high affinity to CD38, we demonstrate that this strategy allows the proper induction of CD38-CARs and CAR-mediated T cell cytotoxicity in a DOX-dose dependent manner. Especially when the DOX dose was limited to 10ng/ml, its removal resulted in a relatively rapid decay of CAR- related off-tumor effects within 24 hours, indicating the active controllability of undesired CAR activity. This Tet-on CAR design also allowed us to induce the maximal anti-MM cytotoxic activity of affinity-optimized CD38-CAR T cells, which already display a low toxicity profile, hereby adding a second level of safety to these cells. Collectively, these results indicate the possibility to utilize this DOX inducible CAR-design to actively regulate the CAR-mediated activities of therapeutic T cells. We therefore conclude that the Tet-on system may be more advantageous above suicide-genes to control the potential toxicities of CAR T cells without the need to destroy them permanently. PMID:29847570

Preprocessing with Photoshop Software on Microscopic Images of A549 Cells in Epithelial-Mesenchymal Transition.

PubMed

Ren, Zhou-Xin; Yu, Hai-Bin; Shen, Jun-Ling; Li, Ya; Li, Jian-Sheng

2015-06-01

To establish a preprocessing method for cell morphometry in microscopic images of A549 cells in epithelial-mesenchymal transition (EMT). Adobe Photoshop CS2 (Adobe Systems, Inc.) was used for preprocessing the images. First, all images were processed for size uniformity and high distinguishability between the cell and background area. Then, a blank image with the same size and grids was established and cross points of the grids were added into a distinct color. The blank image was merged into a processed image. In the merged images, the cells with 1 or more cross points were chosen, and then the cell areas were enclosed and were replaced in a distinct color. Except for chosen cellular areas, all areas were changed into a unique hue. Three observers quantified roundness of cells in images with the image preprocess (IPP) or without the method (Controls), respectively. Furthermore, 1 observer measured the roundness 3 times with the 2 methods, respectively. The results between IPPs and Controls were compared for repeatability and reproducibility. As compared with the Control method, among 3 observers, use of the IPP method resulted in a higher number and a higher percentage of same-chosen cells in an image. The relative average deviation values of roundness, either for 3 observers or 1 observer, were significantly higher in Controls than in IPPs (p < 0.01 or 0.001). The values of intraclass correlation coefficient, both in Single Type or Average, were higher in IPPs than in Controls both for 3 observers and 1 observer. Processed with Adobe Photoshop, a chosen cell from an image was more objective, regular, and accurate, creating an increase of reproducibility and repeatability on morphometry of A549 cells in epithelial to mesenchymal transition.

Central importance of immunoglobulin A in host defense against Giardia spp.

PubMed

Langford, T Dianne; Housley, Michael P; Boes, Marianne; Chen, Jianzhu; Kagnoff, Martin F; Gillin, Frances D; Eckmann, Lars

2002-01-01

The protozoan pathogen Giardia is an important cause of parasitic diarrheal disease worldwide. It colonizes the lumen of the small intestine, suggesting that effective host defenses must act luminally. Immunoglobulin A (IgA) antibodies are presumed to be important for controlling Giardia infection, but direct evidence for this function is lacking. B-cell-independent effector mechanisms also exist and may be equally important for antigiardial host defense. To determine the importance of the immunoglobulin isotypes that are transported into the intestinal lumen, IgA and IgM, for antigiardial host defense, we infected gene-targeted mice lacking IgA-expressing B-cells, IgM-secreting B-cells, or all B-cells as controls with Giardia muris or Giardia lamblia GS/M-83-H7. We found that IgA-deficient mice could not eradicate either G. muris or G. lamblia infection, demonstrating that IgA is required for their clearance. Furthermore, although neither B-cell-deficient nor IgA-deficient mice could clear G. muris infections, IgA-deficient mice controlled infection significantly better than B-cell-deficient mice, suggesting the existence of B-cell-dependent but IgA-independent antigiardial defenses. In contrast, mice deficient for secreted IgM antibodies cleared G. muris infection normally, indicating that they have no unique functions in antigiardial host defense. These data, together with the finding that B-cell-deficient mice have some, albeit limited, residual capacity to control G. muris infection, show that IgA-dependent host defenses are central for eradicating Giardia spp. Moreover, B-cell-dependent but IgA-independent and B-cell-independent antigiardial host defenses exist but are less important for controlling infection.

PGE2 /EP4 Signaling Controls the Transfer of the Mammary Stem Cell State by Lipid Rafts in Extracellular Vesicles.

PubMed

Lin, Meng-Chieh; Chen, Shih-Yin; Tsai, Ho-Min; He, Pei-Lin; Lin, Yen-Chun; Herschman, Harvey; Li, Hua-Jung

2017-02-01

Prostaglandin E 2 (PGE 2 )-initiated signaling contributes to stem cell homeostasis and regeneration. However, it is unclear how PGE 2 signaling controls cell stemness. This study identifies a previously unknown mechanism by which PGE 2 /prostaglandin E receptor 4 (EP 4 ) signaling regulates multiple signaling pathways (e.g., PI3K/Akt signaling, TGFβ signaling, Wnt signaling, EGFR signaling) which maintain the basal mammary stem cell phenotype. A shift of basal mammary epithelial stem cells (MaSCs) from a mesenchymal/stem cell state to a non-basal-MaSC state occurs in response to prostaglandin E receptor 4 (EP 4 ) antagonism. EP 4 antagonists elicit release of signaling components, by controlling their trafficking into extracellular vesicles/exosomes in a lipid raft/caveolae-dependent manner. Consequently, EP 4 antagonism indirectly inactivates, through induced extracellular vesicle/exosome release, pathways required for mammary epithelial stem cell homeostasis, e.g. canonical/noncanonical Wnt, TGFβ and PI3K/Akt pathways. EP 4 antagonism causes signaling receptors and signaling components to shift from non-lipid raft fractions to lipid raft fractions, and to then be released in EP 4 antagonist-induced extracellular vesicles/exosomes, resulting in the loss of the stem cell state by mammary epithelial stem cells. In contrast, luminal mammary epithelial cells can acquire basal stem cell properties following ingestion of EP 4 antagonist-induced stem cell extracellular vesicles/exosomes, and can then form mammary glands. These findings demonstrate that PGE 2 /EP 4 signaling controls homeostasis of mammary epithelial stem cells through regulating extracellular vesicle/exosome release. Reprogramming of mammary epithelial cells can result from EP 4 -mediated stem cell property transfer by extracellular vesicles/exosomes containing caveolae-associated proteins, between mammary basal and luminal epithelial cells. Stem Cells 2017;35:425-444. © 2016 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Human CD34+ Progenitor Cells Freshly Isolated from Umbilical Cord Blood Attenuate Inflammatory Lung Injury following LPS Challenge

PubMed Central

Huang, Xiaojia; Sun, Kai; Zhao, Yidan D.; Vogel, Stephen M.; Song, Yuanling; Mahmud, Nadim; Zhao, You-Yang

2014-01-01

Adult stem cell-based therapy is a promising novel approach for treatment of acute lung injury. Here we investigated the therapeutic potential of freshly isolated human umbilical cord blood CD34+ progenitor cells (fCB-CD34+ cells) in a mouse model of acute lung injury. At 3 h post-lipopolysaccharide (LPS) challenge, fCB-CD34+ cells were transplanted i.v. to mice while CD34− cells or PBS were administered as controls in separate cohorts of mice. We observed that fCB-CD34+ cell treatment inhibited lung vascular injury evident by decreased lung vascular permeability. In contrast, CD34− cells had no effects on lung vascular injury. Lung inflammation determined by myeloperoxidase activity, neutrophil sequestration and expression of pro-inflammatory mediators was attenuated in fCB-CD34+ cell-treated mice at 26 h post-LPS challenge compared to PBS or CD34− cell-treated controls. Importantly, lung inflammation in fCB-CD34+ cell-treated mice was returned to normal levels as seen in basal mice at 52 h post-LPS challenge whereas PBS or CD34− cell-treated control mice exhibited persistent lung inflammation. Accordingly, fCB-CD34+ cell-treated mice exhibited a marked increase of survival rate. Employing in vivo 5-bromo-2′-deoxyuridine incorporation assay, we found a drastic induction of lung endothelial proliferation in fCB-CD34+ cell-treated mice at 52 h post-LPS compared to PBS or CD34− cell-treated controls, which contributed to restoration of vascular integrity and thereby inhibition of lung inflammation. Taken together, these data have demonstrated the protective effects of fCB-CD34+ cell on acute lung injury induced by LPS challenge, suggesting fCB-CD34+ cells are an important source of stem cells for the treatment of acute lung injury. PMID:24558433

Comprehensive analysis of genome-wide DNA methylation across human polycystic ovary syndrome ovary granulosa cell.

PubMed

Xu, Jiawei; Bao, Xiao; Peng, Zhaofeng; Wang, Linlin; Du, Linqing; Niu, Wenbin; Sun, Yingpu

2016-05-10

Polycystic ovary syndrome (PCOS) affects approximately 7% of the reproductive-age women. A growing body of evidence indicated that epigenetic mechanisms contributed to the development of PCOS. The role of DNA modification in human PCOS ovary granulosa cell is still unknown in PCOS progression. Global DNA methylation and hydroxymethylation were detected between PCOS' and controls' granulosa cell. Genome-wide DNA methylation was profiled to investigate the putative function of DNA methylaiton. Selected genes expressions were analyzed between PCOS' and controls' granulosa cell. Our results showed that the granulosa cell global DNA methylation of PCOS patients was significant higher than the controls'. The global DNA hydroxymethylation showed low level and no statistical difference between PCOS and control. 6936 differentially methylated CpG sites were identified between control and PCOS-obesity. 12245 differential methylated CpG sites were detected between control and PCOS-nonobesity group. 5202 methylated CpG sites were significantly differential between PCOS-obesity and PCOS-nonobesity group. Our results showed that DNA methylation not hydroxymethylation altered genome-wide in PCOS granulosa cell. The different methylation genes were enriched in development protein, transcription factor activity, alternative splicing, sequence-specific DNA binding and embryonic morphogenesis. YWHAQ, NCF2, DHRS9 and SCNA were up-regulation in PCOS-obesity patients with no significance different between control and PCOS-nonobesity patients, which may be activated by lower DNA methylaiton. Global and genome-wide DNA methylation alteration may contribute to different genes expression and PCOS clinical pathology.

A new epidemic modeling approach: Multi-regions discrete-time model with travel-blocking vicinity optimal control strategy.

PubMed

Zakary, Omar; Rachik, Mostafa; Elmouki, Ilias

2017-08-01

First, we devise in this paper, a multi-regions discrete-time model which describes the spatial-temporal spread of an epidemic which starts from one region and enters to regions which are connected with their neighbors by any kind of anthropological movement. We suppose homogeneous Susceptible-Infected-Removed (SIR) populations, and we consider in our simulations, a grid of colored cells, which represents the whole domain affected by the epidemic while each cell can represent a sub-domain or region. Second, in order to minimize the number of infected individuals in one region, we propose an optimal control approach based on a travel-blocking vicinity strategy which aims to control only one cell by restricting movements of infected people coming from all neighboring cells. Thus, we show the influence of the optimal control approach on the controlled cell. We should also note that the cellular modeling approach we propose here, can also describes infection dynamics of regions which are not necessarily attached one to an other, even if no empty space can be viewed between cells. The theoretical method we follow for the characterization of the travel-locking optimal controls, is based on a discrete version of Pontryagin's maximum principle while the numerical approach applied to the multi-points boundary value problems we obtain here, is based on discrete progressive-regressive iterative schemes. We illustrate our modeling and control approaches by giving an example of 100 regions.

Primary Sjögren's syndrome is characterized by distinct phenotypic and transcriptional profiles of IgD+ unswitched memory B cells.

PubMed

Roberts, Mustimbo E P; Kaminski, Denise; Jenks, Scott A; Maguire, Craig; Ching, Kathryn; Burbelo, Peter D; Iadarola, Michael J; Rosenberg, Alexander; Coca, Andreea; Anolik, Jennifer; Sanz, Iñaki

2014-09-01

The significance of distinct B cell abnormalities in primary Sjögren's syndrome (SS) remains to be established. We undertook this study to analyze the phenotype and messenger RNA (mRNA) transcript profiles of B cell subsets in patients with primary SS and to compare them with those in sicca syndrome patients and healthy controls. CD19+ B cells from 26 patients with primary SS, 27 sicca syndrome patients, and 22 healthy controls were analyzed by flow cytometry. Gene expression profiles of purified B cell subsets (from 3-5 subjects per group per test) were analyzed using Affymetrix gene arrays. Patients with primary SS had lower frequencies of CD27+IgD- switched memory B cells and CD27+IgD+ unswitched memory B cells compared with healthy controls. Unswitched memory B cell frequencies were also lower in sicca syndrome patients and correlated inversely with serologic hyperactivity in both disease states. Further, unswitched memory B cells in primary SS had lower expression of CD1c and CD21. Gene expression analysis of CD27+ memory B cells separated patients with primary SS from healthy controls and identified a subgroup of sicca syndrome patients with a primary SS-like transcript profile. Moreover, unswitched memory B cell gene expression analysis identified 187 genes differentially expressed between patients with primary SS and healthy controls. A decrease in unswitched memory B cells with serologic hyperactivity is characteristic of both established primary SS and a subgroup of sicca syndrome, which suggests the value of these B cells both as biomarkers of future disease progression and for understanding disease pathogenesis. Overall, the mRNA transcript analysis of unswitched memory B cells suggests that their activation in primary SS takes place through innate immune pathways in the context of attenuated antigen-mediated adaptive signaling. Thus, our findings provide important insight into the mechanisms and potential consequences of decreased unswitched memory B cells in primary SS. Copyright © 2014 by the American College of Rheumatology.

[Effect of CP Metronomic Chemotherapy on RPMI 8226 Cell Proli-feration and Notch1/NF-κB Signaling Pathway In Vitro].

PubMed

Guo, Lie-Ping; Zhou, Fan; Shi, Hao-Tian; Chen, Hai-Min; Lin, Chen-Hui; Chen, Xiao-Ling; Hou, Jian

2016-10-01

To investigate the effect of metronomic chemotherapy of low dose phosphoramide combined with prednisolone (CP metronomic chemotherapy) on proliferation and apoptosis of RPMI 8226 cells, and to explore its regulating effect on Notch1/NF-κB signaling pathways. Experiment was divided into the DMSO control group, and the phosphoramide mustard (PM) group, the prednisolone group, the phosphoramide mustard plus prednisolone group (the CP group). RPMI 8226 cells were treated with different drugs, CCK-8 method was used to detect cell proliferation, flow cytometry was used to detect the cell cycle and apoptosis, reverse transcription PCR was used to detect Notch1 and NF-κB mRNA expression level. Compared with DMSO control group, RPMI8226 cell proliferation inhibition rate in all the PM, prednisolone and CP groups increased significantly with prolonging of time (r of 0.994,0.996,0.999, respectively, P<0.001). And at the same time, the inhibitory rate of cell proliferation was significantly different; the cell inhibitory rate in PM group was lowest, that in CP group was highgest, that in prednissone group was intermediate (P<0.01). After 48 hours, compared with the DMSO control group, the G 1 /G 0 cell proportion in treatment group increased significantly, S phase cell proportion decreased significantly, especially in PM and CP groups. The G 2 /M phase cell proportion increased in PM group, while reduced in the prednisolone and the CP groups. After 48 hours, compared with the DMSO control group, RPMI 8226 cell apoptosis rate increased as follow: in PM, pre-dnisolone and CP group(P<0.01). After 48 hours, compared with the DMSO control group, Notch1 and NF-κB mRNA expression in the prednisolone, the PM and the CP group decreased significantly(P<0.001). CP metronomic chemotherapy can significantly reduce RPMI 8226 cell proliferation, promote RPMI 8226 cell apoptosis, arrest RPMI 8226 cells mainly in the G 1 /G 0 phase, and significantly reduce Notch1 and NF-κB expression levels. It is suggested that Notch1/NF-κB signaling pathways is involved in CP metronomic chemotherapy for MM.

HIV Controllers Exhibit Enhanced Frequencies of Major Histocompatibility Complex Class II Tetramer+ Gag-Specific CD4+ T Cells in Chronic Clade C HIV-1 Infection.

PubMed

Laher, Faatima; Ranasinghe, Srinika; Porichis, Filippos; Mewalal, Nikoshia; Pretorius, Karyn; Ismail, Nasreen; Buus, Søren; Stryhn, Anette; Carrington, Mary; Walker, Bruce D; Ndung'u, Thumbi; Ndhlovu, Zaza M

2017-04-01

Immune control of viral infections is heavily dependent on helper CD4 + T cell function. However, the understanding of the contribution of HIV-specific CD4 + T cell responses to immune protection against HIV-1, particularly in clade C infection, remains incomplete. Recently, major histocompatibility complex (MHC) class II tetramers have emerged as a powerful tool for interrogating antigen-specific CD4 + T cells without relying on effector functions. Here, we defined the MHC class II alleles for immunodominant Gag CD4 + T cell epitopes in clade C virus infection, constructed MHC class II tetramers, and then used these to define the magnitude, function, and relation to the viral load of HIV-specific CD4 + T cell responses in a cohort of untreated HIV clade C-infected persons. We observed significantly higher frequencies of MHC class II tetramer-positive CD4 + T cells in HIV controllers than progressors ( P = 0.0001), and these expanded Gag-specific CD4 + T cells in HIV controllers showed higher levels of expression of the cytolytic proteins granzymes A and B. Importantly, targeting of the immunodominant Gag41 peptide in the context of HLA class II DRB1*1101 was associated with HIV control ( r = -0.5, P = 0.02). These data identify an association between HIV-specific CD4 + T cell targeting of immunodominant Gag epitopes and immune control, particularly the contribution of a single class II MHC-peptide complex to the immune response against HIV-1 infection. Furthermore, these results highlight the advantage of the use of class II tetramers in evaluating HIV-specific CD4 + T cell responses in natural infections. IMPORTANCE Increasing evidence suggests that virus-specific CD4 + T cells contribute to the immune-mediated control of clade B HIV-1 infection, yet there remains a relative paucity of data regarding the role of HIV-specific CD4 + T cells in shaping adaptive immune responses in individuals infected with clade C, which is responsible for the majority of HIV infections worldwide. Understanding the contribution of HIV-specific CD4 + T cell responses in clade C infection is particularly important for developing vaccines that would be efficacious in sub-Saharan Africa, where clade C infection is dominant. Here, we employed MHC class II tetramers designed to immunodominant Gag epitopes and used them to characterize CD4 + T cell responses in HIV-1 clade C infection. Our results demonstrate an association between the frequency of HIV-specific CD4 + T cell responses targeting an immunodominant DRB1*11-Gag41 complex and HIV control, highlighting the important contribution of a single class II MHC-peptide complex to the immune response against HIV-1 infections. Copyright © 2017 American Society for Microbiology.

The cell cycle and acute kidney injury

PubMed Central

Price, Peter M.; Safirstein, Robert L.; Megyesi, Judit

2009-01-01

Acute kidney injury (AKI) activates pathways of cell death and cell proliferation. Although seemingly discrete and unrelated mechanisms, these pathways can now be shown to be connected and even to be controlled by similar pathways. The dependence of the severity of renal-cell injury on cell cycle pathways can be used to control and perhaps to prevent acute kidney injury. This review is written to address the correlation between cellular life and death in kidney tubules, especially in acute kidney injury. PMID:19536080

At-Sea Test and Evaluation Of Oxygen (O2) Analyzers.

DTIC Science & Technology

1981-04-01

Paramagnetic Oxygen Analyzer 2-6 2.4 Thermomagnetic Oxygen Analyzer Sensor 2-8 2.5 Cell Voltage versus Oxygen Concentration at 2-11 Various Cell ...of flue gas out of the stack across the cell and back into the stack. In-situ units place the cell directly in the flue gas path in the uptake. ) The...repetitive failurc of a cell heater temperature control circuit and a control cabinet electron- ic malfunction. Of the five (5) units that remained in

Memory T cells in organ transplantation: progress and challenges

PubMed Central

Espinosa, Jaclyn R.; Samy, Kannan P.; Kirk, Allan D.

2017-01-01

Antigen-experienced T cells, also known as memory T cells, are functionally and phenotypically distinct from naive T cells. Their enhanced expression of adhesion molecules and reduced requirement for co-stimulation enables them to mount potent and rapid recall responses to subsequent antigen encounters. Memory T cells generated in response to prior antigen exposures can cross-react with other nonidentical, but similar, antigens. This heterologous cross-reactivity not only enhances protective immune responses, but also engenders de novo alloimmunity. This latter characteristic is increasingly recognized as a potential barrier to allograft acceptance that is worthy of immunotherapeutic intervention, and several approaches have been investigated. Calcineurin inhibition effectively controls memory T-cell responses to allografts, but this benefit comes at the expense of increased infectious morbidity. Lymphocyte depletion eliminates allospecific T cells but spares memory T cells to some extent, such that patients do not completely lose protective immunity. Co-stimulation blockade is associated with reduced adverse-effect profiles and improved graft function relative to calcineurin inhibition, but lacks efficacy in controlling memory T-cell responses. Targeting the adhesion molecules that are upregulated on memory T cells might offer additional means to control co-stimulation-blockade-resistant memory T-cell responses. PMID:26923209

Epithelial-mesenchymal transition transcription factors control pluripotent adult stem cell migration in vivo in planarians.

PubMed

Abnave, Prasad; Aboukhatwa, Ellen; Kosaka, Nobuyoshi; Thompson, James; Hill, Mark A; Aboobaker, A Aziz

2017-10-01

Migration of stem cells underpins the physiology of metazoan animals. For tissues to be maintained, stem cells and their progeny must migrate and differentiate in the correct positions. This need is even more acute after tissue damage by wounding or pathogenic infection. Inappropriate migration also underpins metastasis. Despite this, few mechanistic studies address stem cell migration during repair or homeostasis in adult tissues. Here, we present a shielded X-ray irradiation assay that allows us to follow stem cell migration in planarians. We demonstrate the use of this system to study the molecular control of stem cell migration and show that snail-1 , snail-2 and zeb-1 EMT transcription factor homologs are necessary for cell migration to wound sites and for the establishment of migratory cell morphology. We also observed that stem cells undergo homeostatic migration to anterior regions that lack local stem cells, in the absence of injury, maintaining tissue homeostasis. This requires the polarity determinant notum Our work establishes planarians as a suitable model for further in-depth study of the processes controlling stem cell migration in vivo . © 2017. Published by The Company of Biologists Ltd.

Optimal control on bladder cancer growth model with BCG immunotherapy and chemotherapy

NASA Astrophysics Data System (ADS)

Dewi, C.; Trisilowati

2015-03-01

In this paper, an optimal control model of the growth of bladder cancer with BCG (Basil Calmate Guerin) immunotherapy and chemotherapy is discussed. The purpose of this optimal control is to determine the number of BCG vaccine and drug should be given during treatment such that the growth of bladder cancer cells can be suppressed. Optimal control is obtained by applying Pontryagin principle. Furthermore, the optimal control problem is solved numerically using Forward-Backward Sweep method. Numerical simulations show the effectiveness of the vaccine and drug in controlling the growth of cancer cells. Hence, it can reduce the number of cancer cells that is not infected with BCG as well as minimize the cost of the treatment.

HLA-B*57 and IFNL4-Related Polymorphisms Are Associated With Protection Against HIV-1 Disease Progression in Controllers

PubMed Central

Dominguez-Molina, Beatriz; Tarancon-Diez, Laura; Hua, Stephane; Abad-Molina, Cristina; Rodriguez-Gallego, Esther; Machmach, Kawthar; Vidal, Francesc; Tural, Cristina; Moreno, Santiago; Goñi, María José; Ramírez de Arellano, Elena; del Val, Margarita; Gonzalez-Escribano, María Francisca; Del Romero, Jorge; Rodriguez, Carmen; Capa, Laura; Viciana, Pompeyo; Alcamí, José; Yu, Xu G.; Walker, Bruce D.; Leal, Manuel; Lichterfeld, Mathias

2017-01-01

Abstract Background. Human immunodeficiency virus type 1 (HIV-1) controllers maintain HIV-1 viremia at low levels (normally <2000 HIV-RNA copies/mL) without antiretroviral treatment. However, some HIV-1 controllers have evidence of immunologic progression with marked CD4+ T-cell decline. We investigated host genetic factors associated with protection against CD4+ T-cell loss in HIV-1 controllers. Methods. We analyzed the association of interferon-lambda 4 (IFNL4)–related polymorphisms and human leukocyte antigen (HLA)-B haplotypes within long-term nonprogressor HIV-1 controllers (LTNP-Cs; defined by maintaining CD4+ T-cells counts >500 cells/mm3 for more than 7 years after HIV-1 diagnosis) vs non-LTNP-Cs who developed CD4+ T-cell counts <500 cells/mm3. Both a Spanish study cohort (n = 140) and an international validation cohort (n = 914) were examined. Additionally, in a subgroup of individuals, HIV-1–specific T-cell responses and soluble cytokines were analyzed. Results. HLA-B*57 was independently associated with the LTNP-C phenotype (odds ratio [OR], 3.056 [1.029–9.069]; P = .044 and OR, 1.924 [1.252–2.957]; P = .003) while IFNL4 genotypes represented independent factors for becoming non-LTNP-C (TT/TT, ss469415590; OR, 0.401 [0.171–0.942]; P = .036 or A/A, rs12980275; OR, 0.637 [0.434–0.934]; P = .021) in the Spanish and validation cohorts, respectively, after adjusting for sex, age at HIV-1 diagnosis, IFNL4-related polymorphisms, and different HLA-B haplotypes. LTNP-Cs showed lower plasma induced protein 10 (P = .019) and higher IFN-γ (P = .02) levels than the HIV-1 controllers with diminished CD4+ T-cell numbers. Moreover, LTNP-Cs exhibited higher quantities of interleukin (IL)2+CD57- and IFN-γ +CD57- HIV-1–specific CD8+ T cells (P = .002 and .041, respectively) than non-LTNP-Cs. Conclusions. We defined genetic markers able to segregate stable HIV-1 controllers from those who experience CD4+ T-cell decline. These findings allow for identification of HIV-1 controllers at risk for immunologic progression and provide avenues for personalized therapeutic interventions and precision medicine for optimizing clinical care of these individuals. PMID:27986689

Clinostat rotation induces apoptosis in luteal cells of the pregnant rat

NASA Technical Reports Server (NTRS)

Yang, Hyunwon; Bhat, Ganapathy K.; Sridaran, Rajagopala

2002-01-01

Recent studies have shown that microgravity induces changes at the cellular level, including apoptosis. However, it is unknown whether microgravity affects luteal cell function. This study was performed to assess whether microgravity conditions generated by clinostat rotation induce apoptosis and affect steroidogenesis by luteal cells. Luteal cells isolated from the corpora lutea of Day 8 pregnant rats were placed in equal numbers in slide flasks (chamber slides). One slide flask was placed in the clinostat and the other served as a stationary control. At 48 h in the clinostat, whereas the levels of progesterone and total cellular protein decreased, the number of shrunken cells increased. To determine whether apoptosis occurred in shrunken cells, Comet and TUNEL assays were performed. At 48 h, the percentage of apoptotic cells in the clinostat increased compared with that in the control. To investigate how the microgravity conditions induce apoptosis, the active mitochondria in luteal cells were detected with JC-1 dye. Cells in the control consisted of many active mitochondria, which were evenly distributed throughout the cell. In contrast, cells in the clinostat displayed fewer active mitochondria, which were distributed either to the outer edge of the cell or around the nucleus. These results suggest that mitochondrial dysfunction induced by clinostat rotation could lead to apoptosis in luteal cells and suppression of progesterone production.

Multi-dose Romidepsin Reactivates Replication Competent SIV in Post-antiretroviral Rhesus Macaque Controllers

PubMed Central

Policicchio, Benjamin B.; Xu, Cuiling; Brocca-Cofano, Egidio; Raehtz, Kevin D.; He, Tianyu; Ma, Dongzhu; Li, Hui; Haret-Richter, George S.; Dunsmore, Tammy; Trichel, Anita; Mellors, John W.; Hahn, Beatrice H.; Shaw, George M.; Ribeiro, Ruy M.; Pandrea, Ivona; Apetrei, Cristian

2016-01-01

Viruses that persist despite seemingly effective antiretroviral treatment (ART) and can reinitiate infection if treatment is stopped preclude definitive treatment of HIV-1 infected individuals, requiring lifelong ART. Among strategies proposed for targeting these viral reservoirs, the premise of the “shock and kill” strategy is to induce expression of latent proviruses [for example with histone deacetylase inhibitors (HDACis)] resulting in elimination of the affected cells through viral cytolysis or immune clearance mechanisms. Yet, ex vivo studies reported that HDACis have variable efficacy for reactivating latent proviruses, and hinder immune functions. We developed a nonhuman primate model of post-treatment control of SIV through early and prolonged administration of ART and performed in vivo reactivation experiments in controller RMs, evaluating the ability of the HDACi romidepsin (RMD) to reactivate SIV and the impact of RMD treatment on SIV-specific T cell responses. Ten RMs were IV-infected with a SIVsmmFTq transmitted-founder infectious molecular clone. Four RMs received conventional ART for >9 months, starting from 65 days post-infection. SIVsmmFTq plasma viremia was robustly controlled to <10 SIV RNA copies/mL with ART, without viral blips. At ART cessation, initial rebound viremia to ~106 copies/mL was followed by a decline to < 10 copies/mL, suggesting effective immune control. Three post-treatment controller RMs received three doses of RMD every 35–50 days, followed by in vivo experimental depletion of CD8+ cells using monoclonal antibody M-T807R1. RMD was well-tolerated and resulted in a rapid and massive surge in T cell activation, as well as significant virus rebounds (~104 copies/ml) peaking at 5–12 days post-treatment. CD8+ cell depletion resulted in a more robust viral rebound (107 copies/ml) that was controlled upon CD8+ T cell recovery. Our results show that RMD can reactivate SIV in vivo in the setting of post-ART viral control. Comparison of the patterns of virus rebound after RMD administration and CD8+ cell depletion suggested that RMD impact on T cells is only transient and does not irreversibly alter the ability of SIV-specific T cells to control the reactivated virus. PMID:27632364

Increase of CD69, CD161 and CD94 on NK cells in women with recurrent spontaneous abortion and in vitro fertilization failure.

PubMed

Ghafourian, Mehri; Karami, Najmeh; Khodadadi, Ali; Nikbakht, Roshan

2014-06-01

Recurrent spontaneous abortion (RSA) and in vitro fertilization (IVF) failure with unknown causes are the controversial issues that are probably related to the immune system. To compare circulating NK cells expressing activation and inhibition surface markers between patients with RSA and IVF failure with those of healthy multiparous and successful IVF control women, respectively. In this case-control study peripheral blood samples were collected from 43 patients who included 23 women with RSA and 20 with IVF failure, plus 43 healthy control women comprising of 36 normal multiparous women and seven women with successful IVF. The expression of CD69, CD94 and CD161 surface markers on CD56+NK cells were assessed using specific monoclonal antibodies by flowcytometry. The percentage of NK cells increased significantly in patients with RSA and in women with IVF failure in comparison to healthy multiparous and successful IVF control groups (p<0.001). The overall expression of CD69, CD94, CD161 were also increased significantly on NK cells in both patient groups compared to control groups (p<0.001). Elevated expression of CD69 and CD161 on NK cells can be considered as immunological risk markers in RSA and IVF failure. However, it is not clear if high expression of CD94 on peripheral blood NK cells is related to abnormal activity of endometrial NK cells.

Amino acid sequence preferences to control cell-specific organization of endothelial cells, smooth muscle cells, and fibroblasts.

PubMed

Kanie, Kei; Kato, Ryuji; Zhao, Yingzi; Narita, Yuji; Okochi, Mina; Honda, Hiroyuki

2011-06-01

Effective surface modification with biocompatible molecules is known to be effective in reducing the life-threatening risks related to artificial cardiovascular implants. In recent strategies in regenerative medicine, the enhancement and support of natural repair systems at the site of injury by designed biocompatible molecules have succeeded in rapid and effective injury repair. Therefore, such a strategy could also be effective for rapid endothelialization of cardiovascular implants to lower the risk of thrombosis and stenosis. To achieve this enhancement of the natural repair system, a biomimetic molecule that mimics proper cellular organization at the implant location is required. In spite of the fact that many reported peptides have cell-attracting properties on material surfaces, there have been few peptides that could control cell-specific adhesion. For the advanced cardiovascular implants, peptides that can mimic the natural mechanism that controls cell-specific organization have been strongly anticipated. To obtain such peptides, we hypothesized the cellular bias toward certain varieties of amino acids and examined the cell preference (in terms of adhesion, proliferation, and protein attraction) of varieties and of repeat length on SPOT peptide arrays. To investigate the role of specific peptides in controlling the organization of various cardiovascular-related cells, we compared endothelial cells (ECs), smooth muscle cells (SMCs), and fibroblasts (FBs). A clear, cell-specific preference was found for amino acids (longer than 5-mer) using three types of cells, and the combinational effect of the physicochemical properties of the residues was analyzed to interpret the mechanism. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd.

Hippocampal place cell instability after lesions of the head direction cell network

NASA Technical Reports Server (NTRS)

Calton, Jeffrey L.; Stackman, Robert W.; Goodridge, Jeremy P.; Archey, William B.; Dudchenko, Paul A.; Taube, Jeffrey S.; Oman, C. M. (Principal Investigator)

2003-01-01

The occurrence of cells that encode spatial location (place cells) or head direction (HD cells) in the rat limbic system suggests that these cell types are important for spatial navigation. We sought to determine whether place fields of hippocampal CA1 place cells would be altered in animals receiving lesions of brain areas containing HD cells. Rats received bilateral lesions of anterodorsal thalamic nuclei (ADN), postsubiculum (PoS), or sham lesions, before place cell recording. Although place cells from lesioned animals did not differ from controls on many place-field characteristics, such as place-field size and infield firing rate, the signal was significantly degraded with respect to measures of outfield firing rate, spatial coherence, and information content. Surprisingly, place cells from lesioned animals were more likely modulated by the directional heading of the animal. Rotation of the landmark cue showed that place fields from PoS-lesioned animals were not controlled by the cue and shifted unpredictably between sessions. Although fields from ADN-lesioned animals tended to have less landmark control than fields from control animals, this impairment was mild compared with cells recorded from PoS-lesioned animals. Removal of the prominent visual cue also led to instability of place-field representations in PoS-lesioned, but not ADN-lesioned, animals. Together, these findings suggest that an intact HD system is not necessary for the maintenance of place fields, but lesions of brain areas that convey the HD signal can degrade this signal, and lesions of the PoS might lead to perceptual or mnemonic deficits, leading to place-field instability between sessions.

Stable knockdown of PASG enhances DNA demethylation but does not accelerate cellular senescence in TIG-7 human fibroblasts

PubMed Central

Suzuki, Toshikazu; Farrar, Jason E.; Yegnasubramanian, Srinivasan; Zahed, Muhammed; Suzuki, Nobuo; Arceci, Robert J.

2009-01-01

Demethylation of 5-methylcytosine in genomic DNA is believed to be one of the mechanisms underlying replicative life-span of mammalian cells. Both proliferation associated SNF2-like gene (PASG, also termed Lsh) and DNA methyltransferase 3B (Dnmt3b) knockout mice result in embryonic genomic hypomethylation and a replicative senescent phenotype. However, it is unclear whether gradual demethylation of DNA during somatic cell division is directly involved in senescence. In this study, we retrovirally transduced TIG-7 human fibroblasts with a shRNA against PASG and compared the rate of change in DNA methylation as well as the replicative life-span to control cells under low (3%) and ambient (20%) oxygen. Expression of PASG protein was decreased by approximately 80% compared to control cells following transduction of PASG shRNA gene. The rate of cell growth was the same in both control and PASG-suppressed cells. The rate of demethylation of DNA was significantly increased in PASG-suppressed cells as compared control cells. However, decreased PASG expression did not shorten the replicative life-span of TIG-7 cells. Culture under low oxygen extended the life-span of TIG-7 cells but did not alter the rate of DNA demethylation. While knockout of PASG during development results in genomic hypomethylation and premature senescence, our results show that while downregulation of PASG expression in a somatic cell also leads to DNA hypomethylation, there is no associated senescent phenotype. These results suggest differences in cellular consequences of hypomethylation mediated by PASG during development compared to that in somatic cells. PMID:18948754