A Motor-Driven Mechanism for Cell-Length Sensing

PubMed Central

Rishal, Ida; Kam, Naaman; Perry, Rotem Ben-Tov; Shinder, Vera; Fisher, Elizabeth M.C.; Schiavo, Giampietro; Fainzilber, Mike

2012-01-01

Summary Size homeostasis is fundamental in cell biology, but it is not clear how large cells such as neurons can assess their own size or length. We examined a role for molecular motors in intracellular length sensing. Computational simulations suggest that spatial information can be encoded by the frequency of an oscillating retrograde signal arising from a composite negative feedback loop between bidirectional motor-dependent signals. The model predicts that decreasing either or both anterograde or retrograde signals should increase cell length, and this prediction was confirmed upon application of siRNAs for specific kinesin and/or dynein heavy chains in adult sensory neurons. Heterozygous dynein heavy chain 1 mutant sensory neurons also exhibited increased lengths both in vitro and during embryonic development. Moreover, similar length increases were observed in mouse embryonic fibroblasts upon partial downregulation of dynein heavy chain 1. Thus, molecular motors critically influence cell-length sensing and growth control. PMID:22773964

Effects of Telomerase and Telomere Length on Epidermal Stem Cell Behavior

NASA Astrophysics Data System (ADS)

Flores, Ignacio; Cayuela, María L.; Blasco, María A.

2005-08-01

A key process in organ homeostasis is the mobilization of stem cells out of their niches. We show through analysis of mouse models that telomere length, as well as the catalytic component of telomerase, Tert, are critical determinants in the mobilization of epidermal stem cells. Telomere shortening inhibited mobilization of stem cells out of their niche, impaired hair growth, and resulted in suppression of stem cell proliferative capacity in vitro. In contrast, Tert overexpression in the absence of changes in telomere length promoted stem cell mobilization, hair growth, and stem cell proliferation in vitro. The effects of telomeres and telomerase on stem cell biology anticipate their role in cancer and aging.

Telomere shortening in hematopoietic stem cell transplantation: a potential mechanism for late graft failure?

PubMed

Awaya, Norihiro; Baerlocher, Gabriela M; Manley, Thomas J; Sanders, Jean E; Mielcarek, Marco; Torok-Storb, Beverly; Lansdorp, Peter M

2002-01-01

Telomeres serve to maintain the structural integrity of chromosomes, yet each somatic cell division is associated with a decrease in telomere length. The cumulative decrease in telomere length can impose an upper limit for the number of cell divisions that can occur before a cell senesces. When studied in vitro with fibroblasts, this limit is referred to as the Hayflick limit and usually occurs after 40 to 80 cell doublings. In theory, a similar replicative potential in a hematopoietic stem cell could support hematopoiesis in a person for more than 100 years. However, stem cells differentiate, and the telomere length differs among chromosomes within a single cell, among cell types, and among age-matched individuals. This variation in telomere length raises the possibility that long-term hematopoiesis by transplanted stem cells could, depending on the telomere length of the engrafted stem cell and the proliferative demand to which it is subjected, reach a Hayflick limit during the life span of the patient. Although significant shortening of telomeres is reported to occur within the first year posttransplantation, as yet no evidence has indicated that this shortening is associated with marrow function. In this review, we summarize reports on telomere shortening in stem cell transplantation recipients and report 2 cases in which graft failure is associated with significant telomere shortening.

Reconstructing the in vivo dynamics of hematopoietic stem cells from telomere length distributions

PubMed Central

Werner, Benjamin; Beier, Fabian; Hummel, Sebastian; Balabanov, Stefan; Lassay, Lisa; Orlikowsky, Thorsten; Dingli, David; Brümmendorf, Tim H; Traulsen, Arne

2015-01-01

We investigate the in vivo patterns of stem cell divisions in the human hematopoietic system throughout life. In particular, we analyze the shape of telomere length distributions underlying stem cell behavior within individuals. Our mathematical model shows that these distributions contain a fingerprint of the progressive telomere loss and the fraction of symmetric cell proliferations. Our predictions are tested against measured telomere length distributions in humans across all ages, collected from lymphocyte and granulocyte sorted telomere length data of 356 healthy individuals, including 47 cord blood and 28 bone marrow samples. We find an increasing stem cell pool during childhood and adolescence and an approximately maintained stem cell population in adults. Furthermore, our method is able to detect individual differences from a single tissue sample, i.e. a single snapshot. Prospectively, this allows us to compare cell proliferation between individuals and identify abnormal stem cell dynamics, which affects the risk of stem cell related diseases. DOI: http://dx.doi.org/10.7554/eLife.08687.001 PMID:26468615

TP53-dependent chromosome instability is associated with transient reductions in telomere length in immortal telomerase-positive cell lines

NASA Technical Reports Server (NTRS)

Schwartz, J. L.; Jordan, R.; Liber, H.; Murnane, J. P.; Evans, H. H.

2001-01-01

Telomere shortening in telomerase-negative somatic cells leads to the activation of the TP53 protein and the elimination of potentially unstable cells. We examined the effect of TP53 gene expression on both telomere metabolism and chromosome stability in immortal, telomerase-positive cell lines. Telomere length, telomerase activity, and chromosome instability were measured in multiple clones isolated from three related human B-lymphoblast cell lines that vary in TP53 expression; TK6 cells express wild-type TP53, WTK1 cells overexpress a mutant form of TP53, and NH32 cells express no TP53 protein. Clonal variations in both telomere length and chromosome stability were observed, and shorter telomeres were associated with higher levels of chromosome instability. The shortest telomeres were found in WTK1- and NH32-derived cells, and these cells had 5- to 10-fold higher levels of chromosome instability. The primary marker of instability was the presence of dicentric chromosomes. Aneuploidy and other stable chromosome alterations were also found in clones showing high levels of dicentrics. Polyploidy was found only in WTK1-derived cells. Both telomere length and chromosome instability fluctuated in the different cell populations with time in culture, presumably as unstable cells and cells with short telomeres were eliminated from the growing population. Our results suggest that transient reductions in telomere lengths may be common in immortal cell lines and that these alterations in telomere metabolism can have a profound effect on chromosome stability. Copyright 2000 Wiley-Liss, Inc.

Fragment Length of Circulating Tumor DNA

PubMed Central

Underhill, Hunter R.; Kitzman, Jacob O.; Hellwig, Sabine; Welker, Noah C.; Daza, Riza; Gligorich, Keith M.; Rostomily, Robert C.; Shendure, Jay

2016-01-01

Malignant tumors shed DNA into the circulation. The transient half-life of circulating tumor DNA (ctDNA) may afford the opportunity to diagnose, monitor recurrence, and evaluate response to therapy solely through a non-invasive blood draw. However, detecting ctDNA against the normally occurring background of cell-free DNA derived from healthy cells has proven challenging, particularly in non-metastatic solid tumors. In this study, distinct differences in fragment length size between ctDNAs and normal cell-free DNA are defined. Human ctDNA in rat plasma derived from human glioblastoma multiforme stem-like cells in the rat brain and human hepatocellular carcinoma in the rat flank were found to have a shorter principal fragment length than the background rat cell-free DNA (134–144 bp vs. 167 bp, respectively). Subsequently, a similar shift in the fragment length of ctDNA in humans with melanoma and lung cancer was identified compared to healthy controls. Comparison of fragment lengths from cell-free DNA between a melanoma patient and healthy controls found that the BRAF V600E mutant allele occurred more commonly at a shorter fragment length than the fragment length of the wild-type allele (132–145 bp vs. 165 bp, respectively). Moreover, size-selecting for shorter cell-free DNA fragment lengths substantially increased the EGFR T790M mutant allele frequency in human lung cancer. These findings provide compelling evidence that experimental or bioinformatic isolation of a specific subset of fragment lengths from cell-free DNA may improve detection of ctDNA. PMID:27428049

Cell cycle phases in the unequal mother/daughter cell cycles of Saccharomyces cerevisiae.

PubMed

Brewer, B J; Chlebowicz-Sledziewska, E; Fangman, W L

1984-11-01

During cell division in the yeast Saccharomyces cerevisiae mother cells produce buds (daughter cells) which are smaller and have longer cell cycles. We performed experiments to compare the lengths of cell cycle phases in mothers and daughters. As anticipated from earlier indirect observations, the longer cell cycle time of daughter cells is accounted for by a longer G1 interval. The S-phase and the G2-phase are of the same duration in mother and daughter cells. An analysis of five isogenic strains shows that cell cycle phase lengths are independent of cell ploidy and mating type.

Long-Term Effects of Radiation Exposure and Metabolic Status on Telomere Length of Peripheral Blood T Cells in Atomic Bomb Survivors.

PubMed

Yoshida, Kengo; Misumi, Munechika; Kubo, Yoshiko; Yamaoka, Mika; Kyoizumi, Seishi; Ohishi, Waka; Hayashi, Tomonori; Kusunoki, Yoichiro

2016-10-01

In a series of studies of atomic bomb survivors, radiation-dose-dependent alterations in peripheral T-cell populations have been reported. For example, reduced size in naïve T-cell pools and impaired proliferation ability of T cells were observed. Because these alterations are also generally observed with human aging, we hypothesized that radiation exposure may accelerate the aging process of the T-cell immune system. To further test this hypothesis, we conducted cross-sectional analyses of telomere length, a hallmark of cellular aging, of naïve and memory CD4 T cells and total CD8 T cells in the peripheral blood of 620 atomic bomb survivors as it relates to age and radiation dose, using fluorescence in situ hybridization with flow cytometry. Since telomere shortening has been recently demonstrated in obesity-related metabolic abnormalities and diseases, the modifying effects of metabolic status were also examined. Our results indicated nonlinear relationships between T-cell telomere length and prior radiation exposure, i.e., longer telomeres with lower dose exposure and a decreasing trend of telomere length with individuals exposed to doses higher than 0.5 Gy. There were associations between shorter T-cell telomeres and higher hemoglobin Alc levels or fatty liver development. In naïve and memory CD4 T cells, radiation dose and high-density lipoprotein (HDL) cholesterol were found to positively interact with telomere length, suggesting that the decreasing trend of telomere length from a higher radiation dose was less conspicuous in individuals with a higher HDL cholesterol. It is therefore likely that radiation exposure perturbs T-cell homeostasis involving telomere length maintenance by multiple biological mechanisms, depending on dose, and that long-term-radiation-induced effects on the maintenance of T-cell telomeres may be modified by the subsequent metabolic conditions of individuals.

Analysis of Cell Division and Elongation Underlying the Developmental Acceleration of Root Growth in Arabidopsis thaliana1

PubMed Central

Beemster, Gerrit T.S.; Baskin, Tobias I.

1998-01-01

To investigate the relation between cell division and expansion in the regulation of organ growth rate, we used Arabidopsis thaliana primary roots grown vertically at 20°C with an elongation rate that increased steadily during the first 14 d after germination. We measured spatial profiles of longitudinal velocity and cell length and calculated parameters of cell expansion and division, including rates of local cell production (cells mm−1 h−1) and cell division (cells cell−1 h−1). Data were obtained for the root cortex and also for the two types of epidermal cell, trichoblasts and atrichoblasts. Accelerating root elongation was caused by an increasingly longer growth zone, while maximal strain rates remained unchanged. The enlargement of the growth zone and, hence, the accelerating root elongation rate, were accompanied by a nearly proportionally increased cell production. This increased production was caused by increasingly numerous dividing cells, whereas their rates of division remained approximately constant. Additionally, the spatial profile of cell division rate was essentially constant. The meristem was longer than generally assumed, extending well into the region where cells elongated rapidly. In the two epidermal cell types, meristem length and cell division rate were both very similar to that of cortical cells, and differences in cell length between the two epidermal cell types originated at the apex of the meristem. These results highlight the importance of controlling the number of dividing cells, both to generate tissues with different cell lengths and to regulate the rate of organ enlargement. PMID:9536070

Differences in the length of the carboxyl terminus mediate functional properties of neurokinin-1 receptor

PubMed Central

Lai, Jian-Ping; Lai, Saien; Tuluc, Florin; Tansky, Morris F.; Kilpatrick, Laurie E.; Leeman, Susan E.; Douglas, Steven D.

2008-01-01

The neurokinin-1 receptor (NK1R) has two naturally occurring forms that differ in the length of the carboxyl terminus: a full-length receptor consisting of 407 aa and a truncated receptor consisting of 311 aa. We examined whether there are differential signaling properties attributable to the carboxyl terminus of this receptor by using stably transfected human embryonic kidney (HEK293) cell lines that express either full-length or truncated NK1R. Substance P (SP) specifically triggered intracellular calcium increase in HEK293 cells expressing full-length NK1R but had no effect in the cells expressing the truncated NK1R. In addition, in cells expressing full-length NK1R, SP activated NF-κB and IL-8 mRNA expression, but in cells expressing the truncated NK1R, SP did not activate NF-κB, and it decreased IL-8 mRNA expression. In cells expressing full-length NK1R, SP stimulated phosphorylation of PKCδ but inhibited phosphorylation of PKCδ in cells expressing truncated NK1R. There are also differences in the timing of SP-induced ERK activation in cells expressing the two different forms of the receptor. Full-length NK1R activation of ERK was rapid (peak within 1–2 min), whereas truncated NK1R-mediated activation was slower (peak at 20–30 min). Thus, the carboxyl terminus of NK1R is the structural basis for differences in the functional properties of the full-length and truncated NK1R. These differences may provide important information toward the design of new NK1R receptor antagonists. PMID:18713853

Telomere Restriction Fragment (TRF) Analysis.

PubMed

Mender, Ilgen; Shay, Jerry W

2015-11-20

While telomerase is expressed in ~90% of primary human tumors, most somatic tissue cells except transiently proliferating stem-like cells do not have detectable telomerase activity (Shay and Wright, 1996; Shay and Wright, 2001). Telomeres progressively shorten with each cell division in normal cells, including proliferating stem-like cells, due to the end replication (lagging strand synthesis) problem and other causes such as oxidative damage, therefore all somatic cells have limited cell proliferation capacity (Hayflick limit) (Hayflick and Moorhead, 1961; Olovnikov, 1973). The progressive telomere shortening eventually leads to growth arrest in normal cells, which is known as replicative senescence (Shay et al. , 1991). Once telomerase is activated in cancer cells, telomere length is stabilized by the addition of TTAGGG repeats to the end of chromosomes, thus enabling the limitless continuation of cell division (Shay and Wright, 1996; Shay and Wright, 2001). Therefore, the link between aging and cancer can be partially explained by telomere biology. There are many rapid and convenient methods to study telomere biology such as Telomere Restriction Fragment (TRF), Telomere Repeat Amplification Protocol (TRAP) (Mender and Shay, 2015b) and Telomere dysfunction Induced Foci (TIF) analysis (Mender and Shay, 2015a). In this protocol paper we describe Telomere Restriction Fragment (TRF) analysis to determine average telomeric length of cells. Telomeric length can be indirectly measured by a technique called Telomere Restriction Fragment analysis (TRF). This technique is a modified Southern blot, which measures the heterogeneous range of telomere lengths in a cell population using the length distribution of the terminal restriction fragments (Harley et al. , 1990; Ouellette et al. , 2000). This method can be used in eukaryotic cells. The description below focuses on the measurement of human cancer cells telomere length. The principle of this method relies on the lack of restriction enzyme recognition sites within TTAGGG tandem telomeric repeats, therefore digestion of genomic DNA, not telomeric DNA, with a combination of 6 base restriction endonucleases reduces genomic DNA size to less than 800 bp.

Constant volume gas cell optical phase-shifter

DOEpatents

Phillion, Donald W.

2002-01-01

A constant volume gas cell optical phase-shifter, particularly applicable for phase-shifting interferometry, contains a sealed volume of atmospheric gas at a pressure somewhat different than atmospheric. An optical window is present at each end of the cell, and as the length of the cell is changed, the optical path length of a laser beam traversing the cell changes. The cell comprises movable coaxial tubes with seals and a volume equalizing opening. Because the cell is constant volume, the pressure, temperature, and density of the contained gas do not change as the cell changes length. This produces an exactly linear relationship between the change in the length of the gas cell and the change in optical phase of the laser beam traversing it. Because the refractive index difference between the gas inside and the atmosphere outside is very much the same, a large motion must be made to change the optical phase by the small fraction of a wavelength that is required by phase-shifting interferometry for its phase step. This motion can be made to great fractional accuracy.

A potential individual cell malignancy indicator: focal length

NASA Astrophysics Data System (ADS)

Wang, Weina; Lear, Kevin L.

2011-03-01

The label-free technique of optofluidic intracavity spectroscopy (OFIS) utilizes the optical transmission spectrum of a cell in a microfluidic Fabry-Pérot (F-P) cavity to distinguish cells from cancerous cell lines and baseline normal blood cells. The classification between canine hemangiosarcoma (HSA) cancer cells and monocytes in canine normal peripheral blood mononuclear cells (PBMCs) had been demonstrated with 95% sensitivity and 98% specificity. Now with a new optical model that treats the cell settled at the bottom of the cavity as a thin lens, the focal length of cells was extracted and used as an individual cell malignancy indicator.

Prostate Cancer Cell Telomere Length Variability and Stromal Cell Telomere Length as Prognostic Markers for Metastasis and Death

PubMed Central

Heaphy, Christopher M.; Yoon, Ghil Suk; Peskoe, Sarah B.; Joshu, Corinne E.; Lee, Thomas K.; Giovannucci, Edward; Mucci, Lorelei A.; Kenfield, Stacey A.; Stampfer, Meir J.; Hicks, Jessica L.; De Marzo, Angelo M.; Platz, Elizabeth A.; Meeker, Alan K.

2013-01-01

Current prognostic indicators are imperfect predictors of outcome in men with clinicallylocalized prostate cancer. Thus, tissue-based markers are urgently needed to improve treatment and surveillance decision-making. Given that shortened telomeres enhance chromosomal instability and such instability is a hallmark of metastatic lesions, we hypothesized that alterations in telomere length in the primary cancer would predict risk of progression to metastasis and prostate cancer death. To test this hypothesis, we conducted a prospective cohort study of 596 surgically treated men who participated in the ongoing Health Professionals Follow-up Study. Men who had the combination of more variable telomere length among prostate cancer cells (cell-to-cell) and shorter telomere length in prostate cancer-associated stromal cells were substantially more likely to progress to metastasis or die of their prostate cancer. These findings point to the translational potential of this telomere biomarker for prognostication and risk stratification for individualized therapeutic and surveillance strategies. PMID:23779129

Acoustic Liners for Turbine Engines

NASA Technical Reports Server (NTRS)

Jones, Michael G (Inventor); Grady, Joseph E (Inventor); Heidmann, James D. (Inventor); Kiser, James D. (Inventor); Miller, Christopher (Inventor)

2016-01-01

An improved acoustic liner for turbine engines is disclosed. The acoustic liner may include a straight cell section including a plurality of cells with straight chambers. The acoustic liner may also include a bent cell section including one or more cells that are bent to extend chamber length without increasing the overall height of the acoustic liner by the entire chamber length. In some cases, holes are placed between cell chambers in addition to bending the cells, or instead of bending the cells.

Size uniformity of animal cells is actively maintained by a p38 MAPK-dependent regulation of G1-length.

PubMed

Liu, Shixuan; Ginzberg, Miriam Bracha; Patel, Nish; Hild, Marc; Leung, Bosco; Li, Zhengda; Chen, Yen-Chi; Chang, Nancy; Wang, Yuan; Tan, Ceryl; Diena, Shulamit; Trimble, William; Wasserman, Larry; Jenkins, Jeremy L; Kirschner, Marc W; Kafri, Ran

2018-03-29

Animal cells within a tissue typically display a striking regularity in their size. To date, the molecular mechanisms that control this uniformity are still unknown. We have previously shown that size uniformity in animal cells is promoted, in part, by size-dependent regulation of G1 length. To identify the molecular mechanisms underlying this process, we performed a large-scale small molecule screen and found that the p38 MAPK pathway is involved in coordinating cell size and cell cycle progression. Small cells display higher p38 activity and spend more time in G1 than larger cells. Inhibition of p38 MAPK leads to loss of the compensatory G1 length extension in small cells, resulting in faster proliferation, smaller cell size and increased size heterogeneity. We propose a model wherein the p38 pathway responds to changes in cell size and regulates G1 exit accordingly, to increase cell size uniformity. © 2017, Liu et al.

Telomere lengthening and other functions of telomerase.

PubMed

Rubtsova, M P; Vasilkova, D P; Malyavko, A N; Naraikina, Yu V; Zvereva, M I; Dontsova, O A

2012-04-01

Telomerase is an enzyme that maintains the length of the telomere. The telomere length specifies the number of divisions a cell can undergo before it finally dies (i.e. the proliferative potential of cells). For example, telomerase is activated in embryonic cell lines and the telomere length is maintained at a constant level; therefore, these cells have an unlimited fission potential. Stem cells are characterized by a lower telomerase activity, which enables only partial compensation for the shortening of telomeres. Somatic cells are usually characterized by the absence of telomerase activity. Telomere shortening leads to the attainment of the Hayflick limit, the transition of cells to a state of senescence. The cells subsequently enter a state of crisis, accompanied by massive cell death. The surviving cells become cancer cells, which are capable both of dividing indefinitely and maintaining telomere length (usually with the aid of telomerase). Telomerase is a reverse transcriptase. It consists of two major components: telomerase RNA (TER) and reverse transcriptase (TERT). TER is a non-coding RNA, and it contains the region which serves as a template for telomere synthesis. An increasing number of articles focussing on the alternative functions of telomerase components have recently started appearing. The present review summarizes data on the structure, biogenesis, and functions of telomerase.

NADH oxidase activity (NOX) and enlargement of HeLa cells oscillate with two different temperature-compensated period lengths of 22 and 24 minutes corresponding to different NOX forms

NASA Technical Reports Server (NTRS)

Wang, S.; Pogue, R.; Morre, D. M.; Morre, D. J.

2001-01-01

NOX proteins are cell surface-associated and growth-related hydroquinone (NADH) oxidases with protein disulfide-thiol interchange activity. A defining characteristic of NOX proteins is that the two enzymatic activities alternate to generate a regular period length of about 24 min. HeLa cells exhibit at least two forms of NOX. One is tumor-associated (tNOX) and is inhibited by putative quinone site inhibitors (e.g., capsaicin or the antitumor sulfonylurea, LY181984). Another is constitutive (CNOX) and refractory to inhibition. The periodic alternation of activities and drug sensitivity of the NADH oxidase activity observed with intact HeLa cells was retained in isolated plasma membranes and with the solubilized and partially purified enzyme. At least two activities were present. One had a period length of 24 min and the other had a period length of 22 min. The lengths of both the 22 and the 24 min periods were temperature compensated (approximately the same when measured at 17, 27 or 37 degrees C) whereas the rate of NADH oxidation approximately doubled with each 10 degrees C rise in temperature. The rate of increase in cell area of HeLa cells when measured by video-enhanced light microscopy also exhibited a complex period of oscillations reflective of both 22 and 24 min period lengths. The findings demonstrate the presence of a novel oscillating NOX activity at the surface of cancer cells with a period length of 22 min in addition to the constitutive NOX of non-cancer cells and tissues with a period length of 24 min.

Antenna Mechanism of Length Control of Actin Cables

PubMed Central

Mohapatra, Lishibanya; Goode, Bruce L.; Kondev, Jane

2015-01-01

Actin cables are linear cytoskeletal structures that serve as tracks for myosin-based intracellular transport of vesicles and organelles in both yeast and mammalian cells. In a yeast cell undergoing budding, cables are in constant dynamic turnover yet some cables grow from the bud neck toward the back of the mother cell until their length roughly equals the diameter of the mother cell. This raises the question: how is the length of these cables controlled? Here we describe a novel molecular mechanism for cable length control inspired by recent experimental observations in cells. This “antenna mechanism” involves three key proteins: formins, which polymerize actin, Smy1 proteins, which bind formins and inhibit actin polymerization, and myosin motors, which deliver Smy1 to formins, leading to a length-dependent actin polymerization rate. We compute the probability distribution of cable lengths as a function of several experimentally tuneable parameters such as the formin-binding affinity of Smy1 and the concentration of myosin motors delivering Smy1. These results provide testable predictions of the antenna mechanism of actin-cable length control. PMID:26107518

Antenna Mechanism of Length Control of Actin Cables.

PubMed

Mohapatra, Lishibanya; Goode, Bruce L; Kondev, Jane

2015-06-01

Actin cables are linear cytoskeletal structures that serve as tracks for myosin-based intracellular transport of vesicles and organelles in both yeast and mammalian cells. In a yeast cell undergoing budding, cables are in constant dynamic turnover yet some cables grow from the bud neck toward the back of the mother cell until their length roughly equals the diameter of the mother cell. This raises the question: how is the length of these cables controlled? Here we describe a novel molecular mechanism for cable length control inspired by recent experimental observations in cells. This "antenna mechanism" involves three key proteins: formins, which polymerize actin, Smy1 proteins, which bind formins and inhibit actin polymerization, and myosin motors, which deliver Smy1 to formins, leading to a length-dependent actin polymerization rate. We compute the probability distribution of cable lengths as a function of several experimentally tuneable parameters such as the formin-binding affinity of Smy1 and the concentration of myosin motors delivering Smy1. These results provide testable predictions of the antenna mechanism of actin-cable length control.

Telomete length in peripheral blood mononuclear cells is associated with folate status in men

USDA-ARS?s Scientific Manuscript database

Human chromosomes are capped by tandem repeats of DNA and associated proteins termed telomeres. The length of the telomeres is reduced with increasing cell divisions except when the enzyme telomerase is active as seen in stem cells and germ cells. Telomere dysfunction has been associated with deve...

Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs.

PubMed

Hayashi, Tetsutaro; Ozaki, Haruka; Sasagawa, Yohei; Umeda, Mana; Danno, Hiroki; Nikaido, Itoshi

2018-02-12

Total RNA sequencing has been used to reveal poly(A) and non-poly(A) RNA expression, RNA processing and enhancer activity. To date, no method for full-length total RNA sequencing of single cells has been developed despite the potential of this technology for single-cell biology. Here we describe random displacement amplification sequencing (RamDA-seq), the first full-length total RNA-sequencing method for single cells. Compared with other methods, RamDA-seq shows high sensitivity to non-poly(A) RNA and near-complete full-length transcript coverage. Using RamDA-seq with differentiation time course samples of mouse embryonic stem cells, we reveal hundreds of dynamically regulated non-poly(A) transcripts, including histone transcripts and long noncoding RNA Neat1. Moreover, RamDA-seq profiles recursive splicing in >300-kb introns. RamDA-seq also detects enhancer RNAs and their cell type-specific activity in single cells. Taken together, we demonstrate that RamDA-seq could help investigate the dynamics of gene expression, RNA-processing events and transcriptional regulation in single cells.

Single Stem Cell Imaging and Analysis Reveals Telomere Length Differences in Diseased Human and Mouse Skeletal Muscles.

PubMed

Tichy, Elisia D; Sidibe, David K; Tierney, Matthew T; Stec, Michael J; Sharifi-Sanjani, Maryam; Hosalkar, Harish; Mubarak, Scott; Johnson, F Brad; Sacco, Alessandra; Mourkioti, Foteini

2017-10-10

Muscle stem cells (MuSCs) contribute to muscle regeneration following injury. In many muscle disorders, the repeated cycles of damage and repair lead to stem cell dysfunction. While telomere attrition may contribute to aberrant stem cell functions, methods to accurately measure telomere length in stem cells from skeletal muscles have not been demonstrated. Here, we have optimized and validated such a method, named MuQ-FISH, for analyzing telomere length in MuSCs from either mice or humans. Our analysis showed no differences in telomere length between young and aged MuSCs from uninjured wild-type mice, but MuSCs isolated from young dystrophic mice exhibited significantly shortened telomeres. In corroboration, we demonstrated that telomere attrition is present in human dystrophic MuSCs, which underscores its importance in diseased regenerative failure. The robust technique described herein provides analysis at a single-cell resolution and may be utilized for other cell types, especially rare populations of cells. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

High-resolution telomere fluorescence in situ hybridization reveals intriguing anomalies in germ cell tumors.

PubMed

Shekhani, Mohammed Talha; Barber, John R; Bezerra, Stephania M; Heaphy, Christopher M; Gonzalez Roibon, Nilda Diana; Taheri, Diana; Reis, Leonardo O; Guner, Gunes; Joshu, Corinne E; Netto, George J; Meeker, Alan K

2016-08-01

Testicular germ cell tumor (TGCT) is the most common malignancy of young men. Most patients are completely cured, which distinguishes these from most other malignancies. Orchiectomy specimens (n=76) were evaluated using high-resolution (single-cell discriminative) telomere-specific fluorescence in situ hybridization (FISH) with simultaneous Oct4 immunofluorescence to describe telomere length phenotype in TGCT neoplastic cells. For the first time, the TGCT precursor lesion, germ cell neoplasia in situ (GCNIS) is also evaluated in depth. The intensity of the signals from cancerous cells was compared to the same patient's reference cells-namely, healthy germ cells (defined as "medium" length) and interstitial/somatic cells (defined as "short" telomere length). We observed short telomeres in most GCNIS and pure seminomas (P=.006 and P=.0005, respectively). In contrast, nonseminomas displayed longer telomeres. Lesion-specific telomere lengths were documented in mixed tumor cases. Embryonal carcinoma (EC) demonstrated the longest telomeres. A fraction of EC displays the telomerase-independent alternative lengthening of telomeres (ALT) phenotype (24% of cases). Loss of ATRX or DAXX nuclear expression was strongly associated with ALT; however, nuclear expression of both proteins was retained in half of ALT-positive ECs. The particular distribution of telomere lengths among TGCT and GCNIS precursors implicate telomeres anomalies in pathogenesis. These results may advise management decisions as well. Copyright © 2016. Published by Elsevier Inc.

Automatic cell identification and visualization using digital holographic microscopy with head mounted augmented reality devices.

PubMed

O'Connor, Timothy; Rawat, Siddharth; Markman, Adam; Javidi, Bahram

2018-03-01

We propose a compact imaging system that integrates an augmented reality head mounted device with digital holographic microscopy for automated cell identification and visualization. A shearing interferometer is used to produce holograms of biological cells, which are recorded using customized smart glasses containing an external camera. After image acquisition, segmentation is performed to isolate regions of interest containing biological cells in the field-of-view, followed by digital reconstruction of the cells, which is used to generate a three-dimensional (3D) pseudocolor optical path length profile. Morphological features are extracted from the cell's optical path length map, including mean optical path length, coefficient of variation, optical volume, projected area, projected area to optical volume ratio, cell skewness, and cell kurtosis. Classification is performed using the random forest classifier, support vector machines, and K-nearest neighbor, and the results are compared. Finally, the augmented reality device displays the cell's pseudocolor 3D rendering of its optical path length profile, extracted features, and the identified cell's type or class. The proposed system could allow a healthcare worker to quickly visualize cells using augmented reality smart glasses and extract the relevant information for rapid diagnosis. To the best of our knowledge, this is the first report on the integration of digital holographic microscopy with augmented reality devices for automated cell identification and visualization.

Prostate stromal cell telomere shortening is associated with risk of prostate cancer in the placebo arm of the Prostate Cancer Prevention Trial*

PubMed Central

Heaphy, Christopher M.; Gaonkar, Gaurav; Peskoe, Sarah B.; Joshu, Corinne E.; De Marzo, Angelo M.; Lucia, M. Scott; Goodman, Phyllis J.; Lippman, Scott M.; Thompson, Ian M.; Platz, Elizabeth A.; Meeker, Alan K.

2015-01-01

Background Telomeres are repetitive nucleoproteins that help maintain chromosomal stability by inhibiting exonucleolytic degradation, prohibiting inappropriate homologous recombination, and preventing chromosomal fusions by suppressing double-strand break signals. We recently observed that men treated for clinically localized prostate cancer with shorter telomeres in their cancer-associated stromal cells, in combination with greater variation in cancer cell telomere lengths, were significantly more likely to progress to distant metastases and die from their disease. Here, we hypothesized that shorter stromal cell telomere length would be associated with prostate cancer risk at time of biopsy. Methods Telomere-specific fluorescence in situ hybridization (FISH) analysis was performed in normal-appearing stromal, basal epithelial, and luminal epithelial cells in biopsies from men randomized to the placebo arm of the Prostate Cancer Prevention Trial. Prostate cancer cases (N=32) were either detected on a biopsy performed for cause or at the end of the study per trial protocol, and controls (N=50), defined as negative for cancer on an end-of-study biopsy performed per trial protocol (e.g. irrespective of indication), were sampled. Logistic regression was used to estimate the association between mean telomere length of the particular cell populations, cell-to-cell telomere length variability, and risk of prostate cancer. Results Men with short stromal cell telomere lengths (below median) had 2.66 (95% CI 1.04-3.06; p=0.04) times the odds of prostate cancer compared with men who had longer lengths (at or above median). Conversely, we did not observe statistically significant associations for short telomere lengths in normal-appearing basal (OR=2.15, 95% CI 0.86-5.39; p=0.10) or luminal (OR=1.15, 95% CI 0.47-2.80; p=0.77) cells. Conclusions These findings suggest that telomere shortening in normal stromal cells is associated with prostate cancer risk. It is essential to extend and validate these findings, while also identifying the cellular milieu that comprises the subset of cells with short telomeres within the prostate tumor microenvironment. PMID:25893825

Prostate stromal cell telomere shortening is associated with risk of prostate cancer in the placebo arm of the Prostate Cancer Prevention Trial.

PubMed

Heaphy, Christopher M; Gaonkar, Gaurav; Peskoe, Sarah B; Joshu, Corinne E; De Marzo, Angelo M; Lucia, M Scott; Goodman, Phyllis J; Lippman, Scott M; Thompson, Ian M; Platz, Elizabeth A; Meeker, Alan K

2015-08-01

Telomeres are repetitive nucleoproteins that help maintain chromosomal stability by inhibiting exonucleolytic degradation, prohibiting inappropriate homologous recombination, and preventing chromosomal fusions by suppressing double-strand break signals. We recently observed that men treated for clinically localized prostate cancer with shorter telomeres in their cancer-associated stromal cells, in combination with greater variation in cancer cell telomere lengths, were significantly more likely to progress to distant metastases, and die from their disease. Here, we hypothesized that shorter stromal cell telomere length would be associated with prostate cancer risk at time of biopsy. Telomere-specific fluorescence in situ hybridization (FISH) analysis was performed in normal-appearing stromal, basal epithelial, and luminal epithelial cells in biopsies from men randomized to the placebo arm of the Prostate Cancer Prevention Trial. Prostate cancer cases (N = 32) were either detected on a biopsy performed for cause or at the end of the study per trial protocol, and controls (N = 50), defined as negative for cancer on an end-of-study biopsy performed per trial protocol (e.g., irrespective of indication), were sampled. Logistic regression was used to estimate the association between mean telomere length of the particular cell populations, cell-to-cell telomere length variability, and risk of prostate cancer. Men with short stromal cell telomere lengths (below median) had 2.66 (95% CI 1.04-3.06; P = 0.04) times the odds of prostate cancer compared with men who had longer lengths (at or above median). Conversely, we did not observe statistically significant associations for short telomere lengths in normal-appearing basal (OR = 2.15, 95% CI 0.86-5.39; P= 0 .10) or luminal (OR = 1.15, 95% CI 0.47-2.80; P = 0.77) cells. These findings suggest that telomere shortening in normal stromal cells is associated with prostate cancer risk. It is essential to extend and validate these findings, while also identifying the cellular milieu that comprises the subset of cells with short telomeres within the prostate tumor microenvironment. © 2015 Wiley Periodicals, Inc.

Acetylcholine, carbachol, and GABA induce no detectable change in the length of isolated outer hair cells.

PubMed

Bobbin, R P; Fallon, M; Puel, J L; Bryant, G; Bledsoe, S C; Zajic, G; Schacht, J

1990-08-01

The mechanical and electrical properties of cochlear outer hair cells (OHCs) are suggested to modulate transduction by inner hair cells. These properties of OHCs are presumably regulated by efferent neurons which use several transmitters including acetylcholine (Ach) and gamma aminobutyric acid (GABA). Since it had been suggested that Ach causes isolated OHCs to shorten visibly, this study was designed to investigate whether GABA also alters the length of OHCs. OHCs were isolated from the guinea pig cochlea by mechanical dispersion after collagenase treatment. Cells were initially selected by strict morphological criteria. In addition they were only included in further studies if they attained a constant length during 10 min of superfusion with buffer solution. Neither GABA (20 microM: 100 microM), Ach (5 mM; 10 microM with 10 microM eserine) or carbachol (10 microM; 100 microM) altered OHC length when applied in iso-osmotic Hank's balanced salt solution (total number of cells tested, 72). If a change in length occurred it must have been smaller than 0.3 microns, our detection ability. In contrast, high potassium and variations in osmolarity changed hair cell length by 3-10% in agreement with other reports.

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.

Probing cooperative force generation in collective cancer invasion

NASA Astrophysics Data System (ADS)

Alobaidi, Amani A.; Xu, Yaopengxiao; Chen, Shaohua; Jiao, Yang; Sun, Bo

2017-08-01

Collective cellular dynamics in the three-dimensional extracellular matrix (ECM) plays a crucial role in many physiological processes such as cancer invasion. Both chemical and mechanical signaling support cell-cell communications on a variety of length scales, leading to collective migratory behaviors. Here we conduct experiments using 3D in vitro tumor models and develop a phenomenological model in order to probe the cooperativity of force generation in the collective invasion of breast cancer cells. In our model, cell-cell communication is characterized by a single parameter that quantifies the correlation length of cellular migration cycles. We devise a stochastic reconstruction method to generate realizations of cell colonies with specific contraction phase correlation functions and correlation length a. We find that as a increases, the characteristic size of regions containing cells with similar contraction phases grows. For small a values, the large fluctuations in individual cell contraction phases smooth out the temporal fluctuations in the time-dependent deformation field in the ECM. For large a values, the periodicity of an individual cell contraction cycle is clearly manifested in the temporal variation of the overall deformation field in the ECM. Through quantitative comparisons of the simulated and experimentally measured deformation fields, we find that the correlation length for collective force generation in the breast cancer diskoid in geometrically micropatterned ECM (DIGME) system is a≈ 25~μ \\text{m} , which is roughly twice the linear size of a single cell. One possible mechanism for this intermediate cell correlation length is the fiber-mediated stress propagation in the 3D ECM network in the DIGME system.

Design and fabrication of segmented-in-series solid oxide fuel cells

NASA Astrophysics Data System (ADS)

Lai, Tammy S.

Segmented-in-series solid oxide fuel cells (SS-SOFC) consist of several thick film cells deposited onto a porous, flattened tubular substrate. SS-SOFCs have a reduced need for gas-tight seals relative to planar SOFCs and can have a short current path compared to tubular SOFCs, limiting electrode ohmic resistance. Like tubular SOFCs, SS-SOFCs are suitable for stationary power generation. Their potentially small cell size makes them candidates for portable applications as well. The goals of this thesis project were to develop SS-SOFCs with 1-2 mm cell lengths and to analyze the effects of cell geometry and support current shunting on performance. Standard SOFC materials were chosen for the active components: yttria stabilized zirconia (YSZ) electrolyte; Ni-YSZ cermet anode; and (La,Sr)MnO 3-based cathode. A Pt-YSZ cermet was used as the interconnect material. Screen printing was the deposition method for all layers due to its low cost and patterning ability. A power density of >900 mW/cm2 was achieved with a cathode sheet resistance of ≈3 O/□ (≈90 mum LSM thickness). A D-optimal study was conducted to find processing conditions yielding substrates with ≥30 vol% porosity and high strength. Uniaxially pressed partially stabilized zirconia (PSZ) with 15 wt% starch pore former met the requirements, though 20 wt% graphite pore former was later found to give a smoother surface that improved screen printed layer quality. Calculations presented in this thesis take into account losses due to cell resistances, electrode ohmic resistances, interconnect resistance, and shunting by a weakly-conductive support material. Power density was maximized at an optimal cell length---it decreased at larger cell lengths due to electrode lateral resistance loss and at smaller cell lengths due to a decreasing fraction of cell active area. Assuming dimensions expected for screen printing and typical area specific resistances (RAS), optimal cell lengths typically ranged from 1 to 3 mm. The calculated and experimental values for the array RAS (active and inactive areas) showed similar dependences on cathode sheet resistance. The impact of shunting current increased with decreasing cell lengths. Shunting current was predicted to decrease array current by ˜10% for a 1.5 mm active cell length, though experimental measurements suggest that the calculation may overestimate the shunting effect.

Increased telomere length and proliferative potential in peripheral blood mononuclear cells of adults of different ages stimulated with concanavalin A

PubMed Central

2013-01-01

Background Recently, a direct correlation with telomere length, proliferative potential and telomerase activity has been found in the process of aging in peripheral blood cells. The objective of the study was to evaluate telomere length and proliferative potential in peripheral blood mononuclear cells (PBMCs) after stimulation with Concanavalin A (ConA) of young adults compared with older adults. Methods Blood samples were obtained from 20 healthy young males (20–25 years old) (group Y) and 20 males (60–65 years old) (group O). We compared PBMC proliferation before and after stimulation with ConA. DNA was isolated from cells separated before and after culture with ConA for telomeric measurement by real-time polymerase chain reaction. Results In vitro stimulation of PBMCs from young subjects induced an increase of telomere length as well as a higher replicative capacity of cell proliferation. Samples from older adults showed higher loss of telomeric DNA (p = 0.03) and higher levels of senescent (≤6.2 kb) telomeric DNA (p = 0.02) and displayed a marked decrease of proliferation capacity. Viability cell counts and CFSE tracking in 72-h-old cell cultures indicated that group O PBMCs (CD8+ and CD4+ T cells) underwent fewer mitotic cycles and had shorter telomeres than group Y (p = 0.04). Conclusions Our findings confirm that telomere length in older-age adults is shorter than in younger subjects. After stimulation with ConA, cells are not restored to the previous telomere length and undergo replicative senescence. This is in sharp contrast to the response observed in young adults after ConA stimulation where cells increase in telomere length and replicative capacity. The mechanisms involved in this phenomenon are not yet clear and merit further investigation. PMID:24063536

Evaluation of telomere length in human cardiac tissues using cardiac quantitative FISH.

PubMed

Sharifi-Sanjani, Maryam; Meeker, Alan K; Mourkioti, Foteini

2017-09-01

Telomere length has been correlated with various diseases, including cardiovascular disease and cancer. The use of currently available telomere-length measurement techniques is often restricted by the requirement of a large amount of cells (Southern-based techniques) or the lack of information on individual cells or telomeres (PCR-based methods). Although several methods have been used to measure telomere length in tissues as a whole, the assessment of cell-type-specific telomere length provides valuable information on individual cell types. The development of fluorescence in situ hybridization (FISH) technologies enables the quantification of telomeres in individual chromosomes, but the use of these methods is dependent on the availability of isolated cells, which prevents their use with fixed archival samples. Here we describe an optimized quantitative FISH (Q-FISH) protocol for measuring telomere length that bypasses the previous limitations by avoiding contributions from undesired cell types. We have used this protocol on small paraffin-embedded cardiac-tissue samples. This protocol describes step-by-step procedures for tissue preparation, permeabilization, cardiac-tissue pretreatment and hybridization with a Cy3-labeled telomeric repeat complementing (CCCTAA) 3 peptide nucleic acid (PNA) probe coupled with cardiac-specific antibody staining. We also describe how to quantify telomere length by means of the fluorescence intensity and area of each telomere within individual nuclei. This protocol provides comparative cell-type-specific telomere-length measurements in relatively small human cardiac samples and offers an attractive technique to test hypotheses implicating telomere length in various cardiac pathologies. The current protocol (from tissue collection to image procurement) takes ∼28 h along with three overnight incubations. We anticipate that the protocol could be easily adapted for use on different tissue types.

Modeling Cancer Cell Growth Dynamics In vitro in Response to Antimitotic Drug Treatment

PubMed Central

Lorz, Alexander; Botesteanu, Dana-Adriana; Levy, Doron

2017-01-01

Investigating the role of intrinsic cell heterogeneity emerging from variations in cell-cycle parameters and apoptosis is a crucial step toward better informing drug administration. Antimitotic agents, widely used in chemotherapy, target exclusively proliferative cells and commonly induce a prolonged mitotic arrest followed by cell death via apoptosis. In this paper, we developed a physiologically motivated mathematical framework for describing cancer cell growth dynamics that incorporates the intrinsic heterogeneity in the time individual cells spend in the cell-cycle and apoptosis process. More precisely, our model comprises two age-structured partial differential equations for the proliferative and apoptotic cell compartments and one ordinary differential equation for the quiescent compartment. To reflect the intrinsic cell heterogeneity that governs the growth dynamics, proliferative and apoptotic cells are structured in “age,” i.e., the amount of time remaining to be spent in each respective compartment. In our model, we considered an antimitotic drug whose effect on the cellular dynamics is to induce mitotic arrest, extending the average cell-cycle length. The prolonged mitotic arrest induced by the drug can trigger apoptosis if the time a cell will spend in the cell cycle is greater than the mitotic arrest threshold. We studied the drug’s effect on the long-term cancer cell growth dynamics using different durations of prolonged mitotic arrest induced by the drug. Our numerical simulations suggest that at confluence and in the absence of the drug, quiescence is the long-term asymptotic behavior emerging from the cancer cell growth dynamics. This pattern is maintained in the presence of small increases in the average cell-cycle length. However, intermediate increases in cell-cycle length markedly decrease the total number of cells and can drive the cancer population to extinction. Intriguingly, a large “switch-on/switch-off” increase in the average cell-cycle length maintains an active cell population in the long term, with oscillating numbers of proliferative cells and a relatively constant quiescent cell number. PMID:28913178

Can Tissue Cilia Lengths and Urine Cilia Proteins Be Markers of Kidney Diseases?

PubMed

Park, Kwon Moo

2018-05-01

The primary cilium is an organelle which consists of a microtubule in the core and a surrounding cilia membrane, and has long been recognized as a "vestigial organelle". However, new evidence demonstrates that the primary cilium has a notable effect on signal transduction in the cell and is associated with some genetic and non-genetic diseases. In the kidney, the primary cilium protrudes into the Bowman's space and the tubular lumen from the apical side of epithelial cells. The length of primary cilia is dynamically altered during the normal cell cycle, being shortened by retraction into the cell body at the entry of cell division and elongated at differentiation. Furthermore, the length of primary cilia is also dynamically changed in the cells, as a result and/or cause, during the progression of various kidney diseases including acute kidney injury and chronic kidney disease. Notably, recent data has demonstrated that the shortening of the primary cilium in the cell is associated with fragmentation, apart from retraction into the cell body, in the progression of diseases and that the fragmented primary cilia are released into the urine. This data reveals that the alteration of primary cilia length could be related to the progression of diseases. This review will consider if primary cilia length alteration is associated with the progression of kidney diseases and if the length of tissue primary cilia and the presence or increase of cilia proteins in the urine is indicative of kidney diseases.

Microscopic optical path length difference and polarization measurement system for cell analysis

NASA Astrophysics Data System (ADS)

Satake, H.; Ikeda, K.; Kowa, H.; Hoshiba, T.; Watanabe, E.

2018-03-01

In recent years, noninvasive, nonstaining, and nondestructive quantitative cell measurement techniques have become increasingly important in the medical field. These cell measurement techniques enable the quantitative analysis of living cells, and are therefore applied to various cell identification processes, such as those determining the passage number limit during cell culturing in regenerative medicine. To enable cell measurement, we developed a quantitative microscopic phase imaging system based on a Mach-Zehnder interferometer that measures the optical path length difference distribution without phase unwrapping using optical phase locking. The applicability of our phase imaging system was demonstrated by successful identification of breast cancer cells amongst normal cells. However, the cell identification method using this phase imaging system exhibited a false identification rate of approximately 7%. In this study, we implemented a polarimetric imaging system by introducing a polarimetric module to one arm of the Mach-Zehnder interferometer of our conventional phase imaging system. This module was comprised of a quarter wave plate and a rotational polarizer on the illumination side of the sample, and a linear polarizer on the optical detector side. In addition, we developed correction methods for the measurement errors of the optical path length and birefringence phase differences that arose through the influence of elements other than cells, such as the Petri dish. As the Petri dish holding the fluid specimens was transparent, it did not affect the amplitude information; however, the optical path length and birefringence phase differences were affected. Therefore, we proposed correction of the optical path length and birefringence phase for the influence of elements other than cells, as a prerequisite for obtaining highly precise phase and polarimetric images.

Mechanisms limiting the performance of large grain polycrystalline silicon solar cells

NASA Technical Reports Server (NTRS)

Culik, J. S.; Alexander, P.; Dumas, K. A.; Wohlgemuth, J. W.

1984-01-01

The open-circuit voltage and short-circuit current of large-grain (1 to 10 mm grain diameter) polycrystalline silicon solar cells is determined by the minority-carrier diffusion length within the bulk of the grains. This was demonstrated by irradiating polycrystalline and single-crystal (Czochralski) silicon solar cells with 1 MeV electrons to reduce their bulk lifetime. The variation of short-circuit current with minority-carrier diffusion length for the polycrystalline solar cells is identical to that of the single-crystal solar cells. The open-circuit voltage versus short-circuit current characteristic of the polycrystalline solar cells for reduced diffusion lengths is also identical to that of the single-crystal solar cells. The open-circuit voltage of the polycrystalline solar cells is a strong function of quasi-neutral (bulk) recombination, and is reduced only slightly, if at all, by grain-boundary recombination.

Microtubule Dynamics Scale with Cell Size to Set Spindle Length and Assembly Timing.

PubMed

Lacroix, Benjamin; Letort, Gaëlle; Pitayu, Laras; Sallé, Jérémy; Stefanutti, Marine; Maton, Gilliane; Ladouceur, Anne-Marie; Canman, Julie C; Maddox, Paul S; Maddox, Amy S; Minc, Nicolas; Nédélec, François; Dumont, Julien

2018-05-21

Successive cell divisions during embryonic cleavage create increasingly smaller cells, so intracellular structures must adapt accordingly. Mitotic spindle size correlates with cell size, but the mechanisms for this scaling remain unclear. Using live cell imaging, we analyzed spindle scaling during embryo cleavage in the nematode Caenorhabditis elegans and sea urchin Paracentrotus lividus. We reveal a common scaling mechanism, where the growth rate of spindle microtubules scales with cell volume, which explains spindle shortening. Spindle assembly timing is, however, constant throughout successive divisions. Analyses in silico suggest that controlling the microtubule growth rate is sufficient to scale spindle length and maintain a constant assembly timing. We tested our in silico predictions to demonstrate that modulating cell volume or microtubule growth rate in vivo induces a proportional spindle size change. Our results suggest that scalability of the microtubule growth rate when cell size varies adapts spindle length to cell volume. Copyright © 2018 Elsevier Inc. All rights reserved.

Recent Advances in Insect Olfaction, Specifically Regarding the Morphology and Sensory Physiology of Antennal Sensilla of the Female Sphinx Moth Manduca sexta

PubMed Central

SHIELDS, VONNIE D.C.; HILDEBRAND, JOHN G.

2008-01-01

The antennal flagellum of female Manduca sexta bears eight sensillum types: two trichoid, two basiconic, one auriculate, two coeloconic, and one styliform complex sensilla. The first type of trichoid sensillum averages 34 μm in length and is innervated by two sensory cells. The second type averages 26 μm in length and is innervated by either one or three sensory cells. The first type of basiconic sensillum averages 22 μm in length, while the second type averages 15 μm in length. Both types are innervated by three bipolar sensory cells. The auriculate sensillum averages 4 μm in length and is innervated by two bipolar sensory cells. The coeloconic type-A and type-B both average 2 μm in length. The former type is innervated by five bipolar sensory cells, while the latter type, by three bipolar sensory cells. The styliform complex sensillum occurs singly on each annulus and averages 38-40 μm in length. It is formed by several contiguous sensilla. Each unit is innervated by three bipolar sensory cells. A total of 2,216 sensilla were found on a single annulus (annulus 21) of the flagellum. Electrophysiological responses from type-A trichoid sensilla to a large panel of volatile odorants revealed three different subsets of olfactory receptor cells (ORCs). Two subsets responded strongly to only a narrow range of odorants, while the third responded strongly to a broad range of odorants. Anterograde labeling of ORCs from type-A trichoid sensilla revealed that their axons projected mainly to two large female glomeruli of the antennal lobe. PMID:11754510

Manipulation of cells' position across a microfluidic channel using a series of continuously varying herringbone structures

NASA Astrophysics Data System (ADS)

Jung, Yugyung; Hyun, Ji-chul; Choi, Jongchan; Atajanov, Arslan; Yang, Sung

2017-12-01

Controlling cells' movement is an important technique in biological analysis that is performed within a microfluidic system. Many external forces are utilized for manipulation of cells, including their position in the channel. These forces can effectively control cells in a desired manner. Most of techniques used to manipulate cells require sophisticated set-ups and equipment to generate desired effect. The exception to this is the use of hydrodynamic force. In this study, a series of continuously varying herringbone structures is proposed for positioning cells in a microfluidic channel using hydrodynamic force. This structure was experimentally developed by changing parameters, such as the length of the herringbone's apex, the length of the herringbone's base and the ratio of the height of the flat channel to the height of the herringbone structure. Results of this study, have demonstrated that the length of the herringbone's apex and the ratio of the heights of the flat channel and the herringbone structure were crucial parameters influencing positioning of cells at 100 μl/h flow rate. The final design was fixed at 170 and 80 μm for the length of herringbone's apex and the length of herringbone's base, respectively. The average position of cells in this device was 34 μm away from the side wall in a 200 μm wide channel. Finally, to substantiate a practical application of the herringbone structure for positioning, cells were randomly introduced into a microfluidic device, containing an array of trapping structures together with a series of herringbone structures along the channel. The cells were moved toward the trapping structure by the herringbone structure and the trapping efficiency was increased. Therefore, it is anticipated that this device will be utilized to continuously control cells' position without application of external forces.

Intrinsic curvature in wool fibres is determined by the relative length of orthocortical and paracortical cells.

PubMed

Harland, Duane P; Vernon, James A; Woods, Joy L; Nagase, Shinobu; Itou, Takashi; Koike, Kenzo; Scobie, David A; Grosvenor, Anita J; Dyer, Jolon M; Clerens, Stefan

2018-03-22

Hair curvature underpins structural diversity and function in mammalian coats, but what causes curl in keratin hair fibres? To obtain structural data to determine one aspect of this question, we used confocal microscopy to provide in situ measurements of the two cell types that make up the cortex of merino wool fibres, which was chosen as a well-characterised model system representative of narrow diameter hairs, such as underhairs. We measured orthocortical and paracortical cross-sectional areas, and cortical cell lengths, within individual fibre snippets of defined uniplanar curvature. This allowed a direct test of two long-standing theories of the mechanism of curvature in hairs. We found evidence contradicting the theory that curvature results from there being more cells on the side of the fibre closest to the outside, or convex edge, of curvature. In all cases, the orthocortical cells close to the outside of curvature were longer than paracortical cells close to the inside of the curvature, which supports the theory that curvature is underpinned by differences in cell type length. However, the latter theory also implies that, for all fibres, curvature should correlate with the proportions of orthocortical and paracortical cells, and we found no evidence for this. In merino wool, it appears that the absolute length of cells of each type and proportion of cells varies from fibre to fibre, and only the difference between the length of the two cell types is important. Implications for curvature in higher diameter hairs, such as guard hairs and those on the human scalp, are discussed. © 2018. Published by The Company of Biologists Ltd.

Tula hantavirus isolate with the full-length ORF for nonstructural protein NSs survives for more consequent passages in interferon-competent cells than the isolate having truncated NSs ORF.

PubMed

Jääskeläinen, Kirsi M; Plyusnina, Angelina; Lundkvist, Ake; Vaheri, Antti; Plyusnin, Alexander

2008-01-11

The competitiveness of two Tula hantavirus (TULV) isolates, TULV/Lodz and TULV/Moravia, was evaluated in interferon (IFN) -competent and IFN-deficient cells. The two isolates differ in the length of the open reading frame (ORF) encoding the nonstructural protein NSs, which has previously been shown to inhibit IFN response in infected cells. In IFN-deficient Vero E6 cells both TULV isolates survived equally well. In contrast, in IFN-competent MRC5 cells TULV/Lodz isolate, that possesses the NSs ORF for the full-length protein of 90 aa, survived for more consequent passages than TULV/Moravia isolate, which contains the ORF for truncated NSs protein (66-67 aa). Our data show that expression of a full-length NSs protein is beneficial for the virus survival and competitiveness in IFN-competent cells and not essential in IFN-deficient cells. These results suggest that the N-terminal aa residues are important for the full activity of the NSs protein.

Strigolactones Stimulate Internode Elongation Independently of Gibberellins1[C][W

PubMed Central

de Saint Germain, Alexandre; Ligerot, Yasmine; Dun, Elizabeth A.; Pillot, Jean-Paul; Ross, John J.; Beveridge, Christine A.; Rameau, Catherine

2013-01-01

Strigolactone (SL) mutants in diverse species show reduced stature in addition to their extensive branching. Here, we show that this dwarfism in pea (Pisum sativum) is not attributable to the strong branching of the mutants. The continuous supply of the synthetic SL GR24 via the root system using hydroponics can restore internode length of the SL-deficient rms1 mutant but not of the SL-response rms4 mutant, indicating that SLs stimulate internode elongation via RMS4. Cytological analysis of internode epidermal cells indicates that SLs control cell number but not cell length, suggesting that SL may affect stem elongation by stimulating cell division. Consequently, SLs can repress (in axillary buds) or promote (in the stem) cell division in a tissue-dependent manner. Because gibberellins (GAs) increase internode length by affecting both cell division and cell length, we tested if SLs stimulate internode elongation by affecting GA metabolism or signaling. Genetic analyses using SL-deficient and GA-deficient or DELLA-deficient double mutants, together with molecular and physiological approaches, suggest that SLs act independently from GAs to stimulate internode elongation. PMID:23943865

Ca2+ and MgATP2- dependence of shortening in skinned single smooth muscle cells.

PubMed

Warshaw, D M; McBride, W J; Hubbard, M S

1987-04-01

Most studies of skinned smooth muscle have been performed in whole tissue preparations. In this study, we report the development of a chemically skinned single smooth muscle cell preparation from the toad, Bufo marinus, stomach. Isolated smooth muscle cells were skinned using saponin. The effect of various ionic environments (i.e., changing free Ca2+ and MgATP2-) on skinned cell contractile response was assessed by measuring cell lengths from populations of cells using a computer-assisted length-measuring system. Comparison of cell length histograms were used to determine the extent of cell shortening in response to a given ionic perturbation. Once skinned, the single cells shortened with a sensitivity to free calcium (ED50 = 1.5 microM Ca2+) that was three orders of magnitude lower than potassium depolarized cells (ED50 = 1.5 mM Ca2+). In addition to the calcium sensitivity, the effect of free MgATP2- on the extent of cell shortening was investigated. The extent of cell shortening was dependent on free MgATP2- with the maximum shortening response occurring at MgATP2- greater than 1 mM.

Variable TERRA abundance and stability in cervical cancer cells.

PubMed

Oh, Bong-Kyeong; Keo, Ponnarath; Bae, Jaeman; Ko, Jung Hwa; Choi, Joong Sub

2017-06-01

Telomeres are transcribed into long non-coding RNA, referred to as telomeric repeat-containing RNA (TERRA), which plays important roles in maintaining telomere integrity and heterochromatin formation. TERRA has been well characterized in HeLa cells, a type of cervical cancer cell. However, TERRA abundance and stability have not been examined in other cervical cancer cells, at least to the best of our knowledge. Thus, in this study, we measured TERRA levels and stability, as well as telomere length in 6 cervical cancer cell lines, HeLa, SiHa, CaSki, HeLa S3, C-33A and SNU-17. We also examined the association between the TERRA level and its stability and telomere length. We found that the TERRA level was several fold greater in the SiHa, CaSki, HeLa S3, C-33A and SNU-17 cells, than in the HeLa cells. An RNA stability assay of actinomycin D-treated cells revealed that TERRA had a short half-life of ~4 h in HeLa cells, which was consistent with previous studies, but was more stable with a longer half-life (>8 h) in the other 5 cell lines. Telomere length varied from 4 to 9 kb in the cells and did not correlate significantly with the TERRA level. On the whole, our data indicate that TERRA abundance and stability vary between different types of cervical cancer cells. TERRA degrades rapidly in HeLa cells, but is maintained stably in other cervical cancer cells that accumulate higher levels of TERRA. TERRA abundance is associated with the stability of RNA in cervical cancer cells, but is unlikely associated with telomere length.

Identifying the transition to the maturation zone in three ecotypes of Arabidopsis thaliana roots.

PubMed

Cajero Sánchez, Wendy; García-Ponce, Berenice; Sánchez, María de la Paz; Álvarez-Buylla, Elena R; Garay-Arroyo, Adriana

2018-01-01

The Arabidopsis thaliana (hereafter Arabidopsis) root has become a useful model for studying how organ morphogenesis emerge from the coordination and balance of cell proliferation and differentiation, as both processes may be observed and quantified in the root at different stages of development. Hence, being able to objectively identify and delimit the different stages of root development has been very important. Up to now, three different zones along the longitudinal axis of the primary root of Arabidopsis, have been identified: the root apical meristematic zone (RAM) with two domains [the proliferative (PD) and the transition domain (TD)], the elongation zone (EZ) and the maturation zone (MZ). We previously reported a method to quantify the length of the cells of the meristematic and the elongation zone, as well as the boundaries or transitions between the root domains along the growing part of the Arabidopsis root. In this study, we provide a more accurate criterion to identify the MZ. Traditionally, the transition between the EZ to the MZ has been established by the emergence of the first root-hair bulge in the epidermis, because this emergence coincides with cell maturation in this cell type. But we have found here that after the emergence of the first root-hair bulge some cells continue to elongate and we have confirmed this in three different Arabidopsis ecotypes. We established the limit between the EZ and the MZ by looking for the closest cortical cell with a longer length than the average cell length of 10 cells after the cortical cell closest to the epidermal cell with the first root-hair bulge in these three ecotypes. In Col-0 and Ws this cell is four cells above the one with the root hair bulge and, in the Ler ecotype, this cell is five cells above. To unambiguously identifying the site at which cells stop elongating and attain their final length and fate at the MZ, we propose to calculate the length of completely elongated cortical cells counting 10 cells starting from the sixth cell above the cortical cell closest to the epidermal cell with the first root-hair bulge. We validated this proposal in the three ecotypes analyzed and consider that this proposal may aid at having a more objective way to characterize root phenotypes and compare among them.

Identifying the transition to the maturation zone in three ecotypes of Arabidopsis thaliana roots

PubMed Central

Cajero Sánchez, Wendy; García-Ponce, Berenice; Sánchez, María de la Paz; Álvarez-Buylla, Elena R.; Garay-Arroyo, Adriana

2018-01-01

ABSTRACT The Arabidopsis thaliana (hereafter Arabidopsis) root has become a useful model for studying how organ morphogenesis emerge from the coordination and balance of cell proliferation and differentiation, as both processes may be observed and quantified in the root at different stages of development. Hence, being able to objectively identify and delimit the different stages of root development has been very important. Up to now, three different zones along the longitudinal axis of the primary root of Arabidopsis, have been identified: the root apical meristematic zone (RAM) with two domains [the proliferative (PD) and the transition domain (TD)], the elongation zone (EZ) and the maturation zone (MZ). We previously reported a method to quantify the length of the cells of the meristematic and the elongation zone, as well as the boundaries or transitions between the root domains along the growing part of the Arabidopsis root. In this study, we provide a more accurate criterion to identify the MZ. Traditionally, the transition between the EZ to the MZ has been established by the emergence of the first root-hair bulge in the epidermis, because this emergence coincides with cell maturation in this cell type. But we have found here that after the emergence of the first root-hair bulge some cells continue to elongate and we have confirmed this in three different Arabidopsis ecotypes. We established the limit between the EZ and the MZ by looking for the closest cortical cell with a longer length than the average cell length of 10 cells after the cortical cell closest to the epidermal cell with the first root-hair bulge in these three ecotypes. In Col-0 and Ws this cell is four cells above the one with the root hair bulge and, in the Ler ecotype, this cell is five cells above. To unambiguously identifying the site at which cells stop elongating and attain their final length and fate at the MZ, we propose to calculate the length of completely elongated cortical cells counting 10 cells starting from the sixth cell above the cortical cell closest to the epidermal cell with the first root-hair bulge. We validated this proposal in the three ecotypes analyzed and consider that this proposal may aid at having a more objective way to characterize root phenotypes and compare among them. PMID:29497470

Telomere shortening associated with chromosome instability is arrested in immortal cells which express telomerase activity.

PubMed Central

Counter, C M; Avilion, A A; LeFeuvre, C E; Stewart, N G; Greider, C W; Harley, C B; Bacchetti, S

1992-01-01

Loss of telomeric DNA during cell proliferation may play a role in ageing and cancer. Since telomeres permit complete replication of eukaryotic chromosomes and protect their ends from recombination, we have measured telomere length, telomerase activity and chromosome rearrangements in human cells before and after transformation with SV40 or Ad5. In all mortal populations, telomeres shortened by approximately 65 bp/generation during the lifespan of the cultures. When transformed cells reached crisis, the length of the telomeric TTAGGG repeats was only approximately 1.5 kbp and many dicentric chromosomes were observed. In immortal cells, telomere length and frequency of dicentric chromosomes stabilized after crisis. Telomerase activity was not detectable in control or extended lifespan populations but was present in immortal populations. These results suggest that chromosomes with short (TTAGGG)n tracts are recombinogenic, critically shortened telomeres may be incompatible with cell proliferation and stabilization of telomere length by telomerase may be required for immortalization. Images PMID:1582420

Cell elongation is an adaptive response for clearing long chromatid arms from the cleavage plane

PubMed Central

Kotadia, Shaila; Montembault, Emilie; Sullivan, William

2012-01-01

Chromosome segregation must be coordinated with cell cleavage to ensure correct transmission of the genome to daughter cells. Here we identify a novel mechanism by which Drosophila melanogaster neuronal stem cells coordinate sister chromatid segregation with cleavage furrow ingression. Cells adapted to a dramatic increase in chromatid arm length by transiently elongating during anaphase/telophase. The degree of cell elongation correlated with the length of the trailing chromatid arms and was concomitant with a slight increase in spindle length and an enlargement of the zone of cortical myosin distribution. Rho guanine-nucleotide exchange factor (Pebble)–depleted cells failed to elongate during segregation of long chromatids. As a result, Pebble-depleted adult flies exhibited morphological defects likely caused by cell death during development. These studies reveal a novel pathway linking trailing chromatid arms and cortical myosin that ensures the clearance of chromatids from the cleavage plane at the appropriate time during cytokinesis, thus preserving genome integrity. PMID:23185030

Diffusion lengths in irradiated N/P InP-on-Si solar cells

NASA Technical Reports Server (NTRS)

Wojtczuk, Steven; Colerico, Claudia; Summers, Geoffrey P.; Walters, Robert J.; Burke, Edward A.

1995-01-01

Indium phosphide (InP) solar cells are being made on silicon (Si) wafers (InP/Si) to take advantage of both the radiation-hardness properties of the InP solar cell and the light weight and low cost of Si wafers compared to InP or germanium (Ge) wafers. The InP/Si cell application is for long duration and/or high radiation orbit space missions. InP/Si cells have higher absolute efficiency after a high radiation dose than gallium arsenide (GaAs) or silicon (Si) solar cells. In this work, base electron diffusion lengths in the N/P cell are extracted from measured AM0 short-circuit photocurrent at various irradiation levels out to an equivalent 1 MeV fluence of 1017 1 MeV electrons/sq cm for a 1 sq cm 12% BOL InP/Si cell. These values are then checked for consistency by comparing measured Voc data with a theoretical Voc model that includes a dark current term that depends on the extracted diffusion lengths.

Numerical prediction of algae cell mixing feature in raceway ponds using particle tracing methods.

PubMed

Ali, Haider; Cheema, Taqi A; Yoon, Ho-Sung; Do, Younghae; Park, Cheol W

2015-02-01

In the present study, a novel technique, which involves numerical computation of the mixing length of algae particles in raceway ponds, was used to evaluate the mixing process. A value of mixing length that is higher than the maximum streamwise distance (MSD) of algae cells indicates that the cells experienced an adequate turbulent mixing in the pond. A coupling methodology was adapted to map the pulsating effects of a 2D paddle wheel on a 3D raceway pond in this study. The turbulent mixing was examined based on the computations of mixing length, residence time, and algae cell distribution in the pond. The results revealed that the use of particle tracing methodology is an improved approach to define the mixing phenomenon more effectively. Moreover, the algae cell distribution aided in identifying the degree of mixing in terms of mixing length and residence time. © 2014 Wiley Periodicals, Inc.

Damage and recovery characteristics of lithium-containing solar cells.

NASA Technical Reports Server (NTRS)

Faith, T. J.

1971-01-01

Damage and recovery characteristics were measured on lithium-containing solar cells irradiated by 1-MeV electrons. Empirical expressions for cell recovery time, diffusion-length damage coefficient immediately after irradiation, and diffusion-length damage coefficient after recovery were derived using results of short-circuit current, diffusion-length, and reverse-bias capacitance measurements. The damage coefficients were expressed in terms of a single lithium density parameter, the lithium gradient. A fluence dependence was also established, this dependence being the same for both the immediate-post-irradiation and post-recovery cases. Cell recovery rates were found to increase linearly with lithium gradient.

Engineering Cyanobacterial Cell Morphology for Enhanced Recovery and Processing of Biomass.

PubMed

Jordan, Adam; Chandler, Jenna; MacCready, Joshua S; Huang, Jingcheng; Osteryoung, Katherine W; Ducat, Daniel C

2017-05-01

Cyanobacteria are emerging as alternative crop species for the production of fuels, chemicals, and biomass. Yet, the success of these microbes depends on the development of cost-effective technologies that permit scaled cultivation and cell harvesting. Here, we investigate the feasibility of engineering cell morphology to improve biomass recovery and decrease energetic costs associated with lysing cyanobacterial cells. Specifically, we modify the levels of Min system proteins in Synechococcus elongatus PCC 7942. The Min system has established functions in controlling cell division by regulating the assembly of FtsZ, a tubulin-like protein required for defining the bacterial division plane. We show that altering the expression of two FtsZ-regulatory proteins, MinC and Cdv3, enables control over cell morphology by disrupting FtsZ localization and cell division without preventing continued cell growth. By varying the expression of these proteins, we can tune the lengths of cyanobacterial cells across a broad dynamic range, anywhere from an ∼20% increased length (relative to the wild type) to near-millimeter lengths. Highly elongated cells exhibit increased rates of sedimentation under low centrifugal forces or by gravity-assisted settling. Furthermore, hyperelongated cells are also more susceptible to lysis through the application of mild physical stress. Collectively, these results demonstrate a novel approach toward decreasing harvesting and processing costs associated with mass cyanobacterial cultivation by altering morphology at the cellular level. IMPORTANCE We show that the cell length of a model cyanobacterial species can be programmed by rationally manipulating the expression of protein factors that suppress cell division. In some instances, we can increase the size of these cells to near-millimeter lengths with this approach. The resulting elongated cells have favorable properties with regard to cell harvesting and lysis. Furthermore, cells treated in this manner continue to grow rapidly at time scales similar to those of uninduced controls. To our knowledge, this is the first reported example of engineering the cell morphology of cyanobacteria or algae to make them more compatible with downstream processing steps that present economic barriers to their use as alternative crop species. Therefore, our results are a promising proof-of-principle for the use of morphology engineering to increase the cost-effectiveness of the mass cultivation of cyanobacteria for various sustainability initiatives. Copyright © 2017 American Society for Microbiology.

The effects of early hypo- and hyperthyroidism on the development of rat cerebellar cortex. III. Kinetics of cell proliferation in the external granular layer.

PubMed

Lauder, J M

1977-04-22

The effects of early hypo- and hyperthyroidism on the rates of cell acquisition and proliferation have been studied in the external granular layer (EGL) of the developing rat cerebellar cortex at 10 days of age using quantitative autoradiographic methods. Both altered thyroid states reduce the rate of cell acquisition in the EGL, but appear to do so for different reasons. Hyperthyroidism shortens the average length of the cell cycle by decreasing the duration of the pre-DNA synthetic phase (G1), indicating that excess thyroxine may exert a direct effect on the EGL. This action involves the early onset of neuronal differentiation (cessation of proliferation)46 which presumably leads to the observed decrease in the rate of cell acquisition (increased doubling time). Such differentiating cells do not, however, leave the proliferative zone or the EGL prematurely, resulting in a reduced labeling index, mitotic index, and growth fraction as non-dividing cells dilute the proliferating cell population. Hypothyroidism, on the other hand, leads to no significant change in the length of the cell cycle or in the mitotic index, but causes a decreased labeling index and growth fraction, as well as a reduced rate of cell acquisition (increased doubling time). No significant change in the amount of cell death in the EGL could be found to explain this apparent discrepancy between the rate of cell proliferation (cell cycle length) and cell acqusiition. The answer to this puzzle appears to lie in the mitotic index, which is not affected to the same extent as the labeling index, although it is also slightly reduced. If cells were to remain longer in mitosis, this could result in a decreased labeling index and growth fraction but nearly normal mitotic index and cell cycle length (as measured using the % labeled mitoses method), since those cells dropping out of the cycling population would be counted as mitoses...

Testing the time-of-flight model for flagellar length sensing.

PubMed

Ishikawa, Hiroaki; Marshall, Wallace F

2017-11-07

Cilia and flagella are microtubule-based organelles that protrude from the surface of most cells, are important to the sensing of extracellular signals, and make a driving force for fluid flow. Maintenance of flagellar length requires an active transport process known as intraflagellar transport (IFT). Recent studies reveal that the amount of IFT injection negatively correlates with the length of flagella. These observations suggest that a length-dependent feedback regulates IFT. However, it is unknown how cells recognize the length of flagella and control IFT. Several theoretical models try to explain this feedback system. We focused on one of the models, the "time-of-flight" model, which measures the length of flagella on the basis of the travel time of IFT protein in the flagellar compartment. We tested the time-of-flight model using Chlamydomonas dynein mutant cells, which show slower retrograde transport speed. The amount of IFT injection in dynein mutant cells was higher than that in control cells. This observation does not support the prediction of the time-of-flight model and suggests that Chlamydomonas uses another length-control feedback system rather than that described by the time-of-flight model. © 2017 Ishikawa and Marshall. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Dynamics of cell proliferation in the adult dentate gyrus of two inbred strains of mice

NASA Technical Reports Server (NTRS)

Hayes, N. L.; Nowakowski, R. S.

2002-01-01

The output potential of proliferating populations in either the developing or the adult nervous system is critically dependent on the length of the cell cycle (T(c)) and the size of the proliferating population. We developed a new approach for analyzing the cell cycle, the 'Saturate and Survive Method' (SSM), that also reveals the dynamic behaviors in the proliferative population and estimates of the size of the proliferating population. We used this method to analyze the proliferating population of the adult dentate gyrus in 60 day old mice of two inbred strains, C57BL/6J and BALB/cByJ. The results show that the number of cells labeled by exposure to BUdR changes dramatically with time as a function of the number of proliferating cells in the population, the length of the S-phase, cell division, the length of the cell cycle, dilution of the S-phase label, and cell death. The major difference between C57BL/6J and BALB/cByJ mice is the size of the proliferating population, which differs by a factor of two; the lengths of the cell cycle and the S-phase and the probability that a newly produced cell will die within the first 10 days do not differ in these two strains. This indicates that genetic regulation of the size of the proliferating population is independent of the genetic regulation of cell death among those newly produced cells. The dynamic changes in the number of labeled cells as revealed by the SSM protocol also indicate that neither single nor repeated daily injections of BUdR accurately measure 'proliferation.'.

HIV-DNA content in different CD4+ T-cell subsets correlates with CD4+ cell :  CD8+ cell ratio or length of efficient treatment.

PubMed

Gibellini, Lara; Pecorini, Simone; De Biasi, Sara; Bianchini, Elena; Digaetano, Margherita; Pinti, Marcello; Carnevale, Gianluca; Borghi, Vanni; Guaraldi, Giovanni; Mussini, Cristina; Cossarizza, Andrea; Nasi, Milena

2017-06-19

HIV establishes a latent infection at different degrees within naïve (TN) or central (TCM) and effector memory (TEM) CD4 T cell. Studying patients in whom HIV production was suppressed by combined antiretroviral therapy, our main aim was to find which factors are related or can influence intracellular viral reservoir in different CD4 T-cell subsets. We enrolled 32 HIV patients successfully treated for more than 2 years, with a CD4 T-cell count more than 500 cells/μl and plasma viremia undetectable from at least 1 year. Proviral HIV-DNA, the amount of cells expressing signal-joint T-cell receptor rearrangement excision circles and telomere length were quantified by droplet digital PCR in highly purified, sorted CD4 T-cell subsets; plasma IL-7 and IL-15 were measured by ELISA. HIV-DNA was significantly lower in TN cells compared with TCM or to TEM. Conversely, TN cells contained more signal-joint T-cell receptor rearrangement excision circles compared with TCM or to TEM; no appreciable changes were observed in telomere length. HIV-DNA content was significantly higher in TN and TCM cells, but not in TEM, from patients with shorter time of treatment, or in those with lower CD4 : CD8 ratio. Length of treatment or recovery of CD4 : CD8 ratio significantly influences viral reservoir in both TN and TCM. Measuring HIV-DNA in purified lymphocyte populations allows a better monitoring of HIV reservoir and could be useful for designing future eradication strategies.

Timing of Tissue-specific Cell Division Requires a Differential Onset of Zygotic Transcription during Metazoan Embryogenesis*

PubMed Central

Wong, Ming-Kin; Guan, Daogang; Ng, Kaoru Hon Chun; Ho, Vincy Wing Sze; An, Xiaomeng; Li, Runsheng; Ren, Xiaoliang

2016-01-01

Metazoan development demands not only precise cell fate differentiation but also accurate timing of cell division to ensure proper development. How cell divisions are temporally coordinated during development is poorly understood. Caenorhabditis elegans embryogenesis provides an excellent opportunity to study this coordination due to its invariant development and widespread division asynchronies. One of the most pronounced asynchronies is a significant delay of cell division in two endoderm progenitor cells, Ea and Ep, hereafter referred to as E2, relative to its cousins that mainly develop into mesoderm organs and tissues. To unravel the genetic control over the endoderm-specific E2 division timing, a total of 822 essential and conserved genes were knocked down using RNAi followed by quantification of cell cycle lengths using in toto imaging of C. elegans embryogenesis and automated lineage. Intriguingly, knockdown of numerous genes encoding the components of general transcription pathway or its regulatory factors leads to a significant reduction in the E2 cell cycle length but an increase in cell cycle length of the remaining cells, indicating a differential requirement of transcription for division timing between the two. Analysis of lineage-specific RNA-seq data demonstrates an earlier onset of transcription in endoderm than in other germ layers, the timing of which coincides with the birth of E2, supporting the notion that the endoderm-specific delay in E2 division timing demands robust zygotic transcription. The reduction in E2 cell cycle length is frequently associated with cell migration defect and gastrulation failure. The results suggest that a tissue-specific transcriptional activation is required to coordinate fate differentiation, division timing, and cell migration to ensure proper development. PMID:27056332

A numerical simulation of finite-length Taylor-Couette flow

NASA Technical Reports Server (NTRS)

Streett, C. L.; Hussaini, M. Y.

1988-01-01

Results from numerical simulations of finite-length Taylor-Couette flow are presented. Included are time-accurate and steady-state studies of the change in the nature of the symmetric two-cell/asymmetric one-cell bifurcation with varying aspect ratio and of the Reynolds number/aspect ratio locus of the two-cell/four-cell bifurcation. Preliminary results from wavy-vortex simulations at low aspect ratios are also presented.

LINDENS: A program for lineament length and density analysis*1

NASA Astrophysics Data System (ADS)

Casas, Antonio M.; Cortés, Angel L.; Maestro, Adolfo; Soriano, M. Asunción; Riaguas, Andres; Bernal, Javier

2000-11-01

Analysis of lineaments from satellite images normally includes the determination of their orientation and density. The spatial variation in the orientation and/or number of lineaments must be obtained by means of a network of cells, the lineaments included in each cell being analysed separately. The program presented in this work, LINDENS, allows the density of lineaments (number of lineaments per km 2 and length of lineaments per km 2) to be estimated. It also provides a tool for classifying the lineaments contained in different cells, so that their orientation can be represented in frequency histograms and/or rose diagrams. The input file must contain the planar coordinates of the beginning and end of each lineament. The density analysis is done by creating a network of square cells, and counting the number of lineaments that are contained within each cell, that have one of their ends within the cell or that cross-cut the cell boundary. The lengths of lineaments are then calculated. To obtain a representative density map the cell size must be fixed according to: (1) the average lineament length; (2) the distance between the lineaments; and (3) the boundaries of zones with low densities due to lithology or outcrop features. An example from the Neogene Duero Basin (Northern Spain) is provided to test the reliability of the density maps obtained with different cell sizes.

Recirculation cells for granular flow in cylindrical rotating tumblers

NASA Astrophysics Data System (ADS)

D'Ortona, Umberto; Thomas, Nathalie; Lueptow, Richard M.

2018-05-01

To better understand the velocity field and flowing layer structure, we have performed a detailed discrete element method study of the flow of monodisperse particles in a partially filled three-dimensional cylindrical rotating tumblers. Similar to what occurs near the poles in spherical and conical tumblers, recirculation cells (secondary flows) develop near the flat endwalls of a cylindrical tumbler in which particles near the surface drift axially toward the endwall, while particles deeper in the flowing layer drift axially toward the midlength of the tumbler. Another recirculation cell with the opposite sense develops next to each endwall recirculation cell, extending to the midlength of the tumbler. For a long enough tumbler, each endwall cell is about one quarter of the tumbler diameter in length. Endwall cells are insensitive to tumbler length and relatively insensitive to rotation speed (so long as the flowing layer remains flat and continuously flowing) or fill level (from 25% to 50% full). However, for shorter tumblers (0.5 to 1.0 length/diameter aspect ratio) the endwall cell size does not change much, while center cells reduce their size and eventually disappear for the shortest tumblers. For longer tumblers (length/diameter aspect ratio larger than 2), a stagnation zone appears in between the central cells. These results provide insight into the mixing of monodisperse particles in rotating cylindrical tumblers as well as the frictional effects of the tumbler endwalls.

Identifying Developmental Zones in Maize Lateral Root Cell Length Profiles using Multiple Change-Point Models

PubMed Central

Moreno-Ortega, Beatriz; Fort, Guillaume; Muller, Bertrand; Guédon, Yann

2017-01-01

The identification of the limits between the cell division, elongation and mature zones in the root apex is still a matter of controversy when methods based on cellular features, molecular markers or kinematics are compared while methods based on cell length profiles have been comparatively underexplored. Segmentation models were developed to identify developmental zones within a root apex on the basis of epidermal cell length profiles. Heteroscedastic piecewise linear models were estimated for maize lateral roots of various lengths of both wild type and two mutants affected in auxin signaling (rtcs and rum-1). The outputs of these individual root analyses combined with morphological features (first root hair position and root diameter) were then globally analyzed using principal component analysis. Three zones corresponding to the division zone, the elongation zone and the mature zone were identified in most lateral roots while division zone and sometimes elongation zone were missing in arrested roots. Our results are consistent with an auxin-dependent coordination between cell flux, cell elongation and cell differentiation. The proposed segmentation models could extend our knowledge of developmental regulations in longitudinally organized plant organs such as roots, monocot leaves or internodes. PMID:29123533

Diffusion lengths of silicon solar cells from luminescence images

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

Wuerfel, P.; Trupke, T.; Puzzer, T.

A method for spatially resolved measurement of the minority carrier diffusion length in silicon wafers and in silicon solar cells is introduced. The method, which is based on measuring the ratio of two luminescence images taken with two different spectral filters, is applicable, in principle, to both photoluminescence and electroluminescence measurements and is demonstrated experimentally by electroluminescence measurements on a multicrystalline silicon solar cell. Good agreement is observed with the diffusion length distribution obtained from a spectrally resolved light beam induced current map. In contrast to the determination of diffusion lengths from one single luminescence image, the method proposed heremore » gives absolute values of the diffusion length and, in comparison, it is much less sensitive to lateral voltage variations across the cell area as caused by local variations of the series resistance. It is also shown that measuring the ratio of two luminescence images allows distinguishing shunts or surface defects from bulk defects.« less

Recovery of infectious classical swine fever virus (CSFV) from full-length genomic cDNA clones by a swine kidney cell line expressing bacteriophage T7 RNA polymerase.

PubMed

van Gennip, H G; van Rijn, P A; Widjojoatmodjo, M N; Moormann, R J

1999-03-01

A new method for the recovery of infectious classical swine fever virus (CSFV) from full-length genomic cDNA clones of the C-strain was developed. Classical reverse genetics is based on transfection of in vitro transcribed RNA to target cells to recover RNA viruses. However, the specific infectivity of such in vitro transcribed RNA in swine kidney cells is usually low. To improve reverse genetics for CSFV, a stable swine kidney cell line was established that expresses cytoplasmic bacteriophage T7 RNA polymerase (SK6.T7). A 200-fold increased virus titre was obtained from SK6.T7 cells transfected with linearized full-length cDNA compared to in vitro transcribed RNA, whereas transfection of circular full-length cDNA resulted in 20-fold increased virus titres. Viruses generated on the SK6.T7 cells are indistinguishable from the viruses generated by the classical reverse genetic procedures. These results show the improved recovery of infectious CSFV directly from full-length cDNAs. Furthermore, the reverse genetic procedures are simplified to a faster, one step protocol. We conclude that the SK6.T7 cell line will be a valuable tool for recovering mutant CSFV and will contribute to future pestivirus research.

Immortal, telomerase-negative cell lines derived from a Li-Fraumeni syndrome patient exhibit telomere length variability and chromosomal and minisatellite instabilities.

PubMed

Tsutsui, Takeki; Kumakura, Shin-Ichi; Tamura, Yukiko; Tsutsui, Takeo W; Sekiguchi, Mizuki; Higuchi, Tokihiro; Barrett, J Carl

2003-05-01

Five immortal cell lines derived from a Li-Fraumeni syndrome patient (MDAH 087) with a germline mutant p53 allele were characterized with respect to telomere length and genomic instability. The remaining wild-type p53 allele is lost in the cell lines. Telomerase activity was undetectable in all immortal cell lines. Five subclones of each cell line and five re-subclones of each of the subclones also showed undetectable telomerase activity. All five immortal cell lines exhibited variability in the mean length of terminal restriction fragments (TRFs). Subclones of each cell line, and re-subclones of the subclones also showed TRF variability, indicating that the variability is owing to clonal heterogeneity. Chromosome aberrations were observed at high frequencies in these cell lines including the subclones and re-subclones, and the principal types of aberrations were breaks, double minute chromosomes and dicentric chromosomes. In addition, minisatellite instability detected by DNA fingerprints was observed in the immortal cell lines. However, all of the cell lines were negative for microsatellite instability. As minisatellite sequences are considered recombinogenic in mammalian cells, these results suggest that recombination rates can be increased in these cell lines. Tumor-derived human cell lines, HT1080 cells and HeLa cells that also lack p53 function, exhibited little genomic instability involving chromosomal and minisatellite instabilities, indicating that chromosomal and minisatellite instabilities observed in the immortal cell lines lacking telomerase activity could not result from loss of p53 function.

Glycoprotein 5 of porcine reproductive and respiratory syndrome virus strain SD16 inhibits viral replication and causes G2/M cell cycle arrest, but does not induce cellular apoptosis in Marc-145 cells

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

Mu, Yang, E-mail: muyang@nwsuaf.edu.cn; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, Ministry of Agriculture of the People's Republic of China, No. 22 Xinong Road, Yangling, Shaanxi 712100; Li, Liangliang, E-mail: lifeiyang2007@126.com

Cell apoptosis is common after infection with porcine reproductive and respiratory syndrome virus (PRRSV). PRRSV GP5 has been reported to induce cell apoptosis. To further understand the role of GP5 in PRRSV induced cell apoptosis, we established Marc-145 cell lines stably expressing full-length GP5, GP5{sup Δ84-96} (aa 84-96 deletion), and GP5{sup Δ97-119} (aa 97-119 deletion). Cell proliferation, cell cycle progression, cell apoptosis and virus replication in these cell lines were evaluated. Neither truncated nor full-length GP5 induced cell apoptosis in Marc-145 cells. However, GP5{sup Δ97-119}, but not full-length or GP5{sup Δ84-96}, induced a cell cycle arrest at the G2/M phasemore » resulting in a reduction in the growth of Marc-145 cells. Additionally, GP5{sup Δ84-96} inhibited the replication of PRRSV in Marc-145 cells through induction of IFN-β. These findings suggest that PRRSV GP5 is not responsible for inducing cell apoptosis in Marc-145 cells under these experimental conditions; however it has other important roles in virus/host cell biology. - Highlights: • Marc-145 cell lines stable expression PRRSV GP5 or truncated GP5 were constructed. • GP5{sup Δ97-119} expression in Marc-145 cell induced cell cycle arrest at G2/M phase. • Expression of GP5 and truncated GP5 could not induce Marc-145 cells apoptosis. • PRRSV replication in Marc-145-GP5{sup Δ84-96} was significantly inhibited.« less

Morphometric analysis of cisplatin-induced neurite outgrowth in N1E-115 neuroblastoma cells.

PubMed

Konings, P N; Philipsen, R L; van den Broek, J H; Ruigt, G S

1994-08-29

Cisplatin, a widely used cytostatic drug for the control of a variety of neoplastic tumors, unexpectedly induced neurite outgrowth in N1E-115 neuroblastoma cells and this phenomenon was studied further in detail with morphometric analysis. As expected, cisplatin dose-dependently reduced cell number. At the same time, however, cisplatin affected the morphology of the neuroblastoma cells that changed from small rounded cell bodies into large flat cell bodies with neurites. The neurite length/cell as a function of cisplatin concentration showed a bell-shaped curve. The maximal effect (1200% of control) on neurite length/cell was observed at 1 microgram/ml cisplatin. In conclusion, cisplatin induced cellular differentiation in N1E-115 neuroblastoma cells at and just above threshold doses for cytostatic activity.

Effects of shoot inversion on stem structure in Pharbitis nil

NASA Technical Reports Server (NTRS)

Prasad, T. K.; Sack, F. D.; Cline, M. G.

1988-01-01

The effects of shoot inversion on stem structure over 72 hr were investigated in Pharbitis nil by analyzing cell number, cell length, and the cross sectional areas of cells, tissues, and regions. An increase in stem diameter can be attributed to an increase in both cell number and cross sectional area of pith (primarily) and vascular tissue (secondarily). Qualitative observations of cell wall thickness in the light microscope did not reveal any significant effects of shoot inversion on this parameter. The inhibition of shoot elongation was accompanied by a significant decrease in cell length in the pith. The results are generally consistent with an ethylene effect on cell dimensions, especially in the pith.

Tired telomeres: Poor global sleep quality, perceived stress, and telomere length in immune cell subsets in obese men and women.

PubMed

Prather, Aric A; Gurfein, Blake; Moran, Patricia; Daubenmier, Jennifer; Acree, Michael; Bacchetti, Peter; Sinclair, Elizabeth; Lin, Jue; Blackburn, Elizabeth; Hecht, Frederick M; Epel, Elissa S

2015-07-01

Poor sleep quality and short sleep duration are associated with increased incidence and progression of a number of chronic health conditions observed at greater frequency among the obese and those experiencing high levels of stress. Accelerated cellular aging, as indexed by telomere attrition in immune cells, is a plausible pathway linking sleep and disease risk. Prior studies linking sleep and telomere length are mixed. One factor may be reliance on leukocytes, which are composed of varied immune cell types, as the sole measure of telomere length. To better clarify these associations, we investigated the relationships of global sleep quality, measured by the Pittsburgh Sleep Quality Index (PSQI), and diary-reported sleep duration with telomere length in different immune cell subsets, including granulocytes, peripheral blood mononuclear cells (PBMCs), CD8+ and CD4+ T lymphocytes, and B lymphocytes in a sample of 87 obese men and women (BMI mean=35.4, SD=3.6; 81.6% women; 62.8% Caucasian). Multiple linear regression analyses were performed adjusting for age, gender, race, education, BMI, sleep apnea risk, and perceived stress. Poorer PSQI global sleep quality was associated with statistically significantly shorter telomere length in lymphocytes but not granulocytes and in particular CD8+ T cells (b=-56.8 base pairs per one point increase in PSQI, SE=20.4, p=0.007) and CD4+ T cells (b=-37.2, SE=15.9, p=0.022). Among separate aspects of global sleep quality, low perceived sleep quality and decrements in daytime function were most related to shorter telomeres. In addition, perceived stress moderated the sleep-CD8+ telomere association. Poorer global sleep quality predicted shorter telomere length in CD8+ T cells among those with high perceived stress but not in low stress participants. These findings provide preliminary evidence that poorer global sleep quality is related to telomere length in several immune cell types, which may serve as a pathway linking sleep and disease risk in obese individuals. Copyright © 2014 Elsevier Inc. All rights reserved.

Maintenance of telomere length in AML.

PubMed

Lansdorp, Peter M

2017-11-28

The importance of telomere length to human health, aging, and cancer continues to be underappreciated. This review examines some basics of telomere biology and relates how telomere function, telomerase activity, and mutations in TERC or TERT are involved in bone marrow failure, leukemias, and other cancers. Given the challenge to obtain accurate data on telomerase activity and telomere length in specific cell types, the situation in acute myeloid leukemia (AML) remains puzzling. In most cancers, telomerase levels are increased after cells have encountered a "telomere crisis," which is typically associated with poor prognosis. Cells emerging from "telomere crisis" have defective DNA damage responses, resulting, for example, from loss of p53. Such cells often express elevated telomerase levels as a result of point mutations in the TERT promoter or amplification of the TERT gene. While telomeres in AML blasts are typically shorter than expected for normal leukocytes, most AML cells do not show evidence of having gone through a "telomere crisis." In chronic myeloid leukemia (CML), the difference between the telomere length in nonmalignant T cells and malignant blasts from the same patient was found to correlate with the remaining duration of the chronic phase. This observation supports that a mitotic clock is ticking in CML stem cells and that disease progression in CML heralds the onset of a "telomere crisis." The presence of very short telomeres in tumor cells was found to predict disease progression in chronic lymphocytic leukemia, myeloma, and various solid tumors. In view of these findings longitudinal studies of telomere length in AML appear worthwhile.

A pilot study showing associations between frequency of CD4(+) memory cell subsets at diagnosis and duration of partial remission in type 1 diabetes.

PubMed

Moya, Rosita; Robertson, Hannah Kathryn; Payne, Dawson; Narsale, Aditi; Koziol, Jim; Davies, Joanna Davida

2016-05-01

In some patients with type 1 diabetes the dose of insulin required to achieve euglycemia is substantially reduced soon after diagnosis. This partial remission is associated with β-cell function and good glucose control. The purpose of this study was to assess whether frequencies of CD4(+) T cell subsets in children newly diagnosed with type 1 diabetes are associated with length of partial remission. We found that the frequency of CD4(+) memory cells, activated Treg cells and CD25(+) cells that express a high density of the IL-7 receptor, CD127 (CD127(hi)) are strongly associated with length of partial remission. Prediction of length of remission via Cox regression is significantly enhanced when CD25(+) CD127(hi) cell frequency is combined with either Insulin Dependent Adjusted A1c (IDAA1c), or glycosylated hemoglobin (HbA1c), or C-peptide levels at diagnosis. CD25(+) CD127(hi) cells do not express Foxp3, LAG-3 and CD49b, indicating that they are neither Treg nor Tr1 cells. Copyright © 2016 Elsevier Inc. All rights reserved.

Realizing the Translational Potential of Telomere Length Variation as a Tissue-Based Prognostic Marker for Prostate Cancer

DTIC Science & Technology

2014-10-01

Telomere Length Variation as a Tissue- Based Prognostic Marker for Prostate Cancer PRINCIPAL INVESTIGATOR: Elizabeth A. Platz CONTRACTING...Translational Potential of Telomere Length Variation as a Tissue- Based Prognostic Marker for Prostate Cancer 5b. GRANT NUMBER W81XWH-12-1-0545 5c...combination of telomere length variability in prostate cancer cells and short telomere length in cancer-associated stromal cells is an independent

Differential segregation in a cell-cell contact interface: the dynamics of the immunological synapse.

PubMed Central

Burroughs, Nigel John; Wülfing, Christoph

2002-01-01

Receptor-ligand couples in the cell-cell contact interface between a T cell and an antigen-presenting cell form distinct geometric patterns and undergo spatial rearrangement within the contact interface. Spatial segregation of the antigen and adhesion receptors occurs within seconds of contact, central aggregation of the antigen receptor then occurring over 1-5 min. This structure, called the immunological synapse, is becoming a paradigm for localized signaling. However, the mechanisms driving its formation, in particular spatial segregation, are currently not understood. With a reaction diffusion model incorporating thermodynamics, elasticity, and reaction kinetics, we examine the hypothesis that differing bond lengths (extracellular domain size) is the driving force behind molecular segregation. We derive two key conditions necessary for segregation: a thermodynamic criterion on the effective bond elasticity and a requirement for the seeding/nucleation of domains. Domains have a minimum length scale and will only spontaneously coalesce/aggregate if the contact area is small or the membrane relaxation distance large. Otherwise, differential attachment of receptors to the cytoskeleton is required for central aggregation. Our analysis indicates that differential bond lengths have a significant effect on synapse dynamics, i.e., there is a significant contribution to the free energy of the interaction, suggesting that segregation by differential bond length is important in cell-cell contact interfaces and the immunological synapse. PMID:12324401

Fracture mechanics modeling of popping event during daughter cell separation.

PubMed

Jiang, Yuxuan; Liang, Xudong; Guo, Ming; Cao, Yanping; Cai, Shengqiang

2018-05-10

Most bacteria cells divide by binary fission which is part of a bacteria cell cycle and requires tight regulations and precise coordination. Fast separation of Staphylococcus Aureus (S. Aureus) daughter cells, named as popping event, has been observed in recent experiments. The popping event was proposed to be driven by mechanical crack propagation in the peripheral ring which connected two daughter cells before their separation. It has also been shown that after the fast separation, a small portion of the peripheral ring was left as a hinge. In the article, we develop a fracture mechanics model for the crack growth in the peripheral ring during S. Aureus daughter cell separation. In particular, using finite element analysis, we calculate the energy release rate associated with the crack growth in the peripheral ring, when daughter cells are inflated by a uniform turgor pressure inside. Our results show that with a fixed inflation of daughter cells, the energy release rate depends on the crack length non-monotonically. The energy release rate reaches a maximum value for a crack of an intermediate length. The non-monotonic relationship between the energy release rate and crack length clearly indicates that the crack propagation in the peripheral ring can be unstable. The computed energy release rate as a function of crack length can also be used to explain the existence of a small portion of peripheral ring remained as hinge after the popping event.

Dense-body aggregates as plastic structures supporting tension in smooth muscle cells.

PubMed

Zhang, Jie; Herrera, Ana M; Paré, Peter D; Seow, Chun Y

2010-11-01

The wall of hollow organs of vertebrates is a unique structure able to generate active tension and maintain a nearly constant passive stiffness over a large volume range. These properties are predominantly attributable to the smooth muscle cells that line the organ wall. Although smooth muscle is known to possess plasticity (i.e., the ability to adapt to large changes in cell length through structural remodeling of contractile apparatus and cytoskeleton), the detailed structural basis for the plasticity is largely unknown. Dense bodies, one of the most prominent structures in smooth muscle cells, have been regarded as the anchoring sites for actin filaments, similar to the Z-disks in striated muscle. Here, we show that the dense bodies and intermediate filaments formed cable-like structures inside airway smooth muscle cells and were able to adjust the cable length according to cell length and tension. Stretching the muscle cell bundle in the relaxed state caused the cables to straighten, indicating that these intracellular structures were connected to the extracellular matrix and could support passive tension. These plastic structures may be responsible for the ability of smooth muscle to maintain a nearly constant tensile stiffness over a large length range. The finding suggests that the structural plasticity of hollow organs may originate from the dense-body cables within the smooth muscle cells.

Quantification of cell cycle kinetics by EdU (5-ethynyl-2′-deoxyuridine)-coupled-fluorescence-intensity analysis

PubMed Central

Cabrita, Marisa; Bekman, Evguenia; Braga, José; Rino, José; Santus, Renè; Filipe, Paulo L.; Sousa, Ana E.; Ferreira, João A.

2017-01-01

We propose a novel single-deoxynucleoside-based assay that is easy to perform and provides accurate values for the absolute length (in units of time) of each of the cell cycle stages (G1, S and G2/M). This flow-cytometric assay takes advantage of the excellent stoichiometric properties of azide-fluorochrome detection of DNA substituted with 5-ethynyl-2′-deoxyuridine (EdU). We show that by pulsing cells with EdU for incremental periods of time maximal EdU-coupled fluorescence is reached when pulsing times match the length of S phase. These pulsing times, allowing labelling for a full S phase of a fraction of cells in asynchronous populations, provide accurate values for the absolute length of S phase. We characterized additional, lower intensity signals that allowed quantification of the absolute durations of G1 and G2 phases. Importantly, using this novel assay data on the lengths of G1, S and G2/M phases are obtained in parallel. Therefore, these parameters can be estimated within a time frame that is shorter than a full cell cycle. This method, which we designate as EdU-Coupled Fluorescence Intensity (E-CFI) analysis, was successfully applied to cell types with distinctive cell cycle features and shows excellent agreement with established methodologies for analysis of cell cycle kinetics. PMID:28465489

The role of telomeres and telomerase in hematologic malignancies and hematopoietic stem cell transplantation

PubMed Central

2014-01-01

Telomeres are specific nucleoprotein structures at the ends of eukaryotic chromosomes. Telomeres and telomere-associated proteins maintain genome stability by protecting the ends of chromosomes from fusion and degradation. In normal somatic cells, the length of the telomeres gradually becomes shortened with cell division. In tumor cells, the shortening of telomeres length is accelerated under the increased proliferation pressure. However, it will be maintained at an extremely short length as the result of activation of telomerase. Significantly shortened telomeres, activation of telomerase, and altered expression of telomere-associated proteins are common features of various hematologic malignancies and are related with progression or chemotherapy resistance in these diseases. In patients who have received hematopoietic stem cell transplantation (HSCT), the telomere length and the telomerase activity of the engrafted donor cells have a significant influence on HSCT outcomes. Transplantation-related factors should be taken into consideration because of their impacts on telomere homeostasis. As activation of telomerase is widespread in tumor cells, it has been employed as a target point in the treatment of neoplastic hematologic disorders. In this review, the characteristics and roles of telomeres and telomerase both in hematologic malignancies and in HSCT will be summarized. The current status of telomerase-targeted therapies utilized in the treatment of hematologic malignancies will also be reviewed. PMID:25139287

The effect of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow‑derived mesenchymal stem cells.

PubMed

Javanmard, F; Azadbakht, M; Pourmoradi, M

2016-01-01

In this study, the role of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow mesenchymal stem cells were investigated. The cells were cultured in treatment medium containing 100 nM of staurosporine for 4 hours; then the cells were affected by hydrostatic pressure (0, 25,50, 100 mmHg). The percentage of cell viability by trypan blue staining and the percentage of cell death by Hoechst/PI differential staining were assessed. We obtained the total neurite length. Expression of β-tubulin III and GFAP (Glial fibrillary acidic protein) proteins were also analyzed by immunocytochemistry. The percentage of cell viability in treatments decreased relative to the increase in hydrostatic pressure and time (p Keywords: bone marrow mesenchymal stem cell, hydrostatic pressure, immunocytochemistry, neural differentiation, neurite length, cell differentiation.

Development of high-efficiency solar cells on silicon web

NASA Technical Reports Server (NTRS)

Meier, D. L.; Greggi, J.; Rai-Choudhury, P.

1986-01-01

Work is reported aimed at identifying and reducing sources of carrier recombination both in the starting web silicon material and in the processed cells. Cross-sectional transmission electron microscopy measurements of several web cells were made and analyzed. The effect of the heavily twinned region on cell efficiency was modeled, and the modeling results compared to measured values for processed cells. The effects of low energy, high dose hydrogen ion implantation on cell efficiency and diffusion length were examined. Cells were fabricated from web silicon known to have a high diffusion length, with a new double layer antireflection coating being applied to these cells. A new contact system, to be used with oxide passivated cells and which greatly reduces the area of contact between metal and silicon, was designed. The application of DLTS measurements to beveled samples was further investigated.

Accumulation of PHA granules in Cupriavidus necator as seen by confocal fluorescence microscopy.

PubMed

Mravec, Filip; Obruca, Stanislav; Krzyzanek, Vladislav; Sedlacek, Petr; Hrubanova, Kamila; Samek, Ota; Kucera, Dan; Benesova, Pavla; Nebesarova, Jana

2016-05-01

Many bacteria are capable of accumulating intracellular granules of polyhydroxyalkanoates (PHA). In this work, we developed confocal microscopy analysis of bacterial cells to study changes in the diameters of cells as well as PHA granules during growth and PHA accumulation in the bacterium Cupriavidus necator H16 (formerly Ralstonia eutropha). The cell envelope was stained by DiD(®) fluorescent probe and PHA granules by Nile Red. Signals from both probes were separated based on their spectral and fluorescence life-time properties. During growth and PHA accumulation, bacterial cells increased their length but the width of the cells remained constant. The volume fraction of PHA granules in cells increased during PHA accumulation, nevertheless, its value did not exceed 40 vol. % regardless of the PHA weight content. It seems that bacterial cultures lengthen the cells in order to control the PHA volume portion. However, since similar changes in cell length were also observed in a PHA non-accumulating mutant, it seems that there is no direct control mechanism, which regulates the prolongation of the cells with respect to PHA granules volume. It is more likely that PHA biosynthesis and the length of cells are influenced by the same external stimuli such as nutrient limitation. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Multi-walled carbon nanotube length as a critical determinant of bioreactivity with primary human pulmonary alveolar cells

PubMed Central

Sweeney, Sinbad; Berhanu, Deborah; Misra, Superb K.; Thorley, Andrew J.; Valsami-Jones, Eugenia; Tetley, Teresa D.

2015-01-01

Multiwalled carbon nanotube (MWCNT) length is suggested to critically determine their pulmonary toxicity. This stems from in vitro and in vivo rodent studies and in vitro human studies using cell lines (typically cancerous). There is little data using primary human lung cells. We addressed this knowledge gap, using highly relevant, primary human alveolar cell models exposed to precisely synthesized and thoroughly characterized MWCNTs. In this work, transformed human alveolar type-I-like epithelial cells (TT1), primary human alveolar type-II epithelial cells (ATII) and alveolar macrophages (AM) were treated with increasing concentrations of MWCNTs before measuring cytotoxicity, inflammatory mediator release and MAP kinase signalling. Strikingly, we observed that short MWCNTs (~0.6 µm in length) induced significantly greater responses from the epithelial cells, whilst AM were particularly susceptible to long MWCNTs (~20 µm). These differences in the pattern of mediator release were associated with alternative profiles of JNK, p38 and ERK1/2 MAP kinase signal transduction within each cell type. This study, using highly relevant target human alveolar cells and well defined and characterized MWCNTs, shows marked cellular responses to the MWCNTs that vary according to the target cell type, as well as the aspect ratio of the MWCNT. PMID:25780270

Cytokinetics of adult rat SVZ after EAE.

PubMed

Sajad, Mir; Chawla, Raman; Zargan, Jamil; Umar, Sadiq; Sadaqat, Mir; Khan, Haider A

2011-01-31

Cytokinetics regulating cell cycle division can be modulated by several endogenous factors. EAE (experimental autoimmune encephalomyelitis) increases proliferation of progenitor cells in the subventricular zone (SVZ). Using cumulative and single S phase labeling with 5-bromo-2-deoxyuridine, we examined cell cycle kinetics of neural progenitor cells in the SVZ after EAE. 20% of the SVZ cell population was proliferating in adjuvant control rats. However, EAE significantly increased them up to 27% and these cells had a cell cycle length (TC) of 15.6h, significantly (P<0.05) shorter than the 19 h TC in non EAE SVZ cells. Few TUNEL (+) cells were detected in the SVZ cells of adjuvant controls. EAE increased (P<0.05) TUNEL (+) nuclei in SVZ suggesting early stage progenitor cell death. Cell cycle phase analysis revealed that EAE substantially shortened the length of the G1 phase (9.6h) compared with the G1 phase of 12.25 h in adjuvant control SVZ cells (P<0.05). This reduction in G1 contributes to EAE-induced reduction of TC because no significant changes were detected on the length of S, G2 and M phases between the two groups. Our results show a surge in proliferating progenitor cells in the SVZ with concomitant increase in apoptotic cell death after EAE. Furthermore, increase in the SVZ proliferation contributes to EAE-induced neurogenesis and this increase is regulated by shortening the G1 phase. Our investigation suggests the activation of quiescent cells in SVZ to generate actively proliferating progenitors. Moreover, the increase in the cell death in proliferating population may contribute towards negative regulation of proliferative cell number and hence diminished regenerative capacity of CNS following EAE. Copyright © 2010 Elsevier B.V. All rights reserved.

Diffusion lengths in irradiated N/P InP-on-Si solar cells

NASA Technical Reports Server (NTRS)

Wojtczuk, Steven; Colerico, Claudia; Summers, Geoffrey P.; Walters, Robert J.; Burke, Edward A.

1996-01-01

Indium phosphide (InP) solar cells were made on silicon (Si) wafers (InP/Si) by to take advantage of both the radiation-hardness properties of the InP solar cell and the light weight and low cost of Si wafers. The InP/Si cell application is for long duration and/or high radiation orbit space missions. Spire has made N/P InP/Si cells of sizes up to 2 cm by 4 cm with beginning-of-life (BOL) AM0 efficiencies over 13% (one-sun, 28C). These InP/Si cells have higher absolute efficiency and power density after a high radiation dose than gallium arsenide (GaAs) or silicon (Si) solar cells after a fluence of about 2e15 1 MeV electrons/sq. cm. In this work, we investigate the minority carrier (electron) base diffusion lengths in the N/P InP/Si cells. A quantum efficiency model was constructed for a 12% BOL AM0 N/P InP/Si cell which agreed well with the absolutely measured quantum efficiency and the sun-simulator measured AM0 photocurrent (30.1 mA/sq. cm). This model was then used to generate a table of AM0 photocurrents for a range of base diffusion lengths. AM0 photocurrents were then measured for irradiations up to 7.7e16 1 MeV electrons/sq. cm (the 12% BOL cell was 8% after the final irradiation). By comparing the measured photocurrents with the predicted photocurrents, base diffusion lengths were assigned at each fluence level. A damage coefficient K of 4e-8 and a starting (unirradiated) base electron diffusion length of 0.8 microns fits the data well. The quantum efficiency was measured again at the end of the experiment to verify that the photocurrent predicted by the model (25.5 mA/sq. cm) agreed with the simulator-measured photocurrent after irradiation (25.7 mA/sq. cm).

The spatial extent of excitatory and inhibitory zones in the receptive field of superficial layer hypercomplex cells

PubMed Central

Sillito, A. M.

1977-01-01

1. An investigation has been made of the extent of inhibitory and excitatory components in the receptive field of superficial layer hypercomplex cells in the cat's striate cortex and the relation of the components to the length preference exhibited by these cells. 2. Maximal responses were produced by an optimal length stimulus moving through a restricted region of the receptive field. The length of this receptive field region was less than the total length of the excitatory zone as mapped with a very short slit. Slits of similar length to the excitatory zone produced a smaller response than an optimal length slit. 3. An increase of slit length so that it passed over receptive field regions either side of the excitatory zone resulted in an elimination of the response. When background discharge levels were increased by the iontophoretic application of D, L-homocysteic acid slits of this length were observed to produce a suppression of the resting discharge as they passed over the receptive field. They did not modify the resting discharge level when it was induced by the iontophoretic application of the GABA antagonist bicuculline. This data is taken to indicate that long slits activate a powerful post-synaptic inhibitory input to the cell. 4. Maximal inhibitory effects were only observed if the testing slit passed over the receptive field centre. That is slits with a gap positioned midway along their length so as to exclude the optimal excitatory response region surprisingly tended to produce excitatory effects rather than the expected inhibitory effects. It appears that simultaneous stimulation of the receptive field centre is a precondition for the inhibitory effect of stimulation of regions either side of the excitatory zone to be activated. 5. It is suggested that the interneurones mediating the inhibitory input to the superficial layer hypercomplex cells are driven both by cells in adjacent hypercolumns with receptive fields spatially displaced to either side of the excitatory zone and by cells in the same column, optimal inhibitory effects only being achieved when both sets of input to the interneurone are activated. PMID:604459

Significantly lengthened telomere in granulosa cells from women with polycystic ovarian syndrome (PCOS).

PubMed

Wei, Duo; Xie, Juanke; Yin, Baoli; Hao, Haoying; Song, Xiaobing; Liu, Qi; Zhang, Cuilian; Sun, Yingpu

2017-07-01

Polycystic ovary syndrome (PCOS) is the most common endocrinopathy among women at reproductive age. However, its etiology remains poorly understood. Recent studies indicated that telomere length was related to PCOS. However, the association between telomere length and PCOS has only been shown in leucocytes and remained controversial across different studies. To clarify the association between telomere length and PCOS, the current study interrogated telomere length not only in leucocytes, but also in follicular granulosa cells, which is essential for folliculogenesis and steroidogenesis. Seventy-five patients with PCOS and 81 controls with mechanical infertility undergoing their first in vitro fertilization cycle were enrolled. Their peripheral blood and granulosa cells were collected on the oocyte retrieval day. Telomere length of both leucocytes in the blood and granulosa cells was assayed by quantitative polymerase chain reaction. No significant difference was found in the leucocyte telomere length between controls and PCOS patients (0.99 ± 0.44 vs. 1.00 ± 0.38, p = 0.93). Interestingly, when comparing telomere length in granulosa cells between controls and PCOS subjects, significantly lengthened telomere length was found in PCOS subjects (1.00 ± 0.37 vs. 1.57±0.67, p < 0.0001). After adjustments for age and body mass index, the p value remained significant (p < 0.0001). This finding reinforced the association between telomere abnormalities and PCOS. Given the importance of telomere length in cellular proliferation, our findings provided novel insights into the pathophysiology of PCOS that abnormalities in telomere length possibly disturb folliculogenesis and subsequently result in PCOS.

The change is length and width of the Sertoli cell nuclei in cytologic smears of testes with depopulation of the seminiferous epithelium.

PubMed

Banek, L; Posinovec, J

1980-09-15

The appearance of the Sertoli cells in cytological smears of tests with depopulation of the seminiferous epithelium is described. The mean values of the lengths and widths of the Sertoli cell nuclei in smears differed significantly between the depopulation and the control group (p < 0.01).

Telomere dynamics, end-to-end fusions and telomerase activation during the human fibroblast immortalization process.

PubMed

Ducray, C; Pommier, J P; Martins, L; Boussin, F D; Sabatier, L

1999-07-22

Loss of telomeric repeats during cell proliferation could play a role in senescence. It has been generally assumed that activation of telomerase prevents further telomere shortening and is essential for cell immortalization. In this study, we performed a detailed cytogenetic and molecular characterization of four SV40 transformed human fibroblastic cell lines by regularly monitoring the size distribution of terminal restriction fragments, telomerase activity and the associated chromosomal instability throughout immortalization. The mean TRF lengths progressively decreased in pre-crisis cells during the lifespan of the cultures. At crisis, telomeres reached a critical size, different among the cell lines, contributing to the peak of dicentric chromosomes, which resulted mostly from telomeric associations. We observed a direct correlation between short telomere length at crisis and chromosomal instability. In two immortal cell lines, although telomerase was detected, mean telomere length still continued to decrease whereas the number of dicentric chromosomes associated was stabilized. Thus telomerase could protect specifically telomeres which have reached a critical size against end-to-end dicentrics, while long telomeres continue to decrease, although at a slower rate as before crisis. This suggests a balance between elongation by telomerase and telomere shortening, towards a stabilized 'optimal' length.

Biophysical force regulation in 3D tumor cell invasion

NASA Astrophysics Data System (ADS)

Wu, Mingming

When embedded within 3D extracellular matrices (ECM), animal cells constantly probe and adapt to the ECM locally (at cell length scale) and exert forces and communicate with other cells globally (up to 10 times of cell length). It is now well accepted that mechanical crosstalk between animal cells and their microenvironment critically regulate cell function such as migration, proliferation and differentiation. Disruption of the cell-ECM crosstalk is implicated in a number of pathologic processes including tumor progression and fibrosis. Central to the problem of cell-ECM crosstalk is the physical force that cells generate. By measuring single cell generated force within 3D collagen matrices, we revealed a mechanical crosstalk mechanism between the tumor cells and the ECM. Cells generate sufficient force to stiffen collagen fiber network, and stiffer matrix, in return promotes larger cell force generation. Our work highlights the importance of fibrous nonlinear elasticity in regulating tumor cell-ECM interaction, and results may have implications in the rapid tissue stiffening commonly found in tumor progression and fibrosis. This work is partially supported by NIH Grants R21RR025801 and R21GM103388.

Improved Method for Linear B-Cell Epitope Prediction Using Antigen’s Primary Sequence

PubMed Central

Raghava, Gajendra P. S.

2013-01-01

One of the major challenges in designing a peptide-based vaccine is the identification of antigenic regions in an antigen that can stimulate B-cell’s response, also called B-cell epitopes. In the past, several methods have been developed for the prediction of conformational and linear (or continuous) B-cell epitopes. However, the existing methods for predicting linear B-cell epitopes are far from perfection. In this study, an attempt has been made to develop an improved method for predicting linear B-cell epitopes. We have retrieved experimentally validated B-cell epitopes as well as non B-cell epitopes from Immune Epitope Database and derived two types of datasets called Lbtope_Variable and Lbtope_Fixed length datasets. The Lbtope_Variable dataset contains 14876 B-cell epitope and 23321 non-epitopes of variable length where as Lbtope_Fixed length dataset contains 12063 B-cell epitopes and 20589 non-epitopes of fixed length. We also evaluated the performance of models on above datasets after removing highly identical peptides from the datasets. In addition, we have derived third dataset Lbtope_Confirm having 1042 epitopes and 1795 non-epitopes where each epitope or non-epitope has been experimentally validated in at least two studies. A number of models have been developed to discriminate epitopes and non-epitopes using different machine-learning techniques like Support Vector Machine, and K-Nearest Neighbor. We achieved accuracy from ∼54% to 86% using diverse s features like binary profile, dipeptide composition, AAP (amino acid pair) profile. In this study, for the first time experimentally validated non B-cell epitopes have been used for developing method for predicting linear B-cell epitopes. In previous studies, random peptides have been used as non B-cell epitopes. In order to provide service to scientific community, a web server LBtope has been developed for predicting and designing B-cell epitopes (http://crdd.osdd.net/raghava/lbtope/). PMID:23667458

Moderate and intense exercise lifestyles attenuate the effects of aging on telomere length and the survival and composition of T cell subpopulations.

PubMed

Silva, Léia Cristina Rodrigues; de Araújo, Adriana Ladeira; Fernandes, Juliana Ruiz; Matias, Manuella de Sousa Toledo; Silva, Paulo Roberto; Duarte, Alberto J S; Garcez Leme, Luiz Eugênio; Benard, Gil

2016-02-01

Studies indicate that exercise might delay human biological aging, but the effects of long-term exercise on T cell function are not well known. We tested the hypothesis that moderate or intense exercise lifestyle may attenuate the effects of aging on the telomere length and the survival and composition of T cell subpopulations. Elderly (65-85 years) with intense training lifestyle (IT, n = 15), moderate training lifestyle (MT, n = 16), and who never trained (NT, n = 15) were studied. Although the three groups presented the age-associated contraction of the TCD4(+)/TCD8(+) naïve compartments and expansion of the memory compartments, both training modalities were associated with lower proportion of terminally differentiated (CD45RA(+)CCR7(neg)) TCD4(+) and TCD8(+) cells, although among the latter cells, the reduction reached statistical significance only with IT. MT was associated with higher proportion of central memory TCD4(+) cells, while IT was associated with higher proportion of effector memory TCD8(+) cells. However, both training lifestyles were unable to modify the proportion of senescent (CD28(neg)) TCD8(+) cells. Telomeres were longer in T cells in both training groups; with IT, telomere length increased mainly in TCD8(+) cells, whereas with MT, a modest increase in telomere length was observed in both TCD8(+) and TCD4(+) cells. Reduced commitment to apoptosis of resting T cells, as assessed by caspase-3 and Bcl-2 expression, was seen predominantly with IT. Measurement of pro-inflammatory cytokines in serum and peripheral blood mononuclear cell (PBMC)'s supernatants did not show chronic low-grade inflammation in any of the groups. In conclusion, MT and IT lifestyles attenuated some of the effects of aging on the immune system.

Different Amounts of DNA in Newborn Cells of Escherichia coli Preclude a Role for the Chromosome in Size Control According to the "Adder" Model.

PubMed

Huls, Peter G; Vischer, Norbert O E; Woldringh, Conrad L

2018-01-01

According to the recently-revived adder model for cell size control, newborn cells of Escherichia coli will grow and divide after having added a constant size or length, ΔL , irrespective of their size at birth. Assuming exponential elongation, this implies that large newborns will divide earlier than small ones. The molecular basis for the constant size increment is still unknown. As DNA replication and cell growth are coordinated, the constant ΔL could be based on duplication of an equal amount of DNA, ΔG , present in newborn cells. To test this idea, we measured amounts of DNA and lengths of nucleoids in DAPI-stained cells growing in batch culture at slow and fast rates. Deeply-constricted cells were divided in two subpopulations of longer and shorter lengths than average; these were considered to represent large and small prospective daughter cells, respectively. While at slow growth, large and small prospective daughter cells contained similar amounts of DNA, fast growing cells with multiforked replicating chromosomes, showed a significantly higher amount of DNA (20%) in the larger cells. This observation precludes the hypothesis that Δ L is based on the synthesis of a constant ΔG . Growth curves were constructed for siblings generated by asymmetric division and growing according to the adder model. Under the assumption that all cells at the same growth rate exhibit the same time between initiation of DNA replication and cell division (i.e., constant C+D -period), the constructions predict that initiation occurs at different sizes ( Li ) and that, at fast growth, large newborn cells transiently contain more DNA than small newborns, in accordance with the observations. Because the state of segregation, measured as the distance between separated nucleoids, was found to be more advanced in larger deeply-constricted cells, we propose that in larger newborns nucleoid separation occurs faster and at a shorter length, allowing them to divide earlier. We propose a composite model in which both differential initiation and segregation leads to an adder-like behavior of large and small newborn cells.

Micro-scale and meso-scale architectural cues cooperate and compete to direct aligned tissue formation

PubMed Central

Gilchrist, Christopher L.; Ruch, David S.; Little, Dianne; Guilak, Farshid

2014-01-01

Tissue and biomaterial microenvironments provide architectural cues that direct important cell behaviors including cell shape, alignment, migration, and resulting tissue formation. These architectural features may be presented to cells across multiple length scales, from nanometers to millimeters in size. In this study, we examined how architectural cues at two distinctly different length scales, “micro-scale” cues on the order of ~1–2 μm, and “meso-scale” cues several orders of magnitude larger (>100 μm), interact to direct aligned neo-tissue formation. Utilizing a micro-photopatterning (μPP) model system to precisely arrange cell-adhesive patterns, we examined the effects of substrate architecture at these length scales on human mesenchymal stem cell (hMSC) organization, gene expression, and fibrillar collagen deposition. Both micro- and meso-scale architectures directed cell alignment and resulting tissue organization, and when combined, meso cues could enhance or compete against micro-scale cues. As meso boundary aspect ratios were increased, meso-scale cues overrode micro-scale cues and controlled tissue alignment, with a characteristic critical width (~500 μm) similar to boundary dimensions that exist in vivo in highly aligned tissues. Meso-scale cues acted via both lateral confinement (in a cell-density-dependent manner) and by permitting end-to-end cell arrangements that yielded greater fibrillar collagen deposition. Despite large differences in fibrillar collagen content and organization between μPP architectural conditions, these changes did not correspond with changes in gene expression of key matrix or tendon-related genes. These findings highlight the complex interplay between geometric cues at multiple length scales and may have implications for tissue engineering strategies, where scaffold designs that incorporate cues at multiple length scales could improve neo-tissue organization and resulting functional outcomes. PMID:25263687

[Construction of dengue virus-specific full-length fully human antibody libraries by mammalian display technology].

PubMed

Wen, Yangming; Lan, Kaijian; Wang, Junjie; Yu, Jingyi; Qu, Yarong; Zhao, Wei; Zhang, Fuchun; Tan, Wanlong; Cao, Hong; Zhou, Chen

2013-06-01

To construct dengue virus-specific full-length fully human antibody libraries using mammalian cell surface display technique. Total RNA was extracted from peripheral blood mononuclear cells (PBMCs) from convalescent patients with dengue fever. The reservoirs of the light chain and heavy chain variable regions (LCκ and VH) of the antibody genes were amplified by RT-PCR and inserted into the vector pDGB-HC-TM separately to construct the light chain and heavy chain libraries. The library DNAs were transfected into CHO cells and the expression of full-length fully human antibodies on the surface of CHO cells was analyzed by flow cytometry. Using 1.2 µg of the total RNA isolated from the PBMCs as the template, the LCκ and VH were amplified and the full-length fully human antibody mammalian display libraries were constructed. The kappa light chain gene library had a size of 1.45×10(4) and the heavy chain gene library had a size of 1.8×10(5). Sequence analysis showed that 8 out of the 10 light chain clones and 7 out of the 10 heavy chain clones randomly picked up from the constructed libraries contained correct open reading frames. FACS analysis demonstrated that all the 15 clones with correct open reading frames expressed full-length antibodies, which could be detected on CHO cell surfaces. After co-transfection of the heavy chain and light chain gene libraries into CHO cells, the expression of full-length antibodies on CHO cell surfaces could be detected by FACS analysis with an expressible diversity of the antibody library reaching 1.46×10(9) [(1.45×10(4)×80%)×(1.8×10(5)×70%)]. Using 1.2 µg of total RNA as template, the LCκ and VH full-length fully human antibody libraries against dengue virus have been successfully constructed with an expressible diversity of 10(9).

Critical telomerase activity for uncontrolled cell growth

NASA Astrophysics Data System (ADS)

Wesch, Neil L.; Burlock, Laura J.; Gooding, Robert J.

2016-08-01

The lengths of the telomere regions of chromosomes in a population of cells are modelled using a chemical master equation formalism, from which the evolution of the average number of cells of each telomere length is extracted. In particular, the role of the telomere-elongating enzyme telomerase on these dynamics is investigated. We show that for biologically relevant rates of cell birth and death, one finds a critical rate, R crit, of telomerase activity such that the total number of cells diverges. Further, R crit is similar in magnitude to the rates of mitosis and cell death. The possible relationship of this result to replicative immortality and its associated hallmark of cancer is discussed.

Evolution of spur-length diversity in Aquilegia petals is achieved solely through cell-shape anisotropy.

PubMed

Puzey, Joshua R; Gerbode, Sharon J; Hodges, Scott A; Kramer, Elena M; Mahadevan, L

2012-04-22

The role of petal spurs and specialized pollinator interactions has been studied since Darwin. Aquilegia petal spurs exhibit striking size and shape diversity, correlated with specialized pollinators ranging from bees to hawkmoths in a textbook example of adaptive radiation. Despite the evolutionary significance of spur length, remarkably little is known about Aquilegia spur morphogenesis and its evolution. Using experimental measurements, both at tissue and cellular levels, combined with numerical modelling, we have investigated the relative roles of cell divisions and cell shape in determining the morphology of the Aquilegia petal spur. Contrary to decades-old hypotheses implicating a discrete meristematic zone as the driver of spur growth, we find that Aquilegia petal spurs develop via anisotropic cell expansion. Furthermore, changes in cell anisotropy account for 99 per cent of the spur-length variation in the genus, suggesting that the true evolutionary innovation underlying the rapid radiation of Aquilegia was the mechanism of tuning cell shape.

Motion of variable-length MreB filaments at the bacterial cell membrane influences cell morphology

PubMed Central

Reimold, Christian; Defeu Soufo, Herve Joel; Dempwolff, Felix; Graumann, Peter L.

2013-01-01

The maintenance of rod-cell shape in many bacteria depends on actin-like MreB proteins and several membrane proteins that interact with MreB. Using superresolution microscopy, we show that at 50-nm resolution, Bacillus subtilis MreB forms filamentous structures of length up to 3.4 μm underneath the cell membrane, which run at angles diverging up to 40° relative to the cell circumference. MreB from Escherichia coli forms at least 1.4-μm-long filaments. MreB filaments move along various tracks with a maximal speed of 85 nm/s, and the loss of ATPase activity leads to the formation of extended and static filaments. Suboptimal growth conditions lead to formation of patch-like structures rather than extended filaments. Coexpression of wild-type MreB with MreB mutated in the subunit interface leads to formation of shorter MreB filaments and a strong effect on cell shape, revealing a link between filament length and cell morphology. Thus MreB has an extended-filament architecture with the potential to position membrane proteins over long distances, whose localization in turn may affect the shape of the cell wall. PMID:23783036

Motion of variable-length MreB filaments at the bacterial cell membrane influences cell morphology.

PubMed

Reimold, Christian; Defeu Soufo, Herve Joel; Dempwolff, Felix; Graumann, Peter L

2013-08-01

The maintenance of rod-cell shape in many bacteria depends on actin-like MreB proteins and several membrane proteins that interact with MreB. Using superresolution microscopy, we show that at 50-nm resolution, Bacillus subtilis MreB forms filamentous structures of length up to 3.4 μm underneath the cell membrane, which run at angles diverging up to 40° relative to the cell circumference. MreB from Escherichia coli forms at least 1.4-μm-long filaments. MreB filaments move along various tracks with a maximal speed of 85 nm/s, and the loss of ATPase activity leads to the formation of extended and static filaments. Suboptimal growth conditions lead to formation of patch-like structures rather than extended filaments. Coexpression of wild-type MreB with MreB mutated in the subunit interface leads to formation of shorter MreB filaments and a strong effect on cell shape, revealing a link between filament length and cell morphology. Thus MreB has an extended-filament architecture with the potential to position membrane proteins over long distances, whose localization in turn may affect the shape of the cell wall.

A numerical model for charge transport and energy conversion of perovskite solar cells.

PubMed

Zhou, Yecheng; Gray-Weale, Angus

2016-02-14

Based on the continuity equations and Poisson's equation, we developed a numerical model for perovskite solar cells. Due to different working mechanisms, the model for perovskite solar cells differs from that of silicon solar cells and Dye Sensitized Solar Cells. The output voltage and current are calculated differently, and in a manner suited in particular to perovskite organohalides. We report a test of our equations against experiment with good agreement. Using this numerical model, it was found that performances of solar cells increase with charge carrier's lifetimes, mobilities and diffusion lengths. The open circuit voltage (Voc) of a solar cell is dependent on light intensities, and charge carrier lifetimes. Diffusion length and light intensity determine the saturated current (Jsc). Additionally, three possible guidelines for the design and fabrication of perovskite solar cells are suggested by our calculations. Lastly, we argue that concentrator perovskite solar cells are promising.

Biological responses according to the shape and size of carbon nanotubes in BEAS-2B and MESO-1 cells

PubMed Central

Haniu, Hisao; Saito, Naoto; Matsuda, Yoshikazu; Tsukahara, Tamotsu; Usui, Yuki; Maruyama, Kayo; Takanashi, Seiji; Aoki, Kaoru; Kobayashi, Shinsuke; Nomura, Hiroki; Tanaka, Manabu; Okamoto, Masanori; Kato, Hiroyuki

2014-01-01

This study aimed to investigate the influence of the shape and size of multi-walled carbon nanotubes (MWCNTs) and cup-stacked carbon nanotubes (CSCNTs) on biological responses in vitro. Three types of MWCNTs – VGCF®-X, VGCF®-S, and VGCF® (vapor grown carbon fibers; with diameters of 15, 80, and 150 nm, respectively) – and three CSCNTs of different lengths (CS-L, 20–80 μm; CS-S, 0.5–20 μm; and CS-M, of intermediate length) were tested. Human bronchial epithelial (BEAS-2B) and malignant pleural mesothelioma cells were exposed to the CNTs (1–50 μg/mL), and cell viability, permeability, uptake, total reactive oxygen species/superoxide production, and intracellular acidity were measured. CSCNTs were less toxic than MWCNTs in both cell types over a 24-hour exposure period. The cytotoxicity of endocytosed MWCNTs varied according to cell type/size, while that of CSCNTs depended on tube length irrespective of cell type. CNT diameter and length influenced cell aggregation and injury extent. Intracellular acidity increased independently of lysosomal activity along with the number of vacuoles in BEAS-2B cells exposed for 24 hours to either CNT (concentration, 10 μg/mL). However, total reactive oxygen species/superoxide generation did not contribute to cytotoxicity. The results demonstrate that CSCNTs could be suitable for biological applications and that CNT shape and size can have differential effects depending on cell type, which can be exploited in the development of highly specialized, biocompatible CNTs. PMID:24790438

Telomere length variation in tumor cells and cancer‐associated fibroblasts: potential biomarker for hepatocellular carcinoma

PubMed Central

Ma, Li‐Jie; Wang, Xiao‐Ying; Duan, Meng; Liu, Long‐Zi; Shi, Jie‐Yi; Dong, Liang‐Qing; Yang, Liu‐Xiao; Wang, Zhi‐Chao; Ding, Zhen‐Bin; Ke, Ai‐Wu; Cao, Ya; Zhang, Xiao‐Ming; Zhou, Jian; Fan, Jia

2017-01-01

Abstract The role of telomere dysfunction and aberrant telomerase activities in hepatocellular carcinoma (HCC) has been overlooked for many years. This study aimed to delineate the variation and prognostic value of telomere length in HCC. Telomere‐specific fluorescence in situ hybridization (FISH) and qPCR were used to evaluate telomere length in HCC cell lines, tumor tissues, and isolated non‐tumor cells within the tumor. Significant telomere attrition was found in tumor cells and cancer‐associated fibroblasts (CAFs) compared to their normal counterparts, but not in intratumor leukocytes or bile duct epithelial cells. Clinical relevance and prognostic value of telomere length were investigated on tissue microarrays of 257 surgically treated HCC patients. Reduced intensity of telomere signals in tumor cells or CAFs correlated with larger tumor size and the presence of vascular invasion (p < 0.05). Shortened telomeres in tumor cells or CAFs associated with reduced survival and increased recurrence, and were identified as independent prognosticators for HCC patients (p < 0.05). These findings were validated in an independent HCC cohort of 371 HCC patients from The Cancer Genome Atlas (TCGA) database, confirming telomere attrition and its prognostic value in HCC. We also showed that telomerase reverse transcriptase promoter (TERTp) mutation correlated with telomere shortening in HCC. Telomere variation in tumor cells and non‐tumor cells within the tumor microenvironment of HCC was a valuable prognostic biomarker for this fatal malignancy. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. PMID:28833123

The Carboxyl Tail of Connexin32 Regulates Gap Junction Assembly in Human Prostate and Pancreatic Cancer Cells*

PubMed Central

Katoch, Parul; Mitra, Shalini; Ray, Anuttoma; Kelsey, Linda; Roberts, Brett J.; Wahl, James K.; Johnson, Keith R.; Mehta, Parmender P.

2015-01-01

Connexins, the constituent proteins of gap junctions, are transmembrane proteins. A connexin (Cx) traverses the membrane four times and has one intracellular and two extracellular loops with the amino and carboxyl termini facing the cytoplasm. The transmembrane and the extracellular loop domains are highly conserved among different Cxs, whereas the carboxyl termini, often called the cytoplasmic tails, are highly divergent. We have explored the role of the cytoplasmic tail of Cx32, a Cx expressed in polarized and differentiated cells, in regulating gap junction assembly. Our results demonstrate that compared with the full-length Cx32, the cytoplasmic tail-deleted Cx32 is assembled into small gap junctions in human pancreatic and prostatic cancer cells. Our results further document that the expression of the full-length Cx32 in cells, which express the tail-deleted Cx32, increases the size of gap junctions, whereas the expression of the tail-deleted Cx32 in cells, which express the full-length Cx32, has the opposite effect. Moreover, we show that the tail is required for the clustering of cell-cell channels and that in cells expressing the tail-deleted Cx32, the expression of cell surface-targeted cytoplasmic tail alone is sufficient to enhance the size of gap junctions. Our live-cell imaging data further demonstrate that gap junctions formed of the tail-deleted Cx32 are highly mobile compared with those formed of full-length Cx32. Our results suggest that the cytoplasmic tail of Cx32 is not required to initiate the assembly of gap junctions but for their subsequent growth and stability. Our findings suggest that the cytoplasmic tail of Cx32 may be involved in regulating the permeability of gap junctions by regulating their size. PMID:25548281

Moderate stem-cell telomere shortening rate postpones cancer onset in a stochastic model

NASA Astrophysics Data System (ADS)

Holbek, Simon; Bendtsen, Kristian Moss; Juul, Jeppe

2013-10-01

Mammalian cells are restricted from proliferating indefinitely. Telomeres at the end of each chromosome are shortened at cell division and when they reach a critical length, the cell will enter permanent cell cycle arrest—a state known as senescence. This mechanism is thought to be tumor suppressing, as it helps prevent precancerous cells from dividing uncontrollably. Stem cells express the enzyme telomerase, which elongates the telomeres, thereby postponing senescence. However, unlike germ cells and most types of cancer cells, stem cells only express telomerase at levels insufficient to fully maintain the length of their telomeres, leading to a slow decline in proliferation potential. It is not yet fully understood how this decline influences the risk of cancer and the longevity of the organism. We here develop a stochastic model to explore the role of telomere dynamics in relation to both senescence and cancer. The model describes the accumulation of cancerous mutations in a multicellular organism and creates a coherent theoretical framework for interpreting the results of several recent experiments on telomerase regulation. We demonstrate that the longest average cancer-free lifespan before cancer onset is obtained when stem cells start with relatively long telomeres that are shortened at a steady rate at cell division. Furthermore, the risk of cancer early in life can be reduced by having a short initial telomere length. Finally, our model suggests that evolution will favor a shorter than optimal average cancer-free lifespan in order to postpone cancer onset until late in life.

Realizing the Translational Potential of Telomere Length Variation as a Tissue Based Prognostic Marker for Prostate Cancer

DTIC Science & Technology

2016-10-01

Award Number: W81XWH-12-1-0545 TITLE: Realizing the Translational Potential of Telomere Length Variation as a Tissue- Based Prognostic Marker for...30 Sep 2015 - 29 Sep 2016 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Realizing the Translational Potential of Telomere Length Variation as a Tissue...HPFS), whether the combination of telomere length variability in prostate cancer cells and short telomere length in cancer-associated stromal cells is

Beat-to-Beat Variation in Periodicity of Local Calcium Releases Contributes to Intrinsic Variations of Spontaneous Cycle Length in Isolated Single Sinoatrial Node Cells

PubMed Central

Monfredi, Oliver; Maltseva, Larissa A.; Spurgeon, Harold A.; Boyett, Mark R.; Lakatta, Edward G.; Maltsev, Victor A.

2013-01-01

Spontaneous, submembrane local Ca2+ releases (LCRs) generated by the sarcoplasmic reticulum in sinoatrial nodal cells, the cells of the primary cardiac pacemaker, activate inward Na+/Ca2+-exchange current to accelerate the diastolic depolarization rate, and therefore to impact on cycle length. Since LCRs are generated by Ca2+ release channel (i.e. ryanodine receptor) openings, they exhibit a degree of stochastic behavior, manifested as notable cycle-to-cycle variations in the time of their occurrence. Aim The present study tested whether variation in LCR periodicity contributes to intrinsic (beat-to-beat) cycle length variability in single sinoatrial nodal cells. Methods We imaged single rabbit sinoatrial nodal cells using a 2D-camera to capture LCRs over the entire cell, and, in selected cells, simultaneously measured action potentials by perforated patch clamp. Results LCRs begin to occur on the descending part of the action potential-induced whole-cell Ca2+ transient, at about the time of the maximum diastolic potential. Shortly after the maximum diastolic potential (mean 54±7.7 ms, n = 14), the ensemble of waxing LCR activity converts the decay of the global Ca2+ transient into a rise, resulting in a late, whole-cell diastolic Ca2+ elevation, accompanied by a notable acceleration in diastolic depolarization rate. On average, cells (n = 9) generate 13.2±3.7 LCRs per cycle (mean±SEM), varying in size (7.1±4.2 µm) and duration (44.2±27.1 ms), with both size and duration being greater for later-occurring LCRs. While the timing of each LCR occurrence also varies, the LCR period (i.e. the time from the preceding Ca2+ transient peak to an LCR’s subsequent occurrence) averaged for all LCRs in a given cycle closely predicts the time of occurrence of the next action potential, i.e. the cycle length. Conclusion Intrinsic cycle length variability in single sinoatrial nodal cells is linked to beat-to-beat variations in the average period of individual LCRs each cycle. PMID:23826247

Paternal age and telomere length in twins: the germ stem cell selection paradigm

PubMed Central

Hjelmborg, Jacob B; Dalgård, Christine; Mangino, Massimo; Spector, Tim D; Halekoh, Ulrich; Möller, Sören; Kimura, Masayuki; Horvath, Kent; Kark, Jeremy D; Christensen, Kaare; Kyvik, Kirsten O; Aviv, Abraham

2015-01-01

Telomere length, a highly heritable trait, is longer in offspring of older fathers. This perplexing feature has been attributed to the longer telomeres in sperm of older men and it might be an ‘epigenetic’ mechanism through which paternal age plays a role in telomere length regulation in humans. Based on two independent (discovery and replication) twin studies, comprising 889 twin pairs, we show an increase in the resemblance of leukocyte telomere length between dizygotic twins of older fathers, which is not seen in monozygotic twins. This phenomenon might result from a paternal age-dependent germ stem cell selection process, whereby the selected stem cells have longer telomeres, are more homogenous with respect to telomere length, and share resistance to aging. PMID:25865872

Computing the Length of the Shortest Telomere in the Nucleus

NASA Astrophysics Data System (ADS)

Dao Duc, K.; Holcman, D.

2013-11-01

The telomere length can either be shortened or elongated by an enzyme called telomerase after each cell division. Interestingly, the shortest telomere is involved in controlling the ability of a cell to divide. Yet, its dynamics remains elusive. We present here a stochastic approach where we model this dynamics using a Markov jump process. We solve the forward Fokker-Planck equation to obtain the steady state distribution and the statistical moments of telomere lengths. We focus specifically on the shortest one and we estimate its length difference with the second shortest telomere. After extracting key parameters such as elongation and shortening dynamics from experimental data, we compute the length of telomeres in yeast and obtain as a possible prediction the minimum concentration of telomerase required to ensure a proper cell division.

Length and activity of the root apical meristem revealed in vivo by infrared imaging.

PubMed

Bizet, François; Hummel, Irène; Bogeat-Triboulot, Marie-Béatrice

2015-03-01

Understanding how cell division and cell elongation influence organ growth and development is a long-standing issue in plant biology. In plant roots, most of the cell divisions occur in a short and specialized region, the root apical meristem (RAM). Although RAM activity has been suggested to be of high importance to understand how roots grow and how the cell cycle is regulated, few experimental and numeric data are currently available. The characterization of the RAM is difficult and essentially based upon cell length measurements through destructive and time-consuming microscopy approaches. Here, a new non-invasive method is described that couples infrared light imaging and kinematic analyses and that allows in vivo measurements of the RAM length. This study provides a detailed description of the RAM activity, especially in terms of cell flux and cell division rate. We focused on roots of hydroponic grown poplars and confirmed our method on maize roots. How the RAM affects root growth rate is studied by taking advantage of the high inter-individual variability of poplar root growth. An osmotic stress was applied and did not significantly affect the RAM length, highlighting its homeostasis in short to middle-term responses. The methodology described here simplifies a lot experimental procedures, allows an increase in the number of individuals that can be taken into account in experiments, and means new experiments can be formulated that allow temporal monitoring of the RAM length. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Current Perspectives of Telomerase Structure and Function in Eukaryotes with Emerging Views on Telomerase in Human Parasites.

PubMed

Dey, Abhishek; Chakrabarti, Kausik

2018-01-24

Replicative capacity of a cell is strongly correlated with telomere length regulation. Aberrant lengthening or reduction in the length of telomeres can lead to health anomalies, such as cancer or premature aging. Telomerase is a master regulator for maintaining replicative potential in most eukaryotic cells. It does so by controlling telomere length at chromosome ends. Akin to cancer cells, most single-cell eukaryotic pathogens are highly proliferative and require persistent telomerase activity to maintain constant length of telomere and propagation within their host. Although telomerase is key to unlimited cellular proliferation in both cases, not much was known about the role of telomerase in human parasites (malaria, Trypanosoma , etc.) until recently. Since telomerase regulation is mediated via its own structural components, interactions with catalytic reverse transcriptase and several factors that can recruit and assemble telomerase to telomeres in a cell cycle-dependent manner, we compare and discuss here recent findings in telomerase biology in cancer, aging and parasitic diseases to give a broader perspective of telomerase function in human diseases.

Finite length Taylor Couette flow

NASA Technical Reports Server (NTRS)

Streett, C. L.; Hussaini, M. Y.

1987-01-01

Axisymmetric numerical solutions of the unsteady Navier-Stokes equations for flow between concentric rotating cylinders of finite length are obtained by a spectral collocation method. These representative results pertain to two-cell/one-cell exchange process, and are compared with recent experiments.

High-throughput full-length single-cell mRNA-seq of rare cells.

PubMed

Ooi, Chin Chun; Mantalas, Gary L; Koh, Winston; Neff, Norma F; Fuchigami, Teruaki; Wong, Dawson J; Wilson, Robert J; Park, Seung-Min; Gambhir, Sanjiv S; Quake, Stephen R; Wang, Shan X

2017-01-01

Single-cell characterization techniques, such as mRNA-seq, have been applied to a diverse range of applications in cancer biology, yielding great insight into mechanisms leading to therapy resistance and tumor clonality. While single-cell techniques can yield a wealth of information, a common bottleneck is the lack of throughput, with many current processing methods being limited to the analysis of small volumes of single cell suspensions with cell densities on the order of 107 per mL. In this work, we present a high-throughput full-length mRNA-seq protocol incorporating a magnetic sifter and magnetic nanoparticle-antibody conjugates for rare cell enrichment, and Smart-seq2 chemistry for sequencing. We evaluate the efficiency and quality of this protocol with a simulated circulating tumor cell system, whereby non-small-cell lung cancer cell lines (NCI-H1650 and NCI-H1975) are spiked into whole blood, before being enriched for single-cell mRNA-seq by EpCAM-functionalized magnetic nanoparticles and the magnetic sifter. We obtain high efficiency (> 90%) capture and release of these simulated rare cells via the magnetic sifter, with reproducible transcriptome data. In addition, while mRNA-seq data is typically only used for gene expression analysis of transcriptomic data, we demonstrate the use of full-length mRNA-seq chemistries like Smart-seq2 to facilitate variant analysis of expressed genes. This enables the use of mRNA-seq data for differentiating cells in a heterogeneous population by both their phenotypic and variant profile. In a simulated heterogeneous mixture of circulating tumor cells in whole blood, we utilize this high-throughput protocol to differentiate these heterogeneous cells by both their phenotype (lung cancer versus white blood cells), and mutational profile (H1650 versus H1975 cells), in a single sequencing run. This high-throughput method can help facilitate single-cell analysis of rare cell populations, such as circulating tumor or endothelial cells, with demonstrably high-quality transcriptomic data.

Correlation between the extent of pneumatization of Agger agger Nasi nasi cells and the anterior-to-posterior length of the frontal recess: A a computer-assisted anatomical study.

PubMed

Altıntaş, Ahmet; Çelik, Mustafa; Yegin, Yakup; Canpolat, Sinan; Olgun, Burak; Tülin Kayhan, Fatma

2017-06-30

To explore the correlation between the volume of the aAgger nNasi (AN) cell bulge and the A-P length of the frontal recess (FR). In total, 120 patients, who underwent septoplasty, were included. All patients underwent preoperative paranasal sinus computed tomography of the paranasal sinuses (PNS CT) imaging. In total, CT data on of all 120 PNSs patients were analyzed in terms of thewith respect to the extent of pneumatization of the AN cell bulge and the A-P dimensions of the FR. Each side was analyzed separately. We included 120 patients,: 78 (65.0%) females and 42 (35.0 %) males. Their average age was 33.7 ± 11.6 years (range: 18-65 years). The mean volume of the AN cell bulge was 0.26 ± 0.4 mm3 on both the right and left sides. The A-P length of the FR was 7.7 ± 2.2 mm. No significant between-side difference in the mean volume of the AN cell bulge was apparent observed (p=0.906). This volume did not differ significantly by age or sex (p=0.844 and p=0.971, respectively). We found no correlation between the volume of the AN cell bulge and the A-P length of the FR (r = 0.098, p=0.192). In the present study, no correlation between AN cell volume and the A-P length of the FR was found. When studying the anatomical complexity of the FR, it is essential to consider the AN cell volume. We suggest that preoperative CT imaging is critical when endoscopic sinus surgery is planned. However, further studies with larger numbers of patients are needed to explore the relationship between AN cell pneumatization and the anatomy of the FR.

Timing the start of division in E. coli: a single-cell study

NASA Astrophysics Data System (ADS)

Reshes, G.; Vanounou, S.; Fishov, I.; Feingold, M.

2008-12-01

We monitor the shape dynamics of individual E. coli cells using time-lapse microscopy together with accurate image analysis. This allows measuring the dynamics of single-cell parameters throughout the cell cycle. In previous work, we have used this approach to characterize the main features of single-cell morphogenesis between successive divisions. Here, we focus on the behavior of the parameters that are related to cell division and study their variation over a population of 30 cells. In particular, we show that the single-cell data for the constriction width dynamics collapse onto a unique curve following appropriate rescaling of the corresponding variables. This suggests the presence of an underlying time scale that determines the rate at which the cell cycle advances in each individual cell. For the case of cell length dynamics a similar rescaling of variables emphasizes the presence of a breakpoint in the growth rate at the time when division starts, τc. We also find that the τc of individual cells is correlated with their generation time, τg, and inversely correlated with the corresponding length at birth, L0. Moreover, the extent of the T-period, τg - τc, is apparently independent of τg. The relations between τc, τg and L0 indicate possible compensation mechanisms that maintain cell length variability at about 10%. Similar behavior was observed for both fast-growing cells in a rich medium (LB) and for slower growth in a minimal medium (M9-glucose). To reveal the molecular mechanisms that lead to the observed organization of the cell cycle, we should further extend our approach to monitor the formation of the divisome.

Positive Feedback Keeps Duration of Mitosis Temporally Insulated from Upstream Cell-Cycle Events.

PubMed

Araujo, Ana Rita; Gelens, Lendert; Sheriff, Rahuman S M; Santos, Silvia D M

2016-10-20

Cell division is characterized by a sequence of events by which a cell gives rise to two daughter cells. Quantitative measurements of cell-cycle dynamics in single cells showed that despite variability in G1-, S-, and G2 phases, duration of mitosis is short and remarkably constant. Surprisingly, there is no correlation between cell-cycle length and mitotic duration, suggesting that mitosis is temporally insulated from variability in earlier cell-cycle phases. By combining live cell imaging and computational modeling, we showed that positive feedback is the molecular mechanism underlying the temporal insulation of mitosis. Perturbing positive feedback gave rise to a sluggish, variable entry and progression through mitosis and uncoupled duration of mitosis from variability in cell cycle length. We show that positive feedback is important to keep mitosis short, constant, and temporally insulated and anticipate it might be a commonly used regulatory strategy to create modularity in other biological systems. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Optimization of yield in magnetic cell separations using nickel nanowires of different lengths.

PubMed

Hultgren, Anne; Tanase, Monica; Felton, Edward J; Bhadriraju, Kiran; Salem, Aliasger K; Chen, Christopher S; Reich, Daniel H

2005-01-01

Ferromagnetic nanowires are shown to perform both high yield and high purity single-step cell separations on cultures of NIH-3T3 mouse fibroblast cells. The nanowires are made by electrochemical deposition in nanoporous templates, permitting detailed control of their chemical and physical properties. When added to fibroblast cell cultures, the nanowires are internalized by the cells via the integrin-mediated adhesion pathway. The effectiveness of magnetic cell separations using Ni nanowires 350 nm in diameter and 5-35 micrometers long in field gradients of 40 T/m was compared to commercially available superparamagnetic beads. The percent yield of the separated populations is found to be optimized when the length of the nanowire is matched to the diameter of the cells in the culture. Magnetic cell separations performed under these conditions achieve 80% purity and 85% yield, a 4-fold increase over the beads. This effect is shown to be robust when the diameter of the cell is changed within the same cell line using mitomycin-C.

Static length changes of cochlear outer hair cells can tune low-frequency hearing

PubMed Central

Ciganović, Nikola; Warren, Rebecca L.; Keçeli, Batu; Jacob, Stefan

2018-01-01

The cochlea not only transduces sound-induced vibration into neural spikes, it also amplifies weak sound to boost its detection. Actuators of this active process are sensory outer hair cells in the organ of Corti, whereas the inner hair cells transduce the resulting motion into electric signals that propagate via the auditory nerve to the brain. However, how the outer hair cells modulate the stimulus to the inner hair cells remains unclear. Here, we combine theoretical modeling and experimental measurements near the cochlear apex to study the way in which length changes of the outer hair cells deform the organ of Corti. We develop a geometry-based kinematic model of the apical organ of Corti that reproduces salient, yet counter-intuitive features of the organ’s motion. Our analysis further uncovers a mechanism by which a static length change of the outer hair cells can sensitively tune the signal transmitted to the sensory inner hair cells. When the outer hair cells are in an elongated state, stimulation of inner hair cells is largely inhibited, whereas outer hair cell contraction leads to a substantial enhancement of sound-evoked motion near the hair bundles. This novel mechanism for regulating the sensitivity of the hearing organ applies to the low frequencies that are most important for the perception of speech and music. We suggest that the proposed mechanism might underlie frequency discrimination at low auditory frequencies, as well as our ability to selectively attend auditory signals in noisy surroundings. PMID:29351276

Positional signaling and expression of ENHANCER OF TRY AND CPC1 are tuned to increase root hair density in response to phosphate deficiency in Arabidopsis thaliana.

PubMed

Savage, Natasha; Yang, Thomas J W; Chen, Chung Ying; Lin, Kai-Lan; Monk, Nicholas A M; Schmidt, Wolfgang

2013-01-01

Phosphate (Pi) deficiency induces a multitude of responses aimed at improving the acquisition of Pi, including an increased density of root hairs. To understand the mechanisms involved in Pi deficiency-induced alterations of the root hair phenotype in Arabidopsis (Arabidopsis thaliana), we analyzed the patterning and length of root epidermal cells under control and Pi-deficient conditions in wild-type plants and in four mutants defective in the expression of master regulators of cell fate, CAPRICE (CPC), ENHANCER OF TRY AND CPC 1 (ETC1), WEREWOLF (WER) and SCRAMBLED (SCM). From this analysis we deduced that the longitudinal cell length of root epidermal cells is dependent on the correct perception of a positional signal ('cortical bias') in both control and Pi-deficient plants; mutants defective in the receptor of the signal, SCM, produced short cells characteristic of root hair-forming cells (trichoblasts). Simulating the effect of cortical bias on the time-evolving probability of cell fate supports a scenario in which a compromised positional signal delays the time point at which non-hair cells opt out the default trichoblast pathway, resulting in short, trichoblast-like non-hair cells. Collectively, our data show that Pi-deficient plants increase root hair density by the formation of shorter cells, resulting in a higher frequency of hairs per unit root length, and additional trichoblast cell fate assignment via increased expression of ETC1.

Chromosomal instability and telomere shortening in long-term culture of hematopoietic stem cells: insights from a cell culture model of RPS14 haploinsufficiency.

PubMed

Thomay, K; Schienke, A; Vajen, B; Modlich, U; Schambach, A; Hofmann, W; Schlegelberger, B; Göhring, G

2014-01-01

The fate of cultivated primary hematopoietic stem cells (HSCs) with respect to genetic instability and telomere attrition has not yet been described in great detail. Thus, knowledge of the genetic constitution of HSCs is important when interpreting results of HSCs in culture. While establishing a cell culture model for myelodysplastic syndrome with a deletion in 5q by performing RPS14 knockdown, we found surprising data that may be of importance for any CD34+ cell culture experiments. We performed cytogenetic analyses and telomere length measurement on transduced CD34+ cells and untransduced control cells to observe the effects of long-term culturing. Initially, CD34+ cells had a normal median telomere length of about 12 kb and showed no signs of chromosomal instability. During follow-up, the median telomere length seemed to decrease and, simultaneously, increased chromosomal instability could be observed - in modified and control cells. One culture showed a clonal monosomy 7 - independent of prior RPS14 knockdown. During further culturing, it seemed that the telomeres re-elongated, and chromosomes stabilized, while TERT expression was not elevated. In summary, irrespective of our results of RPS14 knockdown in the long-term culture of CD34+ cells, it becomes clear that cell culture artefacts inducing telomere shortening and chromosomal instability have to be taken into account and regular cytogenetic analyses should always be performed. © 2013 S. Karger AG, Basel.

Positional Signaling and Expression of ENHANCER OF TRY AND CPC1 Are Tuned to Increase Root Hair Density in Response to Phosphate Deficiency in Arabidopsis thaliana

PubMed Central

Savage, Natasha; Yang, Thomas J. W.; Chen, Chung Ying; Lin, Kai-Lan; Monk, Nicholas A. M.; Schmidt, Wolfgang

2013-01-01

Phosphate (Pi) deficiency induces a multitude of responses aimed at improving the acquisition of Pi, including an increased density of root hairs. To understand the mechanisms involved in Pi deficiency-induced alterations of the root hair phenotype in Arabidopsis (Arabidopsis thaliana), we analyzed the patterning and length of root epidermal cells under control and Pi-deficient conditions in wild-type plants and in four mutants defective in the expression of master regulators of cell fate, CAPRICE (CPC), ENHANCER OF TRY AND CPC 1 (ETC1), WEREWOLF (WER) and SCRAMBLED (SCM). From this analysis we deduced that the longitudinal cell length of root epidermal cells is dependent on the correct perception of a positional signal (‘cortical bias’) in both control and Pi-deficient plants; mutants defective in the receptor of the signal, SCM, produced short cells characteristic of root hair-forming cells (trichoblasts). Simulating the effect of cortical bias on the time-evolving probability of cell fate supports a scenario in which a compromised positional signal delays the time point at which non-hair cells opt out the default trichoblast pathway, resulting in short, trichoblast-like non-hair cells. Collectively, our data show that Pi-deficient plants increase root hair density by the formation of shorter cells, resulting in a higher frequency of hairs per unit root length, and additional trichoblast cell fate assignment via increased expression of ETC1. PMID:24130712

Stroma-induced Jagged1 expression drives PC3 prostate cancer cell migration; disparate effects of RIP-generated proteolytic fragments on cell behaviour and Notch signaling

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

Delury, Craig, E-mail: c.delury@lancaster.ac.uk; Hart, Claire, E-mail: claire.hart@manchester.ac.uk; Brown, Mick, E-mail: michael.brown@ics.manchester.ac.uk

The Notch ligand Jagged1 is subject to regulated intramembrane proteolysis (RIP) which yields a soluble ectodomain (sJag) and a soluble Jagged1 intracellular domain (JICD). The full-length Jagged1 protein enhances prostate cancer (PCa) cell proliferation and is highly expressed in metastatic cells. However, little is known regarding the mechanisms by which Jagged1 or its RIP-generated fragments might promote PCa bone metastasis. In the current study we show that bone marrow stroma (BMS) induces Jagged1 expression in bone metastatic prostate cancer PC3 cells and that this enhanced expression is mechanistically linked to the promotion of cell migration. We also show that RIP-generatedmore » Jagged1 fragments exert disparate effects on PC3 cell behaviour and Notch signaling. In conclusion, the expression of both the full-length ligand and its RIP-generated fragments must be considered in tandem when attempting to regulate Jagged1 as a possible PCa therapy. - Highlights: • Bone marrow stroma induces Jagged1 expression in prostate cancer (PCa) PC3 cells. • This enhanced expression of full-length Jagged1 is required for PC3 cell migration. • Proteolytic fragments of Jagged1 exert disparate effects on PC3 cell behaviour. • Effects of fragments on cell behaviour do not correlate with Notch signaling. • Effects of Jagged1 and its fragments on PCa metastasis likely to be complex.« less

Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells.

PubMed

Hornstein, Benjamin D; Roman, Dany; Arévalo-Soliz, Lirio M; Engevik, Melinda A; Zechiedrich, Lynn

2016-01-01

The ability to produce extremely small and circular supercoiled vectors has opened new territory for improving non-viral gene therapy vectors. In this work, we compared transfection of supercoiled DNA vectors ranging from 383 to 4,548 bp, each encoding shRNA against GFP under control of the H1 promoter. We assessed knockdown of GFP by electroporation into HeLa cells. All of our vectors entered cells in comparable numbers when electroporated with equal moles of DNA. Despite similar cell entry, we found length-dependent differences in how efficiently the vectors knocked down GFP. As vector length increased up to 1,869 bp, GFP knockdown efficiency per mole of transfected DNA increased. From 1,869 to 4,257 bp, GFP knockdown efficiency per mole was steady, then decreased with increasing vector length. In comparing GFP knockdown with equal masses of vectors, we found that the shorter vectors transfect more efficiently per nanogram of DNA transfected. Our results rule out cell entry and DNA mass as determining factors for gene knockdown efficiency via electroporation. The length-dependent effects we have uncovered are likely explained by differences in nuclear translocation or transcription. These data add an important step towards clinical applications of non-viral vector delivery.

Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells

PubMed Central

Hornstein, Benjamin D.; Roman, Dany; Arévalo-Soliz, Lirio M.; Engevik, Melinda A.

2016-01-01

The ability to produce extremely small and circular supercoiled vectors has opened new territory for improving non-viral gene therapy vectors. In this work, we compared transfection of supercoiled DNA vectors ranging from 383 to 4,548 bp, each encoding shRNA against GFP under control of the H1 promoter. We assessed knockdown of GFP by electroporation into HeLa cells. All of our vectors entered cells in comparable numbers when electroporated with equal moles of DNA. Despite similar cell entry, we found length-dependent differences in how efficiently the vectors knocked down GFP. As vector length increased up to 1,869 bp, GFP knockdown efficiency per mole of transfected DNA increased. From 1,869 to 4,257 bp, GFP knockdown efficiency per mole was steady, then decreased with increasing vector length. In comparing GFP knockdown with equal masses of vectors, we found that the shorter vectors transfect more efficiently per nanogram of DNA transfected. Our results rule out cell entry and DNA mass as determining factors for gene knockdown efficiency via electroporation. The length-dependent effects we have uncovered are likely explained by differences in nuclear translocation or transcription. These data add an important step towards clinical applications of non-viral vector delivery. PMID:27918590

Factors Influencing the Production of MFSV Full-Length Clone: Maize Fine Streak Virus Proteins in Drosophila S2 Cells

USDA-ARS?s Scientific Manuscript database

Maize fine streak virus (MFSV) is negative-sense RNA virus member of the genus Nucleorhabdovirus. Our goal is to determine whether Drosophila S2 cells can support the production of a full-length clone of MFSV. We have previously demonstrated that the full-length MFSV nucleoprotein (N) and phosphopro...

The effect of macromolecular crowding on mobility of biomolecules, association kinetics and gene expression in living cells

NASA Astrophysics Data System (ADS)

Tabaka, Marcin; Kalwarczyk, Tomasz; Szymanski, Jedrzej; Hou, Sen; Hołyst, Robert

2014-09-01

We discuss a quantitative influence of macromolecular crowding on biological processes: motion, bimolecular reactions, and gene expression in prokaryotic and eukaryotic cells. We present scaling laws relating diffusion coefficient of an object moving in a cytoplasm of cells to a size of this object and degree of crowding. Such description leads to the notion of the length scale dependent viscosity characteristic for all living cells. We present an application of the length-scale dependent viscosity model to the description of motion in the cytoplasm of both eukaryotic and prokaryotic living cells. We compare the model with all recent data on diffusion of nanoscopic objects in HeLa, and E. coli cells. Additionally a description of the mobility of molecules in cell nucleus is presented. Finally we discuss the influence of crowding on the bimolecular association rates and gene expression in living cells.

RAPAMYCIN INCREASES LENGTH AND MECHANOSENSORY FUNCTION OF PRIMARY CILIA IN RENAL EPITHELIAL AND VASCULAR ENDOTHELIAL CELLS.

PubMed

Sherpa, Rinzhin T; Atkinson, Kimberly F; Ferreira, Viviana P; Nauli, Surya M

2016-12-01

Primary cilia arebiophysically-sensitive organelles responsible for sensing fluid-flow and transducing this stimulus into intracellular responses. Previous studies have shown that the primary cilia mediate flow-induced calcium influx, and sensitivity of cilia function to flow is correlated to cilia length. Cells with abnormal cilia length or function can lead to a host of diseases that are collectively termed as ciliopathies. Rapamycin, a potent inhibitor of mTOR (mammalian target of rapamycin), has been demonstrated to be a potential pharmacological agent against the aberrant mTOR signaling seen in ciliopathies such as polycystic kidney disease (PKD) and tuberous sclerosis complex (TSC). Here we look at the effects of rapamycin on ciliary length and function for the first time. Compared to controls, primary cilia in rapamycin-treated porcine renal epithelial and mouse vascular endothelial cells showed a significant increase in length. Graded increases in fluid-shear stress further indicates that rapamycin enhances cilia sensitivity to fluid flow. Treatment with rapamycin led to G0 arrest in porcine epithelial cells while no significant change in cell cycle were observed in rapamycin-treated mouse epithelial or endothelial cells, indicating a species-specific effect of rapamycin. Given the previousin vitro and in vivo studies establishing rapamycin as a potential therapeutic agent for ciliopathies, such as PKD and TSC, our studies show that rapamycin enhances ciliary function and sensitivity to fluid flow. The results of our studies suggest a potential ciliotherapeutic effect of rapamycin.

Cellular manipulation and patterning using ferromagnetic nanowires

NASA Astrophysics Data System (ADS)

Hultgren, Anne

Ferromagnetic nanowires are demonstrated as an effective tool to apply forces to living cells. Both magnetic cell separations and the magnetic patterning of cells on a substrate will be accomplished through the use of cell-nanowire interactions as well as nanowire-magnetic field interactions. When introduced into cultures of NIH-3T3 cells, the nanowires are internalized by cells via the integrin-mediated adhesion pathway without inflicting any toxic effects on the cell cycle over the course of several days. In addition, the length of the nanowires was found to have an effect on the cell-nanowire interactions when the cells were dissociated from the tissue culture dish. To compare the effectiveness of the nanowires as a means of manipulating cells to the current technology which is based on superparamagnetic beads, magnetic cell separations were performed with electrodeposited Ni nanowires 350 nm in diameter and 5--35 mum long in field gradients of 80 T/m. Single-pass separations of NIH-3T3 cells bound to nanowires achieve up to 81% purity with 85% yield, a dramatic improvement over the 55% purity and 20% yield obtained with the beads. The yield for the separations were found to be dependent on the length of the nanowires, and was maximized when the length of the nanowires equaled the diameter of the cells. This dependence was exploited to perform a size-selective magnetic separation. Substrates containing arrays of micro-magnets, fabricated using photolithography, were placed in cell cultures. These micro-magnet arrays create regions of locally strong magnetic field gradients to trap nanowires in specific locations on the substrate. These substrates were used in conjunction with fluid flow and a weak, externally applied magnetic field to create and control patterns of cells bound to nanowires. Controlled isolation of heterogeneous pairs and groups of cells will enable the study of the biochemistry of cell-cell contacts.

Telomere length variation in tumor cells and cancer-associated fibroblasts: potential biomarker for hepatocellular carcinoma.

PubMed

Ma, Li-Jie; Wang, Xiao-Ying; Duan, Meng; Liu, Long-Zi; Shi, Jie-Yi; Dong, Liang-Qing; Yang, Liu-Xiao; Wang, Zhi-Chao; Ding, Zhen-Bin; Ke, Ai-Wu; Cao, Ya; Zhang, Xiao-Ming; Zhou, Jian; Fan, Jia; Gao, Qiang

2017-12-01

The role of telomere dysfunction and aberrant telomerase activities in hepatocellular carcinoma (HCC) has been overlooked for many years. This study aimed to delineate the variation and prognostic value of telomere length in HCC. Telomere-specific fluorescence in situ hybridization (FISH) and qPCR were used to evaluate telomere length in HCC cell lines, tumor tissues, and isolated non-tumor cells within the tumor. Significant telomere attrition was found in tumor cells and cancer-associated fibroblasts (CAFs) compared to their normal counterparts, but not in intratumor leukocytes or bile duct epithelial cells. Clinical relevance and prognostic value of telomere length were investigated on tissue microarrays of 257 surgically treated HCC patients. Reduced intensity of telomere signals in tumor cells or CAFs correlated with larger tumor size and the presence of vascular invasion (p < 0.05). Shortened telomeres in tumor cells or CAFs associated with reduced survival and increased recurrence, and were identified as independent prognosticators for HCC patients (p < 0.05). These findings were validated in an independent HCC cohort of 371 HCC patients from The Cancer Genome Atlas (TCGA) database, confirming telomere attrition and its prognostic value in HCC. We also showed that telomerase reverse transcriptase promoter (TERTp) mutation correlated with telomere shortening in HCC. Telomere variation in tumor cells and non-tumor cells within the tumor microenvironment of HCC was a valuable prognostic biomarker for this fatal malignancy. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Telomere length profiles in primary human peritoneal mesothelial cells are consistent with senescence.

PubMed

Lopez-Anton, Melisa; Rudolf, András; Baird, Duncan M; Roger, Laureline; Jones, Rhiannon E; Witowski, Janusz; Fraser, Donald J; Bowen, Timothy

2017-06-01

Mesothelial cell (MC) senescence contributes to malignancy and tissue fibrosis. The role of telomere erosion in MC senescence remains controversial, with evidence for both telomere-dependent and telomere-independent mechanisms reported. Single telomere length analysis revealed considerable telomere length heterogeneity in freshly isolated human peritoneal MCs, reflecting a heterogeneous proliferative history and providing high-resolution evidence for telomere-dependent senescence. By contrast the attenuated replicative lifespan, lack of telomere erosion and induction of p16 expression in in vitro-aged cells was consistent with stress-induced senescence. Given the potential pathophysiological impact of senescence in mesothelial tissues, high-resolution MC telomere length analysis may provide clinically useful information. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

Analysis of Average Telomere Length in Human Telomeric Protein Knockout Cells Generated by CRISPR/Cas9.

PubMed

Xu, Jun; Songyang, Zhou; Liu, Dan; Kim, Hyeung

2017-01-01

Telomeres play an important role in ensuring the integrity of the genome. Telomere shortening can lead to loss of genetic information and trigger DNA damage responses. Cultured mammalian cells have served as critical model systems for studying the function of telomere binding proteins and telomerase. Tremendous heterogeneity can be observed both between species and within a single cell population. Recent advances in genome editing (such as the development of the CRISPR/Cas9 platform) have further enabled researchers to carry out loss-of-function analysis of how disrupting key players in telomere maintenance affects telomere length regulation. Here we describe the steps to be carried out in order to analyze the average length of telomeres in CRISPR-engineered human knockout (KO) cells (TRF analysis).

Parameter setting and analysis of a dynamic tubular SOFC model

NASA Astrophysics Data System (ADS)

Jiang, Wei; Fang, Ruixian; Khan, Jamil A.; Dougal, Roger A.

An improved one-dimensional dynamic model of a tubular SOFC stack capable of system simulation in the virtual test bed (VTB) simulation environment is presented in this paper. This model is based on the electrochemical and thermal modeling, accounting for the voltage losses and temperature dynamics. The modeling of an external reformer is also included in this study. A detailed parametric analysis of working conditions and cell configuration of the solid oxide fuel cell (SOFC) stack is the main focus of this paper. The following operating parameters are investigated: pressure ratio, temperature, mass flow rate, external reforming degree and stream to carbon (S/C) ratio. The cell geometric parameters studied include cell diameter and cell length. Elevated operating pressure improves the cell performance. Whereas, higher operating temperature decreases both the Nernst potential and the irreversible losses, resulting in an initial increase then a decrease in cell efficiency. It was found that a higher S/C ratio yields a lower H 2 concentration and partial pressure, which has a negative effect on the Nernst potential. Increased cell diameter is found to increase the power due to a larger activation area at the same time and due to longer current path length there is an increase in the ohmic loss. Increased length of the cell has the undesired affect of an increased pressure drop.

High efficiency solar cells combining a perovskite and a silicon heterojunction solar cells via an optical splitting system

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

Uzu, Hisashi, E-mail: Hisashi.Uzu@kaneka.co.jp, E-mail: npark@skku.edu; Ichikawa, Mitsuru; Hino, Masashi

2015-01-05

We have applied an optical splitting system in order to achieve very high conversion efficiency for a full spectrum multi-junction solar cell. This system consists of multiple solar cells with different band gap optically coupled via an “optical splitter.” An optical splitter is a multi-layered beam splitter with very high reflection in the shorter-wave-length range and very high transmission in the longer-wave-length range. By splitting the incident solar spectrum and distributing it to each solar cell, the solar energy can be managed more efficiently. We have fabricated optical splitters and used them with a wide-gap amorphous silicon (a-Si) solar cellmore » or a CH{sub 3}NH{sub 3}PbI{sub 3} perovskite solar cell as top cells, combined with mono-crystalline silicon heterojunction (HJ) solar cells as bottom cells. We have achieved with a 550 nm cutoff splitter an active area conversion efficiency of over 25% using a-Si and HJ solar cells and 28% using perovskite and HJ solar cells.« less

Determination of lifetimes and recombination currents in p-n junction solar cells, diodes, and transistors

NASA Technical Reports Server (NTRS)

Neugroschel, A.

1981-01-01

New methods are presented and illustrated that enable the accurate determination of the diffusion length of minority carriers in the narrow regions of a solar cell or a diode. Other methods now available are inaccurate for the desired case in which the width of the region is less than the diffusion length. Once the diffusion length is determined by the new methods, this result can be combined with measured dark I-V characteristics and with small-signal admittance characteristics to enable determination of the recombination currents in each quasi-neutral region of the cell - for example, in the emitter, low-doped base, and high-doped base regions of the BSF (back-surface-field) cell. This approach leads to values for the effective surface recombination velocity of the high-low junction forming the back-surface field of BSF cells or the high-low emitter junction of HLE cells. These methods are also applicable for measuring the minority-carrier lifetime in thin epitaxial layers grown on substrates with opposite conductivity type.

Multiple-Path-Length Optical Absorbance Cell

NASA Technical Reports Server (NTRS)

2001-01-01

An optical absorbance cell that offers a selection of multiple optical path lengths has been developed as part of a portable spectrometric instrument that measures absorption spectra of small samples of water and that costs less than does a conventional, non-portable laboratory spectrometer. The instrument is intended, more specifically, for use in studying colored dissolved organic matter (CDOM) in seawater, especially in coastal regions. Accurate characterization of CDOM is necessary for building bio-optical mathematical models of seawater. The multiple path lengths of the absorption cell afford a wide range of sensitivity needed for measuring the optical absorbances associated with the wide range of concentrations of CDOM observed in nature. The instrument operates in the wavelength range of 370 to 725 nm. The major subsystems of the instrument (see figure) include a color-balanced light source; the absorption cell; a peristaltic pump; a high-precision, low-noise fiber optic spectrometer; and a laptop or other personal computer. A fiber-optic cable transmits light from the source to the absorption cell. Other optical fibers transmit light from the absorption cell to the spectrometer,

Genetic Variants Related to Longer Telomere Length are Associated with Increased Risk of Renal Cell Carcinoma.

PubMed

Machiela, Mitchell J; Hofmann, Jonathan N; Carreras-Torres, Robert; Brown, Kevin M; Johansson, Mattias; Wang, Zhaoming; Foll, Matthieu; Li, Peng; Rothman, Nathaniel; Savage, Sharon A; Gaborieau, Valerie; McKay, James D; Ye, Yuanqing; Henrion, Marc; Bruinsma, Fiona; Jordan, Susan; Severi, Gianluca; Hveem, Kristian; Vatten, Lars J; Fletcher, Tony; Koppova, Kvetoslava; Larsson, Susanna C; Wolk, Alicja; Banks, Rosamonde E; Selby, Peter J; Easton, Douglas F; Pharoah, Paul; Andreotti, Gabriella; Freeman, Laura E Beane; Koutros, Stella; Albanes, Demetrius; Mannisto, Satu; Weinstein, Stephanie; Clark, Peter E; Edwards, Todd E; Lipworth, Loren; Gapstur, Susan M; Stevens, Victoria L; Carol, Hallie; Freedman, Matthew L; Pomerantz, Mark M; Cho, Eunyoung; Kraft, Peter; Preston, Mark A; Wilson, Kathryn M; Gaziano, J Michael; Sesso, Howard S; Black, Amanda; Freedman, Neal D; Huang, Wen-Yi; Anema, John G; Kahnoski, Richard J; Lane, Brian R; Noyes, Sabrina L; Petillo, David; Colli, Leandro M; Sampson, Joshua N; Besse, Celine; Blanche, Helene; Boland, Anne; Burdette, Laurie; Prokhortchouk, Egor; Skryabin, Konstantin G; Yeager, Meredith; Mijuskovic, Mirjana; Ognjanovic, Miodrag; Foretova, Lenka; Holcatova, Ivana; Janout, Vladimir; Mates, Dana; Mukeriya, Anush; Rascu, Stefan; Zaridze, David; Bencko, Vladimir; Cybulski, Cezary; Fabianova, Eleonora; Jinga, Viorel; Lissowska, Jolanta; Lubinski, Jan; Navratilova, Marie; Rudnai, Peter; Szeszenia-Dabrowska, Neonila; Benhamou, Simone; Cancel-Tassin, Geraldine; Cussenot, Olivier; Bueno-de-Mesquita, H Bas; Canzian, Federico; Duell, Eric J; Ljungberg, Börje; Sitaram, Raviprakash T; Peters, Ulrike; White, Emily; Anderson, Garnet L; Johnson, Lisa; Luo, Juhua; Buring, Julie; Lee, I-Min; Chow, Wong-Ho; Moore, Lee E; Wood, Christopher; Eisen, Timothy; Larkin, James; Choueiri, Toni K; Lathrop, G Mark; Teh, Bin Tean; Deleuze, Jean-Francois; Wu, Xifeng; Houlston, Richard S; Brennan, Paul; Chanock, Stephen J; Scelo, Ghislaine; Purdue, Mark P

2017-11-01

Relative telomere length in peripheral blood leukocytes has been evaluated as a potential biomarker for renal cell carcinoma (RCC) risk in several studies, with conflicting findings. We performed an analysis of genetic variants associated with leukocyte telomere length to assess the relationship between telomere length and RCC risk using Mendelian randomization, an approach unaffected by biases from temporal variability and reverse causation that might have affected earlier investigations. Genotypes from nine telomere length-associated variants for 10 784 cases and 20 406 cancer-free controls from six genome-wide association studies (GWAS) of RCC were aggregated into a weighted genetic risk score (GRS) predictive of leukocyte telomere length. Odds ratios (ORs) relating the GRS and RCC risk were computed in individual GWAS datasets and combined by meta-analysis. Longer genetically inferred telomere length was associated with an increased risk of RCC (OR=2.07 per predicted kilobase increase, 95% confidence interval [CI]:=1.70-2.53, p<0.0001). As a sensitivity analysis, we excluded two telomere length variants in linkage disequilibrium (R 2 >0.5) with GWAS-identified RCC risk variants (rs10936599 and rs9420907) from the telomere length GRS; despite this exclusion, a statistically significant association between the GRS and RCC risk persisted (OR=1.73, 95% CI=1.36-2.21, p<0.0001). Exploratory analyses for individual histologic subtypes suggested comparable associations with the telomere length GRS for clear cell (N=5573, OR=1.93, 95% CI=1.50-2.49, p<0.0001), papillary (N=573, OR=1.96, 95% CI=1.01-3.81, p=0.046), and chromophobe RCC (N=203, OR=2.37, 95% CI=0.78-7.17, p=0.13). Our investigation adds to the growing body of evidence indicating some aspect of longer telomere length is important for RCC risk. Telomeres are segments of DNA at chromosome ends that maintain chromosomal stability. Our study investigated the relationship between genetic variants associated with telomere length and renal cell carcinoma risk. We found evidence suggesting individuals with inherited predisposition to longer telomere length are at increased risk of developing renal cell carcinoma. Published by Elsevier B.V.

Running rescues defective adult neurogenesis by shortening the length of the cell cycle of neural stem and progenitor cells.

PubMed

Farioli-Vecchioli, Stefano; Mattera, Andrea; Micheli, Laura; Ceccarelli, Manuela; Leonardi, Luca; Saraulli, Daniele; Costanzi, Marco; Cestari, Vincenzo; Rouault, Jean-Pierre; Tirone, Felice

2014-07-01

Physical exercise increases the generation of new neurons in adult neurogenesis. However, only few studies have investigated the beneficial effects of physical exercise in paradigms of impaired neurogenesis. Here, we demonstrate that running fully reverses the deficient adult neurogenesis within the hippocampus and subventricular zone of the lateral ventricle, observed in mice lacking the antiproliferative gene Btg1. We also evaluated for the first time how running influences the cell cycle kinetics of stem and precursor subpopulations of wild-type and Btg1-null mice, using a new method to determine the cell cycle length. Our data show that in wild-type mice running leads to a cell cycle shortening only of NeuroD1-positive progenitor cells. In contrast, in Btg1-null mice, physical exercise fully reactivates the defective hippocampal neurogenesis, by shortening the S-phase length and the overall cell cycle duration of both neural stem (glial fibrillary acidic protein(+) and Sox2(+)) and progenitor (NeuroD1(+)) cells. These events are sufficient and necessary to reactivate the hyperproliferation observed in Btg1-null early-postnatal mice and to expand the pool of adult neural stem and progenitor cells. Such a sustained increase of cell proliferation in Btg1-null mice after running provides a long-lasting increment of proliferation, differentiation, and production of newborn neurons, which rescues the impaired pattern separation previously identified in Btg1-null mice. This study shows that running positively affects the cell cycle kinetics of specific subpopulations of newly generated neurons and suggests that the plasticity of neural stem cells without cell cycle inhibitory control is reactivated by running, with implications for the long-term modulation of neurogenesis. © 2014 AlphaMed Press.

The Length Distribution of Class I-Restricted T Cell Epitopes Is Determined by Both Peptide Supply and MHC Allele-Specific Binding Preference.

PubMed

Trolle, Thomas; McMurtrey, Curtis P; Sidney, John; Bardet, Wilfried; Osborn, Sean C; Kaever, Thomas; Sette, Alessandro; Hildebrand, William H; Nielsen, Morten; Peters, Bjoern

2016-02-15

HLA class I-binding predictions are widely used to identify candidate peptide targets of human CD8(+) T cell responses. Many such approaches focus exclusively on a limited range of peptide lengths, typically 9 aa and sometimes 9-10 aa, despite multiple examples of dominant epitopes of other lengths. In this study, we examined whether epitope predictions can be improved by incorporating the natural length distribution of HLA class I ligands. We found that, although different HLA alleles have diverse length-binding preferences, the length profiles of ligands that are naturally presented by these alleles are much more homogeneous. We hypothesized that this is due to a defined length profile of peptides available for HLA binding in the endoplasmic reticulum. Based on this, we created a model of HLA allele-specific ligand length profiles and demonstrate how this model, in combination with HLA-binding predictions, greatly improves comprehensive identification of CD8(+) T cell epitopes. Copyright © 2016 by The American Association of Immunologists, Inc.

Development and validation of novel AAV2 random libraries displaying peptides of diverse lengths and at diverse capsid positions.

PubMed

Naumer, Matthias; Ying, Ying; Michelfelder, Stefan; Reuter, Antje; Trepel, Martin; Müller, Oliver J; Kleinschmidt, Jürgen A

2012-05-01

Libraries based on the insertion of random peptide ligands into the capsid of adeno-associated virus type 2 (AAV2) have been widely used to improve the efficiency and selectivity of the AAV vector system. However, so far only libraries of 7-mer peptide ligands have been inserted at one well-characterized capsid position. Here, we expanded the combinatorial AAV2 display system to a panel of novel AAV libraries, displaying peptides of 5, 7, 12, 19, or 26 amino acids in length at capsid position 588 or displaying 7-mer peptides at position 453, the most prominently exposed region of the viral capsid. Library selections on two unrelated cell types-human coronary artery endothelial cells and rat cardiomyoblasts-revealed the isolation of cell type-characteristic peptides of different lengths mediating strongly improved target-cell transduction, except for the 26-mer peptide ligands. Characterization of vector selectivity by transduction of nontarget cells and comparative gene-transduction analysis using a panel of 44 human tumor cell lines revealed that insertion of different-length peptides allows targeting of distinct cellular receptors for cell entry with similar efficiency, but with different selectivity. The application of such novel AAV2 libraries broadens the spectrum of targetable receptors by capsid-modified AAV vectors and provides the opportunity to choose the best suited targeting ligand for a certain application from a number of different candidates.

Temperature dependence of damage coefficient in electron irradiated solar cells

NASA Technical Reports Server (NTRS)

Faith, T. J.

1973-01-01

Measurements of light-generated current vs cell temperature on electron-irradiated n/p silicon solar cells show the temperature coefficient of this current to increase with increasing fluence for both 10-ohm and 20-ohm cells. A relationship between minority-carrier diffusion length and light-generated current was derived by combining measurements of these two parameters: vs fluence at room temperature, and vs cell temperature in cells irradiated to a fluence of 1 x 10 to the 15th power e/sq cm. This relationship was used, together with the light-generated current data, to calculate the temperature dependence of the diffusion-length damage coefficient. The results show a strong decrease in the damage coefficient with increasing temperature in the range experienced by solar panels in synchronous earth orbit.

Distinct Responses of Stem Cells to Telomere Uncapping-A Potential Strategy to Improve the Safety of Cell Therapy.

PubMed

Liu, Chang Ching; Ma, Dong Liang; Yan, Ting-Dong; Fan, XiuBo; Poon, Zhiyong; Poon, Lai-Fong; Goh, Su-Ann; Rozen, Steve G; Hwang, William Ying Khee; Tergaonkar, Vinay; Tan, Patrick; Ghosh, Sujoy; Virshup, David M; Goh, Eyleen L K; Li, Shang

2016-10-01

In most human somatic cells, the lack of telomerase activity results in progressive telomere shortening during each cell division. Eventually, DNA damage responses triggered by critically short telomeres induce an irreversible cell cycle arrest termed replicative senescence. However, the cellular responses of human pluripotent stem cells to telomere uncapping remain unknown. We generated telomerase knockout human embryonic stem (ES) cells through gene targeting. Telomerase inactivation in ES cells results in progressive telomere shortening. Telomere DNA damage in ES cells and neural progenitor cells induces rapid apoptosis when telomeres are uncapped, in contrast to fibroblast cells that enter a state of replicative senescence. Significantly, telomerase inactivation limits the proliferation capacity of human ES cells without affecting their pluripotency. By targeting telomerase activity, we can functionally separate the two unique properties of human pluripotent stem cells, namely unlimited self-renewal and pluripotency. We show that the potential of ES cells to form teratomas in vivo is dictated by their telomere length. By controlling telomere length of ES cells through telomerase inactivation, we can inhibit teratoma formation and potentially improve the safety of cell therapies involving terminally differentiated cells as well as specific progenitor cells that do not require sustained cellular proliferation in vivo, and thus sustained telomerase activity. Stem Cells 2016;34:2471-2484. © 2016 AlphaMed Press.

Collective dynamics of cell migration and cell rearrangements

NASA Astrophysics Data System (ADS)

Kabla, Alexandre

Understanding multicellular processes such as embryo development or cancer metastasis requires to decipher the contributions of local cell autonomous behaviours and long range interactions with the tissue environment. A key question in this context concerns the emergence of large scale coordination in cell behaviours, a requirement for collective cell migration or convergent extension. I will present a few examples where physical and mechanical aspects play a significant role in driving tissue scale dynamics.

Soybean cell enlargement oscillates with a temperature-compensated period length of ca. 24 min

NASA Technical Reports Server (NTRS)

Morre, D. J.; Pogue, R.; Morre, D. M.

2001-01-01

Rate of enlargement of epidermal cells from soybean, when measured at intervals of 1 min using a light microscope equipped with a video measurement system, oscillated with a period length of about 24 min. This oscillation parallels the 24-min periodicity observed for the oxidation of NADH by the external plasma membrane NADH oxidase. The increase in length was not only non-linear, but intervals of rapid increase in area alternated with intervals of rapid decrease in area. The length of the period was temperature compensated, and was approximately the same when measured at 14, 24 and 34 degrees C even though the rate of cell enlargement varied over this same range of temperatures. These observations represent the first demonstration of an oscillatory growth behavior correlated with a biochemical activity where the period length of both is independent of temperature (temperature compensated) as is the hallmark of clock-related biological phenomena.

Tumour-cell apoptosis after cisplatin treatment is not telomere dependent.

PubMed

Jeyapalan, Jessie C; Saretzki, Gabriele; Leake, Alan; Tilby, Michael J; von Zglinicki, Thomas

2006-06-01

Cisplatin is a major chemotherapeutic agent, especially for the treatment of neuroblastoma. Telomeres with their sequence (TTAGGG)n are probable targets for cisplatin intrastrand cross-linking, but the role of telomeres in mediating cisplatin cytotoxicity is not clear. After exposure to cisplatin as single dose or continuous treatment, we found no loss of telomeres in either SHSY5Y neuroblastoma cells (telomere length, approximately 4 kbp), HeLa 229 cells (telomere length, 20 kbp) or in the acute lymphoblastic T cell line 1301 (telomere length, approximately 80 kbp). There was no induction of telomeric single strand breaks, telomeric overhangs were not degraded and telomerase activity was down-regulated only after massive onset of apoptosis. In contrast, cisplatin induced a delayed formation of DNA strand breaks and induced DNA damage foci containing gamma-H2A.X at nontelomeric sites. Interstitial DNA damage appears to be more important than telomere loss or telomeric damage as inducer of the signal pathway towards apoptosis and/or growth arrest in cisplatin-treated tumour cells.

Design of landfill daily cells.

PubMed

Panagiotakopoulos, D; Dokas, I

2001-08-01

The objective of this paper is to study the behaviour of the landfill soil-to-refuse (S/R) ratio when size, geometry and operating parameters of the daily cell vary over realistic ranges. A simple procedure is presented (1) for calculating the cell parameters values which minimise the S/R ratio and (2) for studying the sensitivity of this minimum S/R ratio to variations in cell size, final refuse density, working face length, lift height and cover thickness. In countries where daily soil cover is required, savings in landfill space could be realised following this procedure. The sensitivity of minimum S/R to variations in cell dimensions decreases with cell size. Working face length and lift height affect the S/R ratio significantly. This procedure also offers the engineer an additional tool for comparing one large daily cell with two or more smaller ones, at two different working faces within the same landfill.

The carboxyl tail of connexin32 regulates gap junction assembly in human prostate and pancreatic cancer cells.

PubMed

Katoch, Parul; Mitra, Shalini; Ray, Anuttoma; Kelsey, Linda; Roberts, Brett J; Wahl, James K; Johnson, Keith R; Mehta, Parmender P

2015-02-20

Connexins, the constituent proteins of gap junctions, are transmembrane proteins. A connexin (Cx) traverses the membrane four times and has one intracellular and two extracellular loops with the amino and carboxyl termini facing the cytoplasm. The transmembrane and the extracellular loop domains are highly conserved among different Cxs, whereas the carboxyl termini, often called the cytoplasmic tails, are highly divergent. We have explored the role of the cytoplasmic tail of Cx32, a Cx expressed in polarized and differentiated cells, in regulating gap junction assembly. Our results demonstrate that compared with the full-length Cx32, the cytoplasmic tail-deleted Cx32 is assembled into small gap junctions in human pancreatic and prostatic cancer cells. Our results further document that the expression of the full-length Cx32 in cells, which express the tail-deleted Cx32, increases the size of gap junctions, whereas the expression of the tail-deleted Cx32 in cells, which express the full-length Cx32, has the opposite effect. Moreover, we show that the tail is required for the clustering of cell-cell channels and that in cells expressing the tail-deleted Cx32, the expression of cell surface-targeted cytoplasmic tail alone is sufficient to enhance the size of gap junctions. Our live-cell imaging data further demonstrate that gap junctions formed of the tail-deleted Cx32 are highly mobile compared with those formed of full-length Cx32. Our results suggest that the cytoplasmic tail of Cx32 is not required to initiate the assembly of gap junctions but for their subsequent growth and stability. Our findings suggest that the cytoplasmic tail of Cx32 may be involved in regulating the permeability of gap junctions by regulating their size. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Impact of Tumor Cell Cytoskeleton Organization on Invasiveness and Migration: A Microchannel-Based Approach

PubMed Central

Rolli, Claudio G.; Seufferlein, Thomas; Kemkemer, Ralf; Spatz, Joachim P.

2010-01-01

Cell migration is a fundamental feature of the interaction of cells with their surrounding. The cell's stiffness and ability to deform itself are two major characteristics that rule migration behavior especially in three-dimensional tissue. We simulate this situation making use of a micro-fabricated migration chip to test the active invasive behavior of pancreatic cancer cells (Panc-1) into narrow channels. At a channel width of 7 µm cell migration through the channels was significantly impeded due to size exclusion. A striking increase in cell invasiveness was observed once the cells were treated with the bioactive lipid sphingosylphosphorylcholine (SPC) that leads to a reorganization of the cell's keratin network, an enhancement of the cell's deformability, and also an increase in the cell's migration speed on flat surfaces. The migration speed of the highly deformed cells inside the channels was three times higher than of cells on flat substrates but was not affected upon SPC treatment. Cells inside the channels migrated predominantly by smooth sliding while maintaining constant cell length. In contrast, cells on adhesion mediating narrow lines moved in a stepwise way, characterized by fluctuations in cell length. Taken together, with our migration chip we demonstrate that the dimensionality of the environment strongly affects the migration phenotype and we suggest that the spatial cytoskeletal keratin organization correlates with the tumor cell's invasive potential. PMID:20090950

Application of the SEM to the measurement of solar cell parameters

NASA Technical Reports Server (NTRS)

Weizer, V. G.; Andrews, C. W.

1977-01-01

A pair of techniques are described which make use of the SEM to measure, respectively, the minority carrier diffusion length and the metallurgical junction depth in silicon solar cells. The former technique permits the measurement of the true bulk diffusion length through the application of highly doped field layers to the back surfaces of the cells being investigated. The technique yields an absolute value of the diffusion length from a knowledge of the collected fraction of the injected carriers and the cell thickness. It is shown that the secondary emission contrast observed in the SEM on a reverse-biased diode can depict the location of the metallurgical junction if the diode has been prepared with the proper beveled geometry. The SEM provides the required contrast and the option of high magnification, permitting the measurement of extremely shallow junction depths.

Purification of full-length VP22 from cells infected with HSV-1: A two-pronged approach for the solubilization and purification of viral proteins for use in biochemical studies

PubMed Central

Dewberry, Ebony J.; Dunkerley, Eric; Duffy, Carol

2012-01-01

Summary VP22, encoded by the UL49 gene, is one of the most abundant proteins of the herpes simplex virus type 1 (HSV-1) tegument and has been shown to be important for virus replication and spread. However, the exact role(s) played by VP22 in the HSV-1 replication cycle have yet to be delineated. The lack of a procedure to purify full-length VP22 has limited molecular studies on VP22 function. A procedure was developed for the purification of soluble, full-length VP22 from cells infected with HSV-1. A recombinant virus encoding His-tagged VP22 was generated and found to express VP22 at levels comparable to the wild type virus upon infection of Vero cells. By experimenting with a wide variety of cell lysis buffer conditions, several buffers that promote the solubility of full-length VP22 were identified. Buffers that gave the highest levels of solubility were then used in immobilized metal ion affinity chromatography experiments to identify conditions that provided the greatest level of VP22 binding and recovery from cobalt and nickel affinity resins. Using this strategy soluble, full-length VP22 was purified from cells infected with HSV-1. PMID:22569534

Abelson tyrosine-protein kinase 2 regulates myoblast proliferation and controls muscle fiber length

PubMed Central

Lee, Jennifer K; Hallock, Peter T

2017-01-01

Muscle fiber length is nearly uniform within a muscle but widely different among different muscles. We show that Abelson tyrosine-protein kinase 2 (Abl2) has a key role in regulating myofiber length, as a loss of Abl2 leads to excessively long myofibers in the diaphragm, intercostal and levator auris muscles but not limb muscles. Increased myofiber length is caused by enhanced myoblast proliferation, expanding the pool of myoblasts and leading to increased myoblast fusion. Abl2 acts in myoblasts, but as a consequence of expansion of the diaphragm muscle, the diaphragm central tendon is reduced in size, likely contributing to reduced stamina of Abl2 mutant mice. Ectopic muscle islands, each composed of myofibers of uniform length and orientation, form within the central tendon of Abl2+/− mice. Specialized tendon cells, resembling tendon cells at myotendinous junctions, form at the ends of these muscle islands, suggesting that myofibers induce differentiation of tendon cells, which reciprocally regulate myofiber length and orientation. PMID:29231808

Abelson tyrosine-protein kinase 2 regulates myoblast proliferation and controls muscle fiber length.

PubMed

Lee, Jennifer K; Hallock, Peter T; Burden, Steven J

2017-12-12

Muscle fiber length is nearly uniform within a muscle but widely different among different muscles. We show that Abelson tyrosine-protein kinase 2 (Abl2) has a key role in regulating myofiber length, as a loss of Abl2 leads to excessively long myofibers in the diaphragm, intercostal and levator auris muscles but not limb muscles. Increased myofiber length is caused by enhanced myoblast proliferation, expanding the pool of myoblasts and leading to increased myoblast fusion. Abl2 acts in myoblasts, but as a consequence of expansion of the diaphragm muscle, the diaphragm central tendon is reduced in size, likely contributing to reduced stamina of Abl2 mutant mice. Ectopic muscle islands, each composed of myofibers of uniform length and orientation, form within the central tendon of Abl2 +/- mice. Specialized tendon cells, resembling tendon cells at myotendinous junctions, form at the ends of these muscle islands, suggesting that myofibers induce differentiation of tendon cells, which reciprocally regulate myofiber length and orientation.

Physics of active jamming during collective cellular motion in a monolayer.

PubMed

Garcia, Simon; Hannezo, Edouard; Elgeti, Jens; Joanny, Jean-François; Silberzan, Pascal; Gov, Nir S

2015-12-15

Although collective cell motion plays an important role, for example during wound healing, embryogenesis, or cancer progression, the fundamental rules governing this motion are still not well understood, in particular at high cell density. We study here the motion of human bronchial epithelial cells within a monolayer, over long times. We observe that, as the monolayer ages, the cells slow down monotonously, while the velocity correlation length first increases as the cells slow down but eventually decreases at the slowest motions. By comparing experiments, analytic model, and detailed particle-based simulations, we shed light on this biological amorphous solidification process, demonstrating that the observed dynamics can be explained as a consequence of the combined maturation and strengthening of cell-cell and cell-substrate adhesions. Surprisingly, the increase of cell surface density due to proliferation is only secondary in this process. This analysis is confirmed with two other cell types. The very general relations between the mean cell velocity and velocity correlation lengths, which apply for aggregates of self-propelled particles, as well as motile cells, can possibly be used to discriminate between various parameter changes in vivo, from noninvasive microscopy data.

Photonic Devices Based on Surface and Composition-Engineered Infrared Colloidal Nanocrystals

DTIC Science & Technology

2012-01-27

NQD/P3HT solar cells , the need for submicron-phase-separated polymer-NQD blends is therefore expressed by the limiting exciton diffusion length ...P3HT:PbSe are very critical in designing the PM-HJ solar cells : The thickness of P3HT should approximate to the thickness of exciton diffuse length in... cells , luminescent solar concentrators, light emitting diodes, lasers, photonic crystals, CdSe, PbSe, Germanium Jian Xu Pennsylvania State University

Formation of rings from segments of HeLa-cell nuclear deoxyribonucleic acid

PubMed Central

Hardman, Norman

1974-01-01

Duplex segments of HeLa-cell nuclear DNA were generated by cleavage with DNA restriction endonuclease from Haemophilus influenzae. About 20–25% of the DNA segments produced, when partly degraded with exonuclease III and annealed, were found to form rings visible in the electron microscope. A further 5% of the DNA segments formed structures that were branched in configuration. Similar structures were generated from HeLa-cell DNA, without prior treatment with restriction endonuclease, when the complementary polynucleotide chains were exposed by exonuclease III action at single-chain nicks. After exposure of an average single-chain length of 1400 nucleotides per terminus at nicks in HeLa-cell DNA by exonuclease III, followed by annealing, the physical length of ring closures was estimated and found to be 0.02–0.1μm, or 50–300 base pairs. An almost identical distribution of lengths was recorded for the regions of complementary base sequence responsible for branch formation. It is proposed that most of the rings and branches are formed from classes of reiterated base sequence with an average length of 180 base pairs arranged intermittenly in HeLa-cell DNA. From the rate of formation of branched structures when HeLa-cell DNA segments were heat-denatured and annealed, it is estimated that the reiterated sequences are in families containing approximately 2400–24000 copies. ImagesPLATE 2PLATE 1 PMID:4462738

Aphidicolin-induced nuclear elongation in tobacco BY-2 cells.

PubMed

Yasuhara, Hiroki; Kitamoto, Kazuki

2014-05-01

Plant nuclei are known to differentiate into various shapes within a single plant. However, little is known about the mechanisms of nuclear morphogenesis. We found that nuclei of tobacco BY-2 cells were highly elongated on long-term treatment with 5 mg l⁻¹ aphidicolin, an inhibitor of DNA polymerase α. In aphidicolin-treated cells, the nuclear length was correlated with the cell length. During culture in the presence of aphidicolin, the nuclei were elongated in parallel with cell elongation. Nuclear elongation was inhibited by the inhibition of cell elongation with 2,6-dichlorobenzonitrile, a cellulose synthesis inhibitor. However, cell elongation induced in the auxin-depleted medium in the absence of aphidicolin did not cause nuclear elongation, indicating that cell elongation alone is not sufficient for nuclear elongation. Treatment with either latrunculin B or propyzamide inhibited the aphidicolin-induced nuclear elongation, indicating that both actin filaments and microtubules (MTs) are required for nuclear elongation. Observations using BY-YTHCLR2 cells, in which actin filaments, MTs and nuclei were simultaneously visualized, revealed that the longitudinally arranged MT bundles associated with the nucleus play an important role in nuclear elongation, and that actin filaments affect the formation of these MT bundles. In aphidicolin-treated cells, the nuclear DNA contents of the elongated nuclei exceeded 4C, and the nuclear length was highly correlated with the nuclear DNA content. In cells treated with 50 mg l⁻¹ aphidicolin, cells were elongated and nucleus-associated longitudinal MT bundles were formed, but the nuclear DNA contents did not exceed 4C and the nuclei did not elongate. These results indicate that an increase in the nuclear DNA content above 4C is also required for nuclear elongation.

Tankyrase 2 Poly(ADP-Ribose) Polymerase Domain-Deleted Mice Exhibit Growth Defects but Have Normal Telomere Length and Capping

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

Hsiao, Susan J; Poitras, Marc; Cook, Brandoch

Regulation of telomere length maintenance and capping are a critical cell functions in both normal and tumor cells. Tankyrase 2 (Tnks2) is a poly(ADP-ribose) polymerase (PARP) that has been shown to modify itself and TRF1, a telomere-binding protein. We show here by overexpression studies that tankyrase 2, like its closely related homolog tankyrase 1, can function as a positive regulator of telomere length in human cells, dependent on its catalytic PARP activity. To study the role of Tnks2 in vivo, we generated mice with the Tnks2 PARP domain deleted. These mice are viable and fertile but display a growth retardationmore » phenotype. Telomere analysis by quantitative fluorescence in situ hybridization (FISH), flow-FISH, and restriction fragment analysis showed no change in telomere length or telomere capping in these mice. To determine the requirement foTnks2 in long-term maintenance of telomeres, we generated embryonic stem cells with the Tnks2 PARP domain deleted and observed no change, even upon prolonged growth, in telomere length or telomere capping. Together these results suggest that Tnkjs2 has a role in normal growth and development but is not essential for telomere length maintenance or telomere capping in mice.« less

Analysis of the age of Panax ginseng based on telomere length and telomerase activity.

PubMed

Liang, Jiabei; Jiang, Chao; Peng, Huasheng; Shi, Qinghua; Guo, Xiang; Yuan, Yuan; Huang, Luqi

2015-01-23

Ginseng, which is the root of Panax ginseng (Araliaceae), has been used in Oriental medicine as a stimulant and dietary supplement for more than 7,000 years. Older ginseng plants are substantially more medically potent, but ginseng age can be simulated using unscrupulous cultivation practices. Telomeres progressively shorten with each cell division until they reach a critical length, at which point cells enter replicative senescence. However, in some cells, telomerase maintains telomere length. In this study, to determine whether telomere length reflects ginseng age and which tissue is best for such an analysis, we examined telomerase activity in the main roots, leaves, stems, secondary roots and seeds of ginseng plants of known age. Telomere length in the main root (approximately 1 cm below the rhizome) was found to be the best indicator of age. Telomeric terminal restriction fragment (TRF) lengths, which are indicators of telomere length, were determined for the main roots of plants of different ages through Southern hybridization analysis. Telomere length was shown to be positively correlated with plant age, and a simple mathematical model was formulated to describe the relationship between telomere length and age for P. ginseng.

Fabrication of biomimetic nanomaterials and their effect on cell behavior

NASA Astrophysics Data System (ADS)

Porri, Teresa Jane

Cells in vivo respond to an intricate combination of chemical and mechanical signals. The corneal epithelium, a structure which prevents the admission of bacteria and undesirable molecules into the eye, grows on a basement membrane which presents both nanoscale topographic and adhesive chemical signals. An effective approach to biomaterials design takes advantage of the synergistic effects of the multiple cellular inputs which are available to engineer cell-substrate interactions. We have previously demonstrated the effects of nanoscale topography on a variety of corneal epithelial cell behaviors. To gain a better understanding of cell-level control in vivo, we employ a systems-level approach which looks at the effect of nanoscale topography in conjunction with a biomimetic surface chemistry. First, we discuss a novel method of fabricating nanoscale topography through templated electroless deposition of gold into PVP-coated polycarbonate membranes. This technique creates nanowires of gold with an uniform outer diameter that is dependent upon the size of the pores in the membrane used, and a nanowire length that is dependent upon the extent of etching into the polymer membrane. The gold nanowires can be modified with self-assembled monolayers (SAMs) of alkanethiols. Using these substrates, we study the effect of topographic length scale and surface chemistry on cells attached to a discontinuous nanoscale topography, and find a transition in cellular behavior at a length scale (between 600 and 2000 nm inter-wire spacing) that is commensurate with the transition length scale seen on surfaces presenting continuous grooves and ridges. Secondly, we study the effect of non-fouling peptide-modified SAMs on cellular behavior. We examine the effect of co-presented RGD and AG73 peptides and show that cell spreading is a function of the relative ratios of RGD and AG73 present on the surface. Finally, we explore the combinatorial effects of biologically relevant chemistry with anisotropic nanoscale topography with dimensions that vary from the micron to the nanoscale. We show that integrin binding, syndecan binding, and topographic length scale each independently influence epithelial cell response to nanoscale features, lending a high degree of control over cell morphologic responses.

First descriptions of dicyemid mesozoans (Dicyemida: Dicyemidae) from Australian octopus (Octopodidae) and cuttlefish (Sepiidae), including a new record of Dicyemennea in Australian waters.

PubMed

Catalano, Sarah R

2013-09-01

Three new species of dicyemid mesozoans are described for the first time from Australian octopus and cuttlefish species. Dicyemennea floscephalum sp. n. is described from Octopus berrima Stranks et Norman (southern keeled octopus) collected from Spencer Gulf and Gulf St. Vincent, South Australia, Australia and represents the first description of a species of Dicyemennea Whitman, 1883 from Australian waters. Dicyema papuceum sp. n. and D. furuyi sp. n. are described from Sepia papuensis Hoyle (Papuan cuttlefish) collected from Shark Bay, Western Australia, Australia. Dicyemennea floscephalum sp. n. is a medium to large species that reaches approximately 4.9 mm in length. The vermiform stages are characterised by having 23-28 peripheral cells, and a disc-shaped, flower-like calotte in larger individuals. An anterior abortive axial cell is absent in vermiform embryos and verruciform cells were not observed in nematogens and rhombogens. Infusoriform embryos comprise 37 cells; one nucleus is present in each urn cell. Dicyema papuceum sp. n. is a small species that reaches approximately 1.1 mm in length. The vermiform stages are characterised by having 30-33 peripheral cells and a relatively small, cap-shaped calotte. An anterior abortive axial cell is absent in vermiform embryos and verruciform cells were occasionally observed in nematogens. Infusoriform embryos comprise 37 cells; two nuclei are present in each urn cell. Dicyema furuyi sp. n. is a large species that reaches approximately 5.3 mm in length. The vermiform stages are characterised by having 22-24 peripheral cells and an elongate calotte. An anterior abortive axial cell is absent in vermiform embryos and verruciform cells were not observed in nematogens and rhombogens. Infusoriform embryos comprise 37 cells; one nucleus is present in each urn cell. Three secondary nematogens were also observed in the right renal appendages of two host individuals, confirming the occurrence of this form.

Effects of mutations in the Arabidopsis Cold Shock Domain Protein 3 (AtCSP3) gene on leaf cell expansion.

PubMed

Yang, Yongil; Karlson, Dale

2012-08-01

The cold shock domain is among the most evolutionarily conserved nucleic acid binding domains from prokaryotes to higher eukaryotes, including plants. Although eukaryotic cold shock domain proteins have been extensively studied as transcriptional and post-transcriptional regulators during various developmental processes, their functional roles in plants remains poorly understood. In this study, AtCSP3 (At2g17870), which is one of four Arabidopsis thaliana c old s hock domain proteins (AtCSPs), was functionally characterized. Quantitative RT-PCR analysis confirmed high expression of AtCSP3 in reproductive and meristematic tissues. A homozygous atcsp3 loss-of-function mutant exhibits an overall reduced seedling size, stunted and orbicular rosette leaves, reduced petiole length, and curled leaf blades. Palisade mesophyll cells are smaller and more circular in atcsp3 leaves. Cell size analysis indicated that the reduced size of the circular mesophyll cells appears to be generated by a reduction of cell length along the leaf-length axis, resulting in an orbicular leaf shape. It was also determined that leaf cell expansion is impaired for lateral leaf development in the atcsp3 loss-of-function mutant, but leaf cell proliferation is not affected. AtCSP3 loss-of-function resulted in a dramatic reduction of LNG1 transcript, a gene that is involved in two-dimensional leaf polarity regulation. Transient subcellular localization of AtCSP3 in onion epidermal cells confirmed a nucleocytoplasmic localization pattern. Collectively, these data suggest that AtCSP3 is functionally linked to the regulation of leaf length by affecting LNG1 transcript accumulation during leaf development. A putative function of AtCSP3 as an RNA binding protein is also discussed in relation to leaf development.

Trough Coating Solar Cells Without Spillover

NASA Technical Reports Server (NTRS)

Heaps, J. D.

1986-01-01

Problem with trough coating of silicon on ceramic - spillover of molten silicon - overcome by combination of redesigned heaters and tiltable trough. Modifications make it possible to coat virtually any length of ceramic with film of solar-cell-grade silicon. Previously, maximum length coated before spillover occurred was 2 inches (5.1 cm).

Sonic hedgehog controls growth of external genitalia by regulating cell cycle kinetics

PubMed Central

Seifert, Ashley W.; Zheng, Zhengui; Ormerod, Brandi K.; Cohn, Martin J.

2010-01-01

During embryonic development, cells are instructed which position to occupy, they interpret these cues as differentiation programmes, and expand these patterns by growth. Sonic hedgehog (Shh) specifies positional identity in many organs; however, its role in growth is not well understood. In this study, we show that inactivation of Shh in external genitalia extends the cell cycle from 8.5 to 14.4 h, and genital growth is reduced by ∼75%. Transient Shh signalling establishes pattern in the genital tubercle; however, transcriptional levels of G1 cell cycle regulators are reduced. Consequently, G1 length is extended, leading to fewer progenitor cells entering S-phase. Cell cycle genes responded similarly to Shh inactivation in genitalia and limbs, suggesting that Shh may regulate growth by similar mechanisms in different organ systems. The finding that Shh regulates cell number by controlling the length of specific cell cycle phases identifies a novel mechanism by which Shh elaborates pattern during appendage development. PMID:20975695

Crystal growth for high-efficiency silicon solar cells workshop: Summary

NASA Technical Reports Server (NTRS)

Dumas, K. A.

1985-01-01

The state of the art in the growth of silicon crystals for high-efficiency solar cells are reviewed, sheet requirements are defined, and furture areas of research are identified. Silicon sheet material characteristics that limit cell efficiencies and yields were described as well as the criteria for the ideal sheet-growth method. The device engineers wish list to the material engineer included: silicon sheet with long minority carrier lifetime that is uniform throughout the sheet, and which doesn't change during processing; and sheet material that stays flat throughout device processing, has uniform good mechanical strength, and is low cost. Impurities in silicon solar cells depreciate cell performance by reducing diffusion length and degrading junctions. The impurity behavior, degradation mechanisms, and variations in degradation threshold with diffusion length for silicon solar cells were described.

Cell and current collector felt arrangement for solid oxide electrochemical cell combinations

DOEpatents

Reichner, Philip

1988-01-01

A solid electrolyte electrochemical cell combination 1 is made, comprising an annular, axially elongated, inner electrode 2 containing at least one interior gas feed conduit 3; annular solid electrolyte segments 4 around and covering portions of the inner electrode; annular outer electrode segments 6 around and covering portions of the electrolyte segments; electronically conducting, non-porous, interconnection material 5 disposed between electrolyte segments and in contact with the inner electrode, and electronically conducting, porous, metal fiber current collector felts 7 disposed on top of the non-porous interconnect material and outer electrode segments, where both the non-porous interconnect material and the porous metal felts are disposed circumferentially about the cell, transversely to the axial length of the cell and the inner electrode is continuous for the entire axial length of the cell combination.

Characterization of a Setup to test the Impact of High-Amplitude Pressure Waves on Living Cells

PubMed Central

Schmidt, Mischa; Kahlert, Ulf; Wessolleck, Johanna; Maciaczyk, Donata; Merkt, Benjamin; Maciaczyk, Jaroslaw; Osterholz, Jens; Nikkhah, Guido; Steinhauser, Martin O.

2014-01-01

The impact of pressure waves on cells may provide several possible applications in biology and medicine including the direct killing of tumors, drug delivery or gene transfection. In this study we characterize the physical properties of mechanical pressure waves generated by a nanosecond laser pulse in a setup with well-defined cell culture conditions. To systematically characterize the system on the relevant length and time scales (micrometers and nanoseconds) we use photon Doppler velocimetry (PDV) and obtain velocity profiles of the cell culture vessel at the passage of the pressure wave. These profiles serve as input for numerical pressure wave simulations that help to further quantify the pressure conditions on the cellular length scale. On the biological level we demonstrate killing of glioblastoma cells and quantify experimentally the pressure threshold for cell destruction. PMID:24458018

Radiation tolerance of low resistivity, high voltage silicon solar cells

NASA Technical Reports Server (NTRS)

Weizer, V. G.; Weinberg, I.; Swartz, C. K.

1984-01-01

The radiation tolerance of the following three low resistivity, high voltage silicon solar cells was investigated: (1) the COMSAT MSD (multi-step diffused) cell, (2) the MinMIS cell, and (3) the MIND cell. A description of these solar cells is given along with drawings of their configurations. The diffusion length damage coefficients for the cells were calculated and presented. Solar cell spectral response was also discussed. Cells of the MinMIS type were judged to be unsuitable for use in the space radiation environment.

Centrobin-mediated Regulation of the Centrosomal Protein 4.1-associated Protein (CPAP) Level Limits Centriole Length during Elongation Stage*

PubMed Central

Gudi, Radhika; Haycraft, Courtney J.; Bell, P. Darwin; Li, Zihai; Vasu, Chenthamarakshan

2015-01-01

Microtubule-based centrioles in the centrosome mediate accurate bipolar cell division, spindle orientation, and primary cilia formation. Cellular checkpoints ensure that the centrioles duplicate only once in every cell cycle and achieve precise dimensions, dysregulation of which results in genetic instability and neuro- and ciliopathies. The normal cellular level of centrosomal protein 4.1-associated protein (CPAP), achieved by its degradation at mitosis, is considered as one of the major mechanisms that limits centriole growth at a predetermined length. Here we show that CPAP levels and centriole elongation are regulated by centrobin. Exogenous expression of centrobin causes abnormal elongation of centrioles due to massive accumulation of CPAP in the cell. Conversely, CPAP was undetectable in centrobin-depleted cells, suggesting that it undergoes degradation in the absence of centrobin. Only the reintroduction of full-length centrobin, but not its mutant form that lacks the CPAP binding site, could restore cellular CPAP levels in centrobin-depleted cells, indicating that persistence of CPAP requires its interaction with centrobin. Interestingly, inhibition of the proteasome in centrobin-depleted cells restored the cellular and centriolar CPAP expression, suggesting its ubiquitination and proteasome-mediated degradation when centrobin is absent. Intriguingly, however, centrobin-overexpressing cells also showed proteasome-independent accumulation of ubiquitinated CPAP and abnormal, ubiquitin-positive, elongated centrioles. Overall, our results show that centrobin interacts with ubiquitinated CPAP and prevents its degradation for normal centriole elongation function. Therefore, it appears that loss of centrobin expression destabilizes CPAP and triggers its degradation to restrict the centriole length during biogenesis. PMID:25616662

Two new species of dicyemid mesozoans (Dicyemida: Dicyemidae) from Octopus maya Voss & Solis-Ramirez (Octopodidae) off Yucatan, Mexico.

PubMed

Castellanos-Martinez, Sheila; Aguirre-Macedo, M Leopoldina; Furuya, Hidetaka

2016-07-01

Two new dicyemid species are described from the endemic cephalopod Octopus maya Voss & Solis-Ramirez collected off Yucatan, Mexico. The renal sacs of 40 juvenile and adult octopuses from four localities were examined. Dicyema hochbergi n. sp. is a medium-sized species that reaches 2,245 µm in length. The vermiform stages consist of 18-24 peripheral cells, a conical calotte and the extension of the axial cell between the base and middle of the metapolar cells. Infusoriform embryos consist of 39 cells with urn cell containing one germinal cell, two nuclei and solid refringent bodies. Dicyema mexcayae n. sp. is a relatively small species that reaches 1,114 µm in length. The vermiform stages are constituted by 14-16 peripheral cells, an elongate calotte and the axial cell extending forward to the middle of the metapolar cells. The infusoriform embryos consist of 37 cells, two solid refringent bodies and urn cells with two nuclei each. The present study represents the first description of a dicyemid species from O. maya and increases the number of described species from Mexican waters to 11.

Quantifying Electrical Interactions between Cardiomyocytes and Other Cells in Micropatterned Cell Pairs

PubMed Central

Nguyen, Hung; Badie, Nima; McSpadden, Luke; Pedrotty, Dawn; Bursac, Nenad

2014-01-01

Micropatterning is a powerful technique to control cell shape and position on a culture substrate. In this chapter, we describe the method to reproducibly create large numbers of micropatterned heterotypic cell pairs with defined size, shape, and length of cell–cell contact. These cell pairs can be utilized in patch clamp recordings to quantify electrical interactions between cardiomyocytes and non-cardiomyocytes. PMID:25070342

Brg1 coordinates multiple processes during retinogenesis and is a tumor suppressor in retinoblastoma

DOE PAGES

Aldiri, Issam; Ajioka, Itsuki; Xu, Beisi; ...

2015-12-01

Retinal development requires precise temporal and spatial coordination of cell cycle exit, cell fate specification, cell migration and differentiation. When this process is disrupted, retinoblastoma, a developmental tumor of the retina, can form. Epigenetic modulators are central to precisely coordinating developmental events, and many epigenetic processes have been implicated in cancer. Studying epigenetic mechanisms in development is challenging because they often regulate multiple cellular processes; therefore, elucidating the primary molecular mechanisms involved can be difficult. Here we explore the role of Brg1 (Smarca4) in retinal development and retinoblastoma in mice using molecular and cellular approaches. Brg1 was found to regulatemore » retinal size by controlling cell cycle length, cell cycle exit and cell survival during development. Brg1 was not required for cell fate specification but was required for photoreceptor differentiation and cell adhesion/polarity programs that contribute to proper retinal lamination during development. The combination of defective cell differentiation and lamination led to retinal degeneration in Brg1-deficient retinae. Despite the hypocellularity, premature cell cycle exit, increased cell death and extended cell cycle length, retinal progenitor cells persisted in Brg1-deficient retinae, making them more susceptible to retinoblastoma. In conclusion, ChIP-Seq analysis suggests that Brg1 might regulate gene expression through multiple mechanisms.« less

Brg1 coordinates multiple processes during retinogenesis and is a tumor suppressor in retinoblastoma

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

Aldiri, Issam; Ajioka, Itsuki; Xu, Beisi

Retinal development requires precise temporal and spatial coordination of cell cycle exit, cell fate specification, cell migration and differentiation. When this process is disrupted, retinoblastoma, a developmental tumor of the retina, can form. Epigenetic modulators are central to precisely coordinating developmental events, and many epigenetic processes have been implicated in cancer. Studying epigenetic mechanisms in development is challenging because they often regulate multiple cellular processes; therefore, elucidating the primary molecular mechanisms involved can be difficult. Here we explore the role of Brg1 (Smarca4) in retinal development and retinoblastoma in mice using molecular and cellular approaches. Brg1 was found to regulatemore » retinal size by controlling cell cycle length, cell cycle exit and cell survival during development. Brg1 was not required for cell fate specification but was required for photoreceptor differentiation and cell adhesion/polarity programs that contribute to proper retinal lamination during development. The combination of defective cell differentiation and lamination led to retinal degeneration in Brg1-deficient retinae. Despite the hypocellularity, premature cell cycle exit, increased cell death and extended cell cycle length, retinal progenitor cells persisted in Brg1-deficient retinae, making them more susceptible to retinoblastoma. In conclusion, ChIP-Seq analysis suggests that Brg1 might regulate gene expression through multiple mechanisms.« less

Tracking of plus-ends reveals microtubule functional diversity in different cell types

NASA Astrophysics Data System (ADS)

Shaebani, M. Reza; Pasula, Aravind; Ott, Albrecht; Santen, Ludger

2016-07-01

Many cellular processes are tightly connected to the dynamics of microtubules (MTs). While in neuronal axons MTs mainly regulate intracellular trafficking, they participate in cytoskeleton reorganization in many other eukaryotic cells, enabling the cell to efficiently adapt to changes in the environment. We show that the functional differences of MTs in different cell types and regions is reflected in the dynamic properties of MT tips. Using plus-end tracking proteins EB1 to monitor growing MT plus-ends, we show that MT dynamics and life cycle in axons of human neurons significantly differ from that of fibroblast cells. The density of plus-ends, as well as the rescue and catastrophe frequencies increase while the growth rate decreases toward the fibroblast cell margin. This results in a rather stable filamentous network structure and maintains the connection between nucleus and membrane. In contrast, plus-ends are uniformly distributed along the axons and exhibit diverse polymerization run times and spatially homogeneous rescue and catastrophe frequencies, leading to MT segments of various lengths. The probability distributions of the excursion length of polymerization and the MT length both follow nearly exponential tails, in agreement with the analytical predictions of a two-state model of MT dynamics.

Predicting efficiency of solar cells based on transparent conducting electrodes

NASA Astrophysics Data System (ADS)

Kumar, Ankush

2017-01-01

Efficiency of a solar cell is directly correlated with the performance of its transparent conducting electrodes (TCEs) which dictates its two core processes, viz., absorption and collection efficiencies. Emerging designs of a TCE involve active networks of carbon nanotubes, silver nanowires and various template-based techniques providing diverse structures; here, voids are transparent for optical transmittance while the conducting network acts as a charge collector. However, it is still not well understood as to which kind of network structure leads to an optimum solar cell performance; therefore, mostly an arbitrary network is chosen as a solar cell electrode. Herein, we propose a new generic approach for understanding the role of TCEs in determining the solar cell efficiency based on analysis of shadowing and recombination losses. A random network of wires encloses void regions of different sizes and shapes which permit light transmission; two terms, void fraction and equivalent radius, are defined to represent the TCE transmittance and wire spacings, respectively. The approach has been applied to various literature examples and their solar cell performance has been compared. To obtain high-efficiency solar cells, optimum density of the wires and their aspect ratio as well as active layer thickness are calculated. Our findings show that a TCE well suitable for one solar cell may not be suitable for another. For high diffusion length based solar cells, the void fraction of the network should be low while for low diffusion length based solar cells, the equivalent radius should be lower. The network with less wire spacing compared to the diffusion length behaves similar to continuous film based TCEs (such as indium tin oxide). The present work will be useful for architectural as well as material engineering of transparent electrodes for improvisation of solar cell performance.

Inadequate Reference Datasets Biased toward Short Non-epitopes Confound B-cell Epitope Prediction*

PubMed Central

Rahman, Kh. Shamsur; Chowdhury, Erfan Ullah; Sachse, Konrad; Kaltenboeck, Bernhard

2016-01-01

X-ray crystallography has shown that an antibody paratope typically binds 15–22 amino acids (aa) of an epitope, of which 2–5 randomly distributed amino acids contribute most of the binding energy. In contrast, researchers typically choose for B-cell epitope mapping short peptide antigens in antibody binding assays. Furthermore, short 6–11-aa epitopes, and in particular non-epitopes, are over-represented in published B-cell epitope datasets that are commonly used for development of B-cell epitope prediction approaches from protein antigen sequences. We hypothesized that such suboptimal length peptides result in weak antibody binding and cause false-negative results. We tested the influence of peptide antigen length on antibody binding by analyzing data on more than 900 peptides used for B-cell epitope mapping of immunodominant proteins of Chlamydia spp. We demonstrate that short 7–12-aa peptides of B-cell epitopes bind antibodies poorly; thus, epitope mapping with short peptide antigens falsely classifies many B-cell epitopes as non-epitopes. We also show in published datasets of confirmed epitopes and non-epitopes a direct correlation between length of peptide antigens and antibody binding. Elimination of short, ≤11-aa epitope/non-epitope sequences improved datasets for evaluation of in silico B-cell epitope prediction. Achieving up to 86% accuracy, protein disorder tendency is the best indicator of B-cell epitope regions for chlamydial and published datasets. For B-cell epitope prediction, the most effective approach is plotting disorder of protein sequences with the IUPred-L scale, followed by antibody reactivity testing of 16–30-aa peptides from peak regions. This strategy overcomes the well known inaccuracy of in silico B-cell epitope prediction from primary protein sequences. PMID:27189949

Microtubules restrict plastid sedimentation in protonemata of the moss Ceratodon

NASA Technical Reports Server (NTRS)

Schwuchow, J.; Sack, F. D.

1994-01-01

Apical cells of protonemata of the moss Ceratodon purpureus are unusual among plant cells with sedimentation in that only some amyloplasts sediment and these do not fall completely to the bottom of vertical cells. To determine whether the cytoskeleton restricts plastid sedimentation, the effects of amiprophos-methyl (APM) and cytochalasin D (CD) on plastid position were quantified. APM treatments of 30-60 min increased the plastid sedimentation that is normally seen along the length of untreated or control cells. Longer APM treatments often resulted in more dramatic plastid sedimentation, and in some cases almost all plastids sedimented to the lowermost point in the cell. In contrast, the microfilament inhibitor CD did not affect longitudinal plastid sedimentation compared to untreated cells, although it did disturb or eliminate plastid zonation in the tip. These data suggest that microtubules restrict the sedimentation of plastids along the length of the cell and that microtubules are load-bearing for all the plastids in the apical cell. This demonstrates the importance of the cytoskeleton in maintaining organelle position and cell organization against the force of gravity.

Mathematical analysis of the Photovoltage Decay (PVD) method for minority carrier lifetime measurements

NASA Technical Reports Server (NTRS)

Vonroos, O. H.

1982-01-01

When the diffusion length of minority carriers becomes comparable with or larger than the thickness of a p-n junction solar cell, the characteristic decay of the photon-generated voltage results from a mixture of contributions with different time constants. The minority carrier recombination lifetime tau and the time constant l(2)/D, where l is essentially the thickness of the cell and D the minority carrier diffusion length, determine the signal as a function of time. It is shown that for ordinary solar cells (n(+)-p junctions), particularly when the diffusion length L of the minority carriers is larger than the cell thickness l, the excess carrier density decays according to exp (-t/tau-pi(2)Dt/4l(2)), tau being the lifetime. Therefore, tau can be readily determined by the photovoltage decay method once D and L are known.

Cloned cows with short telomeres deliver healthy offspring with normal-length telomeres.

PubMed

Miyashita, Norikazu; Kubo, Yasuaki; Yonai, Miharu; Kaneyama, Kanako; Saito, Norio; Sawai, Ken; Minamihashi, Akira; Suzuki, Toshiyuki; Kojima, Toshiyuki; Nagai, Takashi

2011-10-01

Dolly, the first mammal cloned from a somatic cell, had shorter telomeres than age-matched controls and died at an early age because of disease. To investigate longevity and lifetime performance in cloned animals, we produced cloned cows with short telomeres using oviductal epithelial cells as donor cells. At 5 years of age, despite the presence of short telomeres, all cloned cows delivered multiple healthy offspring following artificial insemination with conventionally processed spermatozoa from noncloned bulls, and their milk production was comparable to that of donor cows. Moreover, this study revealed that the offspring had normal-length telomeres in their leukocytes and major organs. Thus, cloned animals have normal functional germ lines, and therefore germ line function can completely restore telomere lengths in clone gametes by telomerase activity, resulting in healthy offspring with normal-length telomeres.

Superresolution Imaging of Dynamic MreB Filaments in B. subtilis—A Multiple-Motor-Driven Transport?

PubMed Central

Olshausen, Philipp v.; Defeu Soufo, Hervé Joël; Wicker, Kai; Heintzmann, Rainer; Graumann, Peter L.; Rohrbach, Alexander

2013-01-01

The cytoskeletal protein MreB is an essential component of the bacterial cell-shape generation system. Using a superresolution variant of total internal reflection microscopy with structured illumination, as well as three-dimensional stacks of deconvolved epifluorescence microscopy, we found that inside living Bacillus subtilis cells, MreB forms filamentous structures of variable lengths, typically not longer than 1 μm. These filaments move along their orientation and mainly perpendicular to the long bacterial axis, revealing a maximal velocity at an intermediate length and a decreasing velocity with increasing filament length. Filaments move along straight trajectories but can reverse or alter their direction of propagation. Based on our measurements, we provide a mechanistic model that is consistent with all observations. In this model, MreB filaments mechanically couple several motors that putatively synthesize the cell wall, whereas the filaments’ traces mirror the trajectories of the motors. On the basis of our mechanistic model, we developed a mathematical model that can explain the nonlinear velocity length dependence. We deduce that the coupling of cell wall synthesis motors determines the MreB filament transport velocity, and the filament mechanically controls a concerted synthesis of parallel peptidoglycan strands to improve cell wall stability. PMID:24010660

Cell shape can mediate the spatial organization of the bacterial cytoskeleton

NASA Astrophysics Data System (ADS)

Wang, Siyuan; Wingreen, Ned

2013-03-01

The bacterial cytoskeleton guides the synthesis of cell wall and thus regulates cell shape. Since spatial patterning of the bacterial cytoskeleton is critical to the proper control of cell shape, it is important to ask how the cytoskeleton spatially self-organizes in the first place. In this work, we develop a quantitative model to account for the various spatial patterns adopted by bacterial cytoskeletal proteins, especially the orientation and length of cytoskeletal filaments such as FtsZ and MreB in rod-shaped cells. We show that the combined mechanical energy of membrane bending, membrane pinning, and filament bending of a membrane-attached cytoskeletal filament can be sufficient to prescribe orientation, e.g. circumferential for FtsZ or helical for MreB, with the accuracy of orientation increasing with the length of the cytoskeletal filament. Moreover, the mechanical energy can compete with the chemical energy of cytoskeletal polymerization to regulate filament length. Notably, we predict a conformational transition with increasing polymer length from smoothly curved to end-bent polymers. Finally, the mechanical energy also results in a mutual attraction among polymers on the same membrane, which could facilitate tight polymer spacing or bundling. The predictions of the model can be verified through genetic, microscopic, and microfluidic approaches.

Superresolution imaging of dynamic MreB filaments in B. subtilis--a multiple-motor-driven transport?

PubMed

Olshausen, Philipp V; Defeu Soufo, Hervé Joël; Wicker, Kai; Heintzmann, Rainer; Graumann, Peter L; Rohrbach, Alexander

2013-09-03

The cytoskeletal protein MreB is an essential component of the bacterial cell-shape generation system. Using a superresolution variant of total internal reflection microscopy with structured illumination, as well as three-dimensional stacks of deconvolved epifluorescence microscopy, we found that inside living Bacillus subtilis cells, MreB forms filamentous structures of variable lengths, typically not longer than 1 μm. These filaments move along their orientation and mainly perpendicular to the long bacterial axis, revealing a maximal velocity at an intermediate length and a decreasing velocity with increasing filament length. Filaments move along straight trajectories but can reverse or alter their direction of propagation. Based on our measurements, we provide a mechanistic model that is consistent with all observations. In this model, MreB filaments mechanically couple several motors that putatively synthesize the cell wall, whereas the filaments' traces mirror the trajectories of the motors. On the basis of our mechanistic model, we developed a mathematical model that can explain the nonlinear velocity length dependence. We deduce that the coupling of cell wall synthesis motors determines the MreB filament transport velocity, and the filament mechanically controls a concerted synthesis of parallel peptidoglycan strands to improve cell wall stability. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Targeting Prostate Cancer with Bifunctional Modulators of the Androgen Receptor

DTIC Science & Technology

2013-10-01

linker lengths were prepared and assessed to determine the impact of linker length on the activity of the molecules. HEK293T cells were transfected as...M. Determination of...15 derivatives were determined experimentally by radiolabeled competition binding assays using an extract 16 from Hi5 insect cells expressing an N

Flagellar Synchronization Is a Simple Alternative to Cell Cycle Synchronization for Ciliary and Flagellar Studies

PubMed Central

Dutta, Soumita

2017-01-01

ABSTRACT The unicellular green alga Chlamydomonas reinhardtii is an ideal model organism for studies of ciliary function and assembly. In assays for biological and biochemical effects of various factors on flagellar structure and function, synchronous culture is advantageous for minimizing variability. Here, we have characterized a method in which 100% synchronization is achieved with respect to flagellar length but not with respect to the cell cycle. The method requires inducing flagellar regeneration by amputation of the entire cell population and limiting regeneration time. This results in a maximally homogeneous distribution of flagellar lengths at 3 h postamputation. We found that time-limiting new protein synthesis during flagellar synchronization limits variability in the unassembled pool of limiting flagellar protein and variability in flagellar length without affecting the range of cell volumes. We also found that long- and short-flagella mutants that regenerate normally require longer and shorter synchronization times, respectively. By minimizing flagellar length variability using a simple method requiring only hours and no changes in media, flagellar synchronization facilitates the detection of small changes in flagellar length resulting from both chemical and genetic perturbations in Chlamydomonas. This method increases our ability to probe the basic biology of ciliary size regulation and related disease etiologies. IMPORTANCE Cilia and flagella are highly conserved antenna-like organelles that found in nearly all mammalian cell types. They perform sensory and motile functions contributing to numerous physiological and developmental processes. Defects in their assembly and function are implicated in a wide range of human diseases ranging from retinal degeneration to cancer. Chlamydomonas reinhardtii is an algal model system for studying mammalian cilium formation and function. Here, we report a simple synchronization method that allows detection of small changes in ciliary length by minimizing variability in the population. We find that this method alters the key relationship between cell size and the amount of protein accumulated for flagellar growth. This provides a rapid alternative to traditional methods of cell synchronization for uncovering novel regulators of cilia. PMID:28289724

Flagellar Synchronization Is a Simple Alternative to Cell Cycle Synchronization for Ciliary and Flagellar Studies.

PubMed

Dutta, Soumita; Avasthi, Prachee

2017-01-01

The unicellular green alga Chlamydomonas reinhardtii is an ideal model organism for studies of ciliary function and assembly. In assays for biological and biochemical effects of various factors on flagellar structure and function, synchronous culture is advantageous for minimizing variability. Here, we have characterized a method in which 100% synchronization is achieved with respect to flagellar length but not with respect to the cell cycle. The method requires inducing flagellar regeneration by amputation of the entire cell population and limiting regeneration time. This results in a maximally homogeneous distribution of flagellar lengths at 3 h postamputation. We found that time-limiting new protein synthesis during flagellar synchronization limits variability in the unassembled pool of limiting flagellar protein and variability in flagellar length without affecting the range of cell volumes. We also found that long- and short-flagella mutants that regenerate normally require longer and shorter synchronization times, respectively. By minimizing flagellar length variability using a simple method requiring only hours and no changes in media, flagellar synchronization facilitates the detection of small changes in flagellar length resulting from both chemical and genetic perturbations in Chlamydomonas . This method increases our ability to probe the basic biology of ciliary size regulation and related disease etiologies. IMPORTANCE Cilia and flagella are highly conserved antenna-like organelles that found in nearly all mammalian cell types. They perform sensory and motile functions contributing to numerous physiological and developmental processes. Defects in their assembly and function are implicated in a wide range of human diseases ranging from retinal degeneration to cancer. Chlamydomonas reinhardtii is an algal model system for studying mammalian cilium formation and function. Here, we report a simple synchronization method that allows detection of small changes in ciliary length by minimizing variability in the population. We find that this method alters the key relationship between cell size and the amount of protein accumulated for flagellar growth. This provides a rapid alternative to traditional methods of cell synchronization for uncovering novel regulators of cilia.

Dense Pattern Optical Multipass Cell

NASA Technical Reports Server (NTRS)

Silver, Joel A. (Inventor)

2009-01-01

A multiple pass optical cell and method comprising providing a pair of opposed cylindrical mirrors having curved axes with substantially equal focal lengths, positioning an entrance hole for introducing light into the cell and an exit hole for extracting light from the cell, wherein the entrance hole and exit hole are coextensive or non-coextensive, introducing light into the cell through the entrance hole, and extracting light from the cell through the exit hole.

Dense pattern optical multipass cell

DOEpatents

Silver, Joel A [Santa Fe, NM

2009-01-13

A multiple pass optical cell and method comprising providing a pair of opposed cylindrical mirrors having curved axes with substantially equal focal lengths, positioning an entrance hole for introducing light into the cell and an exit hole for extracting light from the cell, wherein the entrance hole and exit hole are coextensive or non-coextensive, introducing light into the cell through the entrance hole, and extracting light from the cell through the exit hole.

Active Cells for Multifunctional Structures

DTIC Science & Technology

2014-09-24

techniques to explore a variety of cell designs.  Designed a simplified active cell using Nitinol as the actuation method and relying on Joule heating...for contraction of the cell.  Developed manufacturing techniques for reliably creating Nitinol spring coils in a variety of diameters and gauges...design of the active cells to maximum the stroked length of the active cells by tuning the stiffness of a passive spring in parallel with the Nitinol

Neurite outgrowth of murine cerebellar granule cells can be enhanced by aniracetam with or without alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA).

PubMed

Fushiki, S; Matsumoto, K; Nagata, A

1995-10-27

To assess the neurotrophic effects of a nootropic drug, aniracetam, we studied neurite extension of mouse cerebellar granule cells in culture with low or with high K+ under different combinations of drugs and then immunohistochemically stained the cells with an antibody against L1, a neural cell adhesion molecule on cerebellar granule cells. Quantitative analyses using parameters of the total neurite length, maximal neurite length and number of branches disclosed that aniracetam, in the presence of high K+ and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), significantly enhanced neurite extension of cultured granule neurons. Aniracetam alone also stimulated neurite extension of cerebellar granule cells at a longer period of culture with low K+ showing a bell-shaped dose response curve with maximal effects at 10 microM. Aniracetam may influence remodeling of the neural network after injury.

Chronic nerve compression alters Schwann cell myelin architecture in a murine model

PubMed Central

Gupta, Ranjan; Nassiri, Nima; Hazel, Antony; Bathen, Mary; Mozaffar, Tahseen

2011-01-01

Introduction Myelinating Schwann cells compartmentalize their outermost layer to form actin-rich channels known as Cajal bands. Here, we investigate changes in Schwann cell architecture and cytoplasmic morphology in a novel mouse model of carpal tunnel syndrome. Methods Chronic nerve compression (CNC) injury was created in wild-type and slow-Wallerian degeneration (WldS) mice. Over 12 weeks, nerves were electrodiagnostically assessed, and Schwann cell morphology was thoroughly evaluated. Results A decline in nerve conduction velocity and increase in g-ratio is observed without early axonal damage. Schwann cells display shortened internodal lengths and severely disrupted Cajal bands. Quite surprisingly, the latter is reconstituted without improvements to nerve conduction velocity. Discussion Chronic entrapment injuries like carpal tunnel syndrome are primarily mediated by the Schwann cell response, wherein decreases in internodal length and myelin thickness disrupt the efficiency of impulse propagation. Restitution of Cajal bands is not sufficient for remyelination post-CNC injury. PMID:22246880

Alternative lengthening of telomeres can be maintained by preferential elongation of lagging strands

PubMed Central

Min, Jaewon; Wright, Woodring E.

2017-01-01

Abstract Alternative lengthening of telomeres (ALT) is a telomerase independent telomere maintenance mechanism that occurs in ∼15% of cancers. The potential mechanism of ALT is homology-directed telomere synthesis, but molecular mechanisms of how ALT maintains telomere length in human cancer is poorly understood. Here, we generated TERC (telomerase RNA) gene knockouts in telomerase positive cell lines that resulted in long-term surviving clones acquiring the ALT pathway but at a very low frequency. By comparing these ALT cells with parental telomerase positive cells, we observed that ALT cells possess excessively long telomeric overhangs derived from telomere elongation processes that mostly occur during S phase. ALT cells exhibited preferential elongation of the telomeric lagging strands, whereas telomerase positive cells exhibited similar elongation between leading and lagging strands. We propose that the ALT pathway preferentially occurs at telomeric lagging strands leading to heterogeneous telomere lengths observed in most ALT cancers. PMID:28082393

Roughness threshold for cell attachment and proliferation on plasma micro-nanotextured polymeric surfaces: the case of primary human skin fibroblasts and mouse immortalized 3T3 fibroblasts

NASA Astrophysics Data System (ADS)

Bourkoula, A.; Constantoudis, V.; Kontziampasis, D.; Petrou, P. S.; Kakabakos, S. E.; Tserepi, A.; Gogolides, E.

2016-08-01

Poly(methyl methacrylate) surfaces have been micro-nanotextured in oxygen plasmas with increasing ion energy, leading to micro-nanotopography characterized by increased root mean square roughness, correlation length and fractal dimension. Primary human skin fibroblasts and mouse immortalized 3T3 fibroblasts were cultured on these surfaces and the number of adhering cells, their proliferation rate and morphology (cytoplasm and nucleus area) were evaluated as a function of roughness height, correlation length, and fractal dimension. A roughness threshold behavior was observed for both types of cells leading to dramatic cell number decrease above this threshold, which is almost similar for the two types of cells, despite their differences in size and stiffness. The results are discussed based on two theoretical models, which are reconciled and unified when the elastic moduli and the size of the cells are taken into account.

Gigantism in a bacterium, Epulopiscium fishelsoni, correlates with complex patterns in arrangement, quantity, and segregation of DNA.

PubMed

Bresler, V; Montgomery, W L; Fishelson, L; Pollak, P E

1998-11-01

Epulopiscium fishelsoni, gut symbiont of the brown surgeonfish (Acanthurus nigrofuscus) in the Red Sea, attains a larger size than any other eubacterium, varies 10- to 20-fold in length (and >2, 000-fold in volume), and undergoes a complex daily life cycle. In early morning, nucleoids contain highly condensed DNA in elongate, chromosome-like structures which are physically separated from the general cytoplasm. Cell division involves production of two (rarely three) nucleoids within a cell, deposition of cell walls around expanded nucleoids, and emergence of daughter cells from the parent cell. Fluorescence measurements of DNA, RNA, and other cell components indicate the following. DNA quantity is proportional to cell volume over cell lengths of approximately 30 micrometers to >500 micrometers. For cells of a given size, nucleoids of cells with two nucleoids (binucleoid) contain approximately equal amounts of DNA. And each nucleoid of a binucleoid cell contains one-half the DNA of the single nucleoid in a uninucleoid cell of the same size. The life cycle involves approximately equal subdivision of DNA among daughter cells, formation of apical caps of condensed DNA from previously decondensed and diffusely distributed DNA, and "pinching" of DNA near the middle of the cell in the absence of new wall formation. Mechanisms underlying these patterns remain unclear, but formation of daughter nucleoids and cells occurs both during diurnal periods of host feeding and bacterial cell growth and during nocturnal periods of host inactivity when mean bacterial cell size declines.

Comparison of telomere length and association with progenitor cell markers in lacrimal gland between Sjögren syndrome and non-Sjögren syndrome dry eye patients

PubMed Central

Kawashima, Motoko; Maida, Yoshiko; Kamoi, Mizuka; Ogawa, Yoko; Shimmura, Shigeto; Masutomi, Kenkichi; Tsubota, Kazuo

2011-01-01

Purpose Indicators of aging such as disruption of telomeric function due to shortening may be more frequent in dysfunctional lacrimal gland. The aims of this study were to 1) determine the viability of quantitative fluorescence in situ hybridization of telomeres (telo-FISH) for the assessment of telomere length in lacrimal gland in Sjögren and non- Sjögren syndrome patients; and 2) investigate the relationship between progenitor cell markers and telomere length in both groups. Methods Quantitative fluorescence in situ hybridization with a peptide nucleic acid probe complementary to the telomere repeat sequence was performed on frozen sections from human lacrimal gland tissues. The mean fluorescence intensity of telomere spots was automatically quantified by image analysis as relative telomere length in lacrimal gland epithelial cells. Immunostaining for p63, nucleostemin, ATP-binding cassette, sub-family G, member 2 (ABCG2), and nestin was also performed. Results Telomere intensity in the Sjögren syndrome group (6,785.0±455) was significantly lower than that in the non-Sjögren syndrome group (7,494.7±477; p=0.02). Among the samples from the non-Sjögren syndrome group, immunostaining revealed that p63 was expressed in 1–3 acinar cells in each acinar unit and continuously in the basal layer of duct cells. In contrast, in the Sjögren syndrome group, p63 and nucleostemin showed a lower level of expression. ABCG2 was expressed in acinar cells in both sjogren and non-Sjogren syndrome. Conclusions The results of this study indicate that 1) telo-FISH is a viable method of assessing telomere length in lacrimal gland, and 2) telomere length in Sjögren syndrome is shorter and associated with lower levels of expression of p63 and nucleostemin than in non-Sjögren syndrome. PMID:21655359

Physics of active jamming during collective cellular motion in a monolayer

PubMed Central

Garcia, Simon; Hannezo, Edouard; Elgeti, Jens; Joanny, Jean-François; Silberzan, Pascal; Gov, Nir S.

2015-01-01

Although collective cell motion plays an important role, for example during wound healing, embryogenesis, or cancer progression, the fundamental rules governing this motion are still not well understood, in particular at high cell density. We study here the motion of human bronchial epithelial cells within a monolayer, over long times. We observe that, as the monolayer ages, the cells slow down monotonously, while the velocity correlation length first increases as the cells slow down but eventually decreases at the slowest motions. By comparing experiments, analytic model, and detailed particle-based simulations, we shed light on this biological amorphous solidification process, demonstrating that the observed dynamics can be explained as a consequence of the combined maturation and strengthening of cell−cell and cell−substrate adhesions. Surprisingly, the increase of cell surface density due to proliferation is only secondary in this process. This analysis is confirmed with two other cell types. The very general relations between the mean cell velocity and velocity correlation lengths, which apply for aggregates of self-propelled particles, as well as motile cells, can possibly be used to discriminate between various parameter changes in vivo, from noninvasive microscopy data. PMID:26627719

Near Field Imaging of Gallium Nitride Nanowires for Characterization of Minority Carrier Diffusion

DTIC Science & Technology

2009-12-01

diffusion length in nanowires is critical to potential applications in solar cells , spectroscopic sensing, and/or lasers and light emitting diodes (LED...technique has been successfully demonstrated with thin film solar cell materials [4, 5]. In these experiments, the diffusion length was measured using a...minority carrier diffusion length . This technique has been used in the near-field collection mode to image the diffusion of holes in n-type GaN

Socioeconomic status and length of hospital stay in children with vaso-occlusive crises of sickle cell disease.

PubMed Central

Ellison, Angela M.; Bauchner, Howard

2007-01-01

OBJECTIVE: To examine the association between socioeconomic status and length of hospital stay for vaso-occlusive crises in children with sickle cell disease. METHODS: 19,174 discharges (aged 1-20 years), with a primary diagnosis of sickle cell disease with crisis were analyzed from the Healthcare Cost and Utilization Project Kid Inpatient Database 2000. Socioeconomic status was assessed using an area-based measure, median household income by ZIP code and an individual-level measure, insurance status. We adjusted for age, gender, hospital location/teaching status, presence of pneumonia, number of diagnoses on record and number of procedures performed. Negative binomial regression models using generalized estimating equations (GEE) were used to assess length of stay. RESULTS: Socioeconomic status as measured by income was not associated with length of stay (incidence rate ratio (highest versus lowest category) = 1.04 (95% CI: 0.98, 1.11)). In contrast, socioeconomic status as measured by insurance was associated with length of stay [adjusted incidence rate ratio = 1.04 (95% CI: 1.01, 1.08)), although the magnitude of this difference is small and not likely to be clinically important. CONCLUSIONS: We found no evidence to suggest that socioeconomic status has any clinically important effect on length of hospital stay in children with vaso-occlusive crises in sickle cell disease. PMID:17393942

Increased pericarp cell length underlies a major quantitative trait locus for grain weight in hexaploid wheat.

PubMed

Brinton, Jemima; Simmonds, James; Minter, Francesca; Leverington-Waite, Michelle; Snape, John; Uauy, Cristobal

2017-08-01

Crop yields must increase to address food insecurity. Grain weight, determined by grain length and width, is an important yield component, but our understanding of the underlying genes and mechanisms is limited. We used genetic mapping and near isogenic lines (NILs) to identify, validate and fine-map a major quantitative trait locus (QTL) on wheat chromosome 5A associated with grain weight. Detailed phenotypic characterisation of developing and mature grains from the NILs was performed. We identified a stable and robust QTL associated with a 6.9% increase in grain weight. The positive interval leads to 4.0% longer grains, with differences first visible 12 d after fertilization. This grain length effect was fine-mapped to a 4.3 cM interval. The locus also has a pleiotropic effect on grain width (1.5%) during late grain development that determines the relative magnitude of the grain weight increase. Positive NILs have increased maternal pericarp cell length, an effect which is independent of absolute grain length. These results provide direct genetic evidence that pericarp cell length affects final grain size and weight in polyploid wheat. We propose that combining genes that control distinct biological mechanisms, such as cell expansion and proliferation, will enhance crop yields. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Somatic mosaicism of androgen receptor CAG repeats in colorectal carcinoma epithelial cells from men.

PubMed

Di Fabio, Francesco; Alvarado, Carlos; Gologan, Adrian; Youssef, Emad; Voda, Linda; Mitmaker, Elliot; Beitel, Lenore K; Gordon, Philip H; Trifiro, Mark

2009-06-01

The X-linked human androgen receptor gene (AR) contains an exonic polymorphic trinucleotide CAG. The length of this encoded CAG tract inversely affects AR transcriptional activity. Colorectal carcinoma is known to express the androgen receptor, but data on somatic CAG repeat lengths variations in malignant and normal epithelial cells are still sporadic. Using laser capture microdissection (LCM), epithelial cells from colorectal carcinoma and normal-appearing mucosa were collected from the fresh tissue of eight consecutive male patients undergoing surgery (mean age, 70 y; range, 54-82). DNA isolated from each LCM sample underwent subsequent PCR and DNA sequencing to precisely determine AR CAG repeat lengths and the presence of microsatellite instability (MSI). Different AR CAG repeat lengths were observed in colorectal carcinoma (ranging from 0 to 36 CAG repeats), mainly in the form of multiple shorter repeat lengths. This genetic heterogeneity (somatic mosaicism) was also found in normal-appearing colorectal mucosa. Half of the carcinoma cases examined tended to have a higher number of AR CAG repeat lengths with a wider range of repeat size variation compared to normal mucosa. MSI carcinomas tended to have longer median AR CAG repeat lengths (n = 17) compared to microsatellite stable carcinomas (n = 14), although the difference was not significant (P = 0.31, Mann-Whitney test). Multiple unique somatic mutations of the AR CAG repeats occur in colorectal mucosa and in carcinoma, predominantly resulting in shorter alleles. Colorectal epithelial cells carrying AR alleles with shorter CAG repeat lengths may be more androgen-sensitive and therefore have a growth advantage.

Expression of connective tissue growth factor (CTGF/CCN2) in breast cancer cells is associated with increased migration and angiogenesis.

PubMed

Chien, Wenwen; O'Kelly, James; Lu, Daning; Leiter, Amanda; Sohn, Julia; Yin, Dong; Karlan, Beth; Vadgama, Jay; Lyons, Karen M; Koeffler, H Phillip

2011-06-01

Connective tissue growth factor (CTGF/CCN2) belongs to the CCN family of matricellular proteins, comprising Cyr61, CTGF, NovH and WISP1-3. The CCN proteins contain an N-terminal signal peptide followed by four conserved domains sharing sequence similarities with the insulin-like growth factor binding proteins, von Willebrand factor type C repeat, thrombospondin type 1 repeat, and a C-terminal growth factor cysteine knot domain. To investigate the role of CCN2 in breast cancer, we transfected MCF-7 cells with full-length CCN2, and with four mutant constructs in which one of the domains had been deleted. MCF-7 cells stably expressing full-length CCN2 demonstrated reduced cell proliferation, increased migration in Boyden chamber assays and promoted angiogenesis in chorioallantoic membrane assays compared to control cells. Deletion of the C-terminal cysteine knot domain, but not of any other domain-deleted mutants, abolished activities mediated by full-length CCN2. We have dissected the role of CCN2 in breast tumorigenesis on a structural basis.

Correlations of catalytic combustor performance parameters

NASA Technical Reports Server (NTRS)

Bulzan, D. L.

1978-01-01

Correlations for combustion efficiency percentage drop and the minimum required adiabatic reaction temperature necessary to meet emissions goals of 13.6 g CO/kg fuel and 1.64 g HC/kg fuel are presented. Combustion efficiency was found to be a function of the cell density, cell circumference, reactor length, reference velocity, and adiabatic reaction temperature. The percentage pressure drop at an adiabatic reaction temperature of 1450 K was found to be proportional to the reference velocity to the 1.5 power and to the reactor length. It is inversely proportional to the pressure, cell hydraulic diameter, and fractional open area. The minimum required adiabatic reaction temperature was found to increase with reference velocity and decrease with cell circumference, cell density and reactor length. A catalyst factor was introduced into the correlations to account for differences between catalysts. Combustion efficiency, the percentage pressure drop, and the minimum required adiabatic reaction temperature were found to be a function of the catalyst factor. The data was from a 12 cm-diameter test rig with noble metal reactors using propane fuel at an inlet temperature of 800 K.

In Vivo Protein Dynamics on the Nanometer Length Scale and Nanosecond Time Scale

DOE PAGES

Anunciado, Divina B.; Nyugen, Vyncent P.; Hurst, Gregory B.; ...

2017-04-07

Selectively labeled GroEL protein was produced in living deuterated bacterial cells to enhance its neutron scattering signal above that of the intracellular milieu. Quasi-elastic neutron scattering shows that the in-cell diffusion coefficient of GroEL was (4.7 ± 0.3) × 10 –12 m 2/s, a factor of 4 slower than its diffusion coefficient in buffer solution. Furthermore, for internal protein dynamics we see a relaxation time of (65 ± 6) ps, a factor of 2 slower compared to the protein in solution. Comparison to the literature suggests that the effective diffusivity of proteins depends on the length and time scale beingmore » probed. Retardation of in-cell diffusion compared to the buffer becomes more significant with the increasing probe length scale, suggesting that intracellular diffusion of biomolecules is nonuniform over the cellular volume. This approach outlined here enables investigation of protein dynamics within living cells to open up new lines of research using “in-cell neutron scattering” to study the dynamics of complex biomolecular systems.« less

In Vivo Protein Dynamics on the Nanometer Length Scale and Nanosecond Time Scale

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

Anunciado, Divina B.; Nyugen, Vyncent P.; Hurst, Gregory B.

Selectively labeled GroEL protein was produced in living deuterated bacterial cells to enhance its neutron scattering signal above that of the intracellular milieu. Quasi-elastic neutron scattering shows that the in-cell diffusion coefficient of GroEL was (4.7 ± 0.3) × 10 –12 m 2/s, a factor of 4 slower than its diffusion coefficient in buffer solution. Furthermore, for internal protein dynamics we see a relaxation time of (65 ± 6) ps, a factor of 2 slower compared to the protein in solution. Comparison to the literature suggests that the effective diffusivity of proteins depends on the length and time scale beingmore » probed. Retardation of in-cell diffusion compared to the buffer becomes more significant with the increasing probe length scale, suggesting that intracellular diffusion of biomolecules is nonuniform over the cellular volume. This approach outlined here enables investigation of protein dynamics within living cells to open up new lines of research using “in-cell neutron scattering” to study the dynamics of complex biomolecular systems.« less

Physical activity and telomere length: Impact of aging and potential mechanisms of action

PubMed Central

Arsenis, Nicole C.; You, Tongjian; Ogawa, Elisa F.; Tinsley, Grant M.; Zuo, Li

2017-01-01

Telomeres protect the integrity of information-carrying DNA by serving as caps on the terminal portions of chromosomes. Telomere length decreases with aging, and this contributes to cell senescence. Recent evidence supports that telomere length of leukocytes and skeletal muscle cells may be positively associated with healthy living and inversely correlated with the risk of several age-related diseases, including cancer, cardiovascular disease, obesity, diabetes, chronic pain, and stress. In observational studies, higher levels of physical activity or exercise are related to longer telomere lengths in various populations, and athletes tend to have longer telomere lengths than non-athletes. This relationship is particularly evident in older individuals, suggesting a role of physical activity in combating the typical age-induced decrements in telomere length. To date, a small number of exercise interventions have been executed to examine the potential influence of chronic exercise on telomere length, but these studies have not fully established such relationship. Several potential mechanisms through which physical activity or exercise could affect telomere length are discussed, including changes in telomerase activity, oxidative stress, inflammation, and decreased skeletal muscle satellite cell content. Future research is needed to mechanistically examine the effects of various modalities of exercise on telomere length in middle-aged and older adults, as well as in specific clinical populations. PMID:28410238

Physical activity and telomere length: Impact of aging and potential mechanisms of action.

PubMed

Arsenis, Nicole C; You, Tongjian; Ogawa, Elisa F; Tinsley, Grant M; Zuo, Li

2017-07-04

Telomeres protect the integrity of information-carrying DNA by serving as caps on the terminal portions of chromosomes. Telomere length decreases with aging, and this contributes to cell senescence. Recent evidence supports that telomere length of leukocytes and skeletal muscle cells may be positively associated with healthy living and inversely correlated with the risk of several age-related diseases, including cancer, cardiovascular disease, obesity, diabetes, chronic pain, and stress. In observational studies, higher levels of physical activity or exercise are related to longer telomere lengths in various populations, and athletes tend to have longer telomere lengths than non-athletes. This relationship is particularly evident in older individuals, suggesting a role of physical activity in combating the typical age-induced decrements in telomere length. To date, a small number of exercise interventions have been executed to examine the potential influence of chronic exercise on telomere length, but these studies have not fully established such relationship. Several potential mechanisms through which physical activity or exercise could affect telomere length are discussed, including changes in telomerase activity, oxidative stress, inflammation, and decreased skeletal muscle satellite cell content. Future research is needed to mechanistically examine the effects of various modalities of exercise on telomere length in middle-aged and older adults, as well as in specific clinical populations.

Telomerase Activity in Human Ovarian Carcinoma

NASA Astrophysics Data System (ADS)

Counter, Christopher M.; Hirte, Hal W.; Bacchetti, Silvia; Harley, Calvin B.

1994-04-01

Telomeres fulfill the dual function of protecting eukaryotic chromosomes from illegitimate recombination and degradation and may aid in chromosome attachment to the nuclear membrane. We have previously shown that telomerase, the enzyme which synthesizes telomeric DNA, is not detected in normal somatic cells and that telomeres shorten with replicative age. In cells immortalized in vitro, activation of telomerase apparently stabilizes telomere length, preventing a critical destabilization of chromosomes, and cell proliferation continues even when telomeres are short. In vivo, telomeres of most tumors are shorter than telomeres of control tissues, suggesting an analogous role for the enzyme. To assess the relevance of telomerase and telomere stability in the development and progression of tumors, we have measured enzyme activity and telomere length in metastatic cells of epithelial ovarian carcinoma. We report that extremely short telomeres are maintained in these cells and that tumor cells, but not isogenic nonmalignant cells, express telomerase. Our findings suggest that progression of malignancy is ultimately dependent upon activation of telomerase and that telomerase inhibitors may be effective antitumor drugs.

Androgen receptor polyglutamine repeat length affects receptor activity and C2C12 cell development.

PubMed

Sheppard, Ryan L; Spangenburg, Espen E; Chin, Eva R; Roth, Stephen M

2011-10-20

Testosterone (T) has an anabolic effect on skeletal muscle and is believed to exert its local effects via the androgen receptor (AR). The AR harbors a polymorphic stretch of glutamine repeats demonstrated to inversely affect receptor transcriptional activity in prostate and kidney cells. The effects of AR glutamine repeat length on skeletal muscle are unknown. In this study we examined the effect of AR CAG repeat length on AR function in C2C12 cells. AR expression vectors harboring 14, 24, and 33 CAG repeats were used to assess AR transcriptional activity. C2C12 cell proliferation, differentiation, gene expression, myotube formation, and myonuclear fusion index were assessed. Transcriptional activity increased with increasing repeat length and in response to testosterone (AR14 = 3.91 ± 0.26, AR24 = 25.21 ± 1.72, AR33 = 36.08 ± 3.22 relative light units; P < 0.001). Ligand activation was increased for AR33 (2.10 ± 0.04) compared with AR14 (1.54 ± 0.09) and AR24 (1.57 ± 0.05, P < 0.001). AR mRNA expression was elevated in each stably transfected line. AR33 cell proliferation (20,512.3 ± 1,024.0) was decreased vs. AR14 (27,604.17 ± 1,425.3; P < 0.001) after 72 h. Decreased CK activity in AR14 cells (54.9 ± 2.9 units/μg protein) in comparison to AR33 (70.8 ± 8.1) (P < 0.05) was noted. The myonuclear fusion index was lower for AR14 (15.21 ± 3.24%) and AR33 (9.97 ± 3.14%) in comparison to WT (35.07 ± 5.60%, P < 0.001). AR14 and AR33 cells also displayed atypical myotube morphology. RT-PCR revealed genotype differences in myostatin and myogenin expression. We conclude that AR polyglutamine repeat length is directly associated with transcriptional activity and alters the growth and development of C2C12 cells. This polymorphism may contribute to the heritability of muscle mass in humans.

Visualization and quantitative analysis of extrachromosomal telomere-repeat DNA in individual human cells by Halo-FISH

PubMed Central

Komosa, Martin; Root, Heather; Meyn, M. Stephen

2015-01-01

Current methods for characterizing extrachromosomal nuclear DNA in mammalian cells do not permit single-cell analysis, are often semi-quantitative and frequently biased toward the detection of circular species. To overcome these limitations, we developed Halo-FISH to visualize and quantitatively analyze extrachromosomal DNA in single cells. We demonstrate Halo-FISH by using it to analyze extrachromosomal telomere-repeat (ECTR) in human cells that use the Alternative Lengthening of Telomeres (ALT) pathway(s) to maintain telomere lengths. We find that GM847 and VA13 ALT cells average ∼80 detectable G/C-strand ECTR DNA molecules/nucleus, while U2OS ALT cells average ∼18 molecules/nucleus. In comparison, human primary and telomerase-positive cells contain <5 ECTR DNA molecules/nucleus. ECTR DNA in ALT cells exhibit striking cell-to-cell variations in number (<20 to >300), range widely in length (<1 to >200 kb) and are composed of primarily G- or C-strand telomere-repeat DNA. Halo-FISH enables, for the first time, the simultaneous analysis of ECTR DNA and chromosomal telomeres in a single cell. We find that ECTR DNA comprises ∼15% of telomere-repeat DNA in GM847 and VA13 cells, but <4% in U2OS cells. In addition to its use in ALT cell analysis, Halo-FISH can facilitate the study of a wide variety of extrachromosomal DNA in mammalian cells. PMID:25662602

Expression, purification, and characterization of recombinant human and murine milk fat globule-epidermal growth factor-factor 8.

PubMed

Castellanos, Erick R; Ciferri, Claudio; Phung, Wilson; Sandoval, Wendy; Matsumoto, Marissa L

2016-08-01

Milk fat globule-epidermal growth factor-factor 8 (MFG-E8), as its name suggests, is a major glycoprotein component of milk fat globules secreted by the mammary epithelium. Although its role in milk fat production is unclear, MFG-E8 has been shown to act as a bridge linking apoptotic cells to phagocytes for removal of these dying cells. MFG-E8 is capable of bridging these two very different cell types via interactions through both its epidermal growth factor (EGF)-like domain(s) and its lectin-type C domains. The EGF-like domain interacts with αVβ3 and αVβ5 integrins on the surface of phagocytes, whereas the C domains bind phosphatidylserine found on the surface of apoptotic cells. In an attempt to purify full-length, recombinant MFG-E8 expressed in either insect cells or CHO cells, we find that it is highly aggregated. Systematic truncation of the domain architecture of MFG-E8 indicates that the C domains are mainly responsible for the aggregation propensity. Addition of Triton X-100 to the conditioned cell culture media allowed partial recovery of non-aggregated, full-length MFG-E8. A more comprehensive detergent screen identified CHAPS as a stabilizer of MFG-E8 and allowed purification of a significant portion of non-aggregated, full-length protein. The CHAPS-stabilized recombinant MFG-E8 retained its natural ability to bind both αVβ3 and αVβ5 integrins and phosphatidylserine suggesting that it is properly folded and active. Herein we describe an efficient purification method for production of non-aggregated, full-length MFG-E8. Copyright © 2016 Elsevier Inc. All rights reserved.

Numerically bridging lamellipodial and filopodial activity during cell spreading reveals a potentially novel trigger of focal adhesion maturation.

PubMed

Loosli, Y; Vianay, B; Luginbuehl, R; Snedeker, J G

2012-05-01

We present a novel approach to modeling cell spreading, and use it to reveal a potentially central mechanism regulating focal adhesion maturation in various cell phenotypes. Actin bundles that span neighboring focal complexes at the lamellipodium-lamellum interface were assumed to be loaded by intracellular forces in proportion to bundle length. We hypothesized that the length of an actin bundle (with the corresponding accumulated force at its adhesions) may thus regulate adhesion maturation to ensure cell mechanical stability and morphological integrity. We developed a model to test this hypothesis, implementing a "top-down" approach to simplify certain cellular processes while explicitly incorporating complexity of other key subcellular mechanisms. Filopodial and lamellipodial activities were treated as modular processes with functional spatiotemporal interactions coordinated by rules regarding focal adhesion turnover and actin bundle dynamics. This theoretical framework was able to robustly predict temporal evolution of cell area and cytoskeletal organization as reported from a wide range of cell spreading experiments using micropatterned substrates. We conclude that a geometric/temporal modeling framework can capture the key functional aspects of the rapid spreading phase and resultant cytoskeletal complexity. Hence the model is used to reveal mechanistic insight into basic cell behavior essential for spreading. It demonstrates that actin bundles spanning nascent focal adhesions such that they are aligned to the leading edge may accumulate centripetal endogenous forces along their length, and could thus trigger focal adhesion maturation in a force-length dependent fashion. We suggest that this mechanism could be a central "integrating" factor that effectively coordinates force-mediated adhesion maturation at the lamellipodium-lamellum interface.

Characterizing emergency admissions of patients with sickle cell crisis in NHS brent: observational study

PubMed Central

Green, Stuart A; AlJuburi, Ghida; Majeed, Azeem; Okoye, Ogo; Amobi, Carole; Banarsee, Ricky; Phekoo, Karen J

2012-01-01

Objectives To characterize emergency admissions for patients with sickle cell crisis in NHS Brent and to determine which patients and practices may benefit most from primary care intervention. Design Observational study Setting Emergency departments attended by residents of the London borough of Brent Participants Patients with sickle cell disease registered with a general practitioner (GP) in the borough of Brent Main outcome measures Analysis of admissions between January 2008 and July 2010 that included length of stay (average and <2 days versus ≥2 days) by age group and registered GP practice. Results Thirty six percent of sickle cell disease admission spells resulted in a length of stay of less than two days. Seventy four percent of total bed days are associated with patients with more than one admission during the period of analysis, i.e. multiple admissions. Two general practices in Brent were identified as having the highest number of patients admitted to the emergency department for sickle cell crisis and may benefit most from primary care intervention. Discussion Patients with short length of stay and multiple admissions may be potentially amenable to primary care intervention. The practices which have the highest numbers of sickle cell disease patients who frequently seek emergency care will be earmarked for an education intervention designed to help further engage general practitioners in the care and management of their sickle cell patients. PMID:22768371

Characterizing emergency admissions of patients with sickle cell crisis in NHS brent: observational study.

PubMed

Green, Stuart A; Aljuburi, Ghida; Majeed, Azeem; Okoye, Ogo; Amobi, Carole; Banarsee, Ricky; Phekoo, Karen J

2012-06-01

To characterize emergency admissions for patients with sickle cell crisis in NHS Brent and to determine which patients and practices may benefit most from primary care intervention. Observational study Emergency departments attended by residents of the London borough of Brent Patients with sickle cell disease registered with a general practitioner (GP) in the borough of Brent Analysis of admissions between January 2008 and July 2010 that included length of stay (average and <2 days versus ≥2 days) by age group and registered GP practice. Thirty six percent of sickle cell disease admission spells resulted in a length of stay of less than two days. Seventy four percent of total bed days are associated with patients with more than one admission during the period of analysis, i.e. multiple admissions. Two general practices in Brent were identified as having the highest number of patients admitted to the emergency department for sickle cell crisis and may benefit most from primary care intervention. Patients with short length of stay and multiple admissions may be potentially amenable to primary care intervention. The practices which have the highest numbers of sickle cell disease patients who frequently seek emergency care will be earmarked for an education intervention designed to help further engage general practitioners in the care and management of their sickle cell patients.

Contraction of gut smooth muscle cells assessed by fluorescence imaging.

PubMed

Tokita, Yohei; Akiho, Hirotada; Nakamura, Kazuhiko; Ihara, Eikichi; Yamamoto, Masahiro

2015-03-01

Here we discuss the development of a novel cell imaging system for the evaluation of smooth muscle cell (SMC) contraction. SMCs were isolated from the circular and longitudinal muscular layers of mouse small intestine by enzymatic digestion. SMCs were stimulated by test agents, thereafter fixed in acrolein. Actin in fixed SMCs was stained with phalloidin and cell length was determined by measuring diameter at the large end of phalloidin-stained strings within the cells. The contractile response was taken as the decrease in the average length of a population of stimulated-SMCs. Various mediators and chemically identified compounds of daikenchuto (DKT), pharmaceutical-grade traditional Japanese prokinetics, were examined. Verification of the integrity of SMC morphology by phalloidin and DAPI staining and semi-automatic measurement of cell length using an imaging analyzer was a reliable method by which to quantify the contractile response. Serotonin, substance P, prostaglandin E2 and histamine induced SMC contraction in concentration-dependent manner. Two components of DKT, hydroxy-α-sanshool and hydroxy-β-sanshool, induced contraction of SMCs. We established a novel cell imaging technique to evaluate SMC contractility. This method may facilitate investigation into SMC activity and its role in gastrointestinal motility, and may assist in the discovery of new prokinetic agents. Copyright © 2015 Japanese Pharmacological Society. Production and hosting by Elsevier B.V. All rights reserved.

Local homogeneity of cell cycle length in developing mouse cortex

NASA Technical Reports Server (NTRS)

Cai, L.; Hayes, N. L.; Nowakowski, R. S.

1997-01-01

We have measured the amount of variation in the length of the cell cycle for cells in the pseudostratified ventricular epithelium (PVE) of the developing cortex of mice on embryonic day 14. Our measurements were made in three cortical regions (i.e., the neocortex, archicortex, and periarchicortex) using three different methods: the cumulative labeling method (CLM), the percent labeled mitoses (PLM) method, and a comparison of the time needed for the PLM to ascend from 0 to 100% with the time needed for the PLM to descend from 100 to 0%. These 3 different techniques provide different perspectives on the cytokinetic parameters. Theoretically, CLM gives an estimate for a maximum value of the total length of the cell cycle (TC), whereas PLM gives an estimate of a minimum value of TC. The difference between these two estimates indicates that the range for TC is +/-1% of the mean TC for periarchicortex, +/-7% for neocortex, and +/-8% for archicortex. This was confirmed by a lengthening of the PLM descent time in comparison with its ascent time. The sharpness of the transitions and the flatness of the plateau of the PLM curves indicate that 99% of the proliferating cells are within this narrow estimated range for TC; hence, only approximately 1% deviate outside of a relatively restricted range from the average TC of the population. In the context of the possible existence within the cortical PVE of two populations with markedly dissimilar cell cycle kinetics from the mean, one such population must comprise approximately 99% of the total population, and the other, if it exists, is only approximately 1% of the total. This seems to be true for all three cortical regions. The narrow range of TC indicates a homogeneity in the cell cycle length for proliferating cells in three different cortical regions, despite the fact that progenitor cells of different lineages may be present. It further predicts the existence of almost synchronous interkinetic nuclear movements of the proliferating cells in the ventricular zone during early development of the cerebral cortex.

Controlling cell volume for efficient PHB production by Halomonas.

PubMed

Jiang, Xiao-Ran; Yao, Zhi-Hao; Chen, Guo-Qiang

2017-11-01

Bacterial morphology is decided by cytoskeleton protein MreB and cell division protein FtsZ encoded by essential genes mreB and ftsZ, respectively. Inactivating mreB and ftsZ lead to increasing cell sizes and cell lengths, respectively, yet seriously reduce cell growth ability. Here we develop a temperature-responsible plasmid expression system for compensated expression of relevant gene(s) in mreB or ftsZ disrupted recombinants H. campaniensis LS21, allowing mreB or ftsZ disrupted recombinants to grow normally at 30°C in a bioreactor for 12h so that a certain cell density can be reached, followed by 36h cell size expansions or cell shape elongations at elevated 37°C at which the mreB and ftsZ encoded plasmid pTKmf failed to replicate in the recombinants and thus lost themselves. Finally, 80% PHB yield increase was achieved via controllable morphology manipulated H. campaniensis LS21. It is concluded that controllable expanding cell volumes (widths or lengths) provides more spaces for accumulating more inclusion body polyhydroxybutyrate (PHB) and the resulting cell gravity precipitation benefits the final separation of cells and product during downstream. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Surface photovoltage method extended to silicon solar cell junction

NASA Technical Reports Server (NTRS)

Wang, E. Y.; Baraona, C. R.; Brandhorst, H. W., Jr.

1974-01-01

The conventional surface photovoltage (SPV) method is extended to the measurement of the minority carrier diffusion length in diffused semiconductor junctions of the type used in a silicon solar cell. The minority carrier diffusion values obtained by the SPV method agree well with those obtained by the X-ray method. Agreement within experimental error is also obtained between the minority carrier diffusion lengths in solar cell diffusion junctions and in the same materials with n-regions removed by etching, when the SPV method was used in the measurements.

Telomere length analysis.

PubMed

Canela, Andrés; Klatt, Peter; Blasco, María A

2007-01-01

Most somatic cells of long-lived species undergo telomere shortening throughout life. Critically short telomeres trigger loss of cell viability in tissues, which has been related to alteration of tissue function and loss of regenerative capabilities in aging and aging-related diseases. Hence, telomere length is an important biomarker for aging and can be used in the prognosis of aging diseases. These facts highlight the importance of developing methods for telomere length determination that can be employed to evaluate telomere length during the human aging process. Telomere length quantification methods have improved greatly in accuracy and sensitivity since the development of the conventional telomeric Southern blot. Here, we describe the different methodologies recently developed for telomere length quantification, as well as their potential applications for human aging studies.

Effects of the red tide dinoflagellate, Karenia brevis, on early development of the eastern oyster Crassostrea virginica and northern quahog Mercenaria mercenaria.

PubMed

Rolton, Anne; Vignier, Julien; Soudant, Philippe; Shumway, Sandra E; Bricelj, V Monica; Volety, Aswani K

2014-10-01

The brevetoxin-producing dinoflagellate, Karenia brevis, adversely affects many shellfish species including the commercially and ecologically important bivalve molluscs, the northern quahog (=hard clam) Mercenaria mercenaria and eastern oyster Crassostrea virginica, in the Gulf of Mexico, USA. This study assessed the effects of exposure of these bivalves to K. brevis during their early development. In separate experiments, embryos of 2-4 cell stage of M. mercenaria and C. virginica were exposed to both whole and lysed K. brevis cells isolated from Manasota Key, Florida. Low bloom concentrations of 500 to 3000 cells mL(-1) were simulated for 96 h. Shell length, percent abnormality (and normality), and percent mortality of resulting larvae were measured. Percentages were recorded after 6, 24, and 96 h of exposure; larval shell length was measured at 24 and 96 h. For both quahogs and oysters, the effects of exposing embryos to K. brevis on all larval responses were generally dose- and time-dependent. Percent mortalities and abnormalities of both clam and oyster embryos increased significantly after only 6h of exposure to whole cells of K. brevis. For clams, these parameters were significantly higher in whole and lysed treatments (at 3000 cells mL(-1)) than in controls. Percent mortalities of oysters were significantly higher in the whole-cell treatment (3000 cells mL(-1)) than under control conditions. After 24h of exposure, mean larval shell length of both bivalve species was significantly reduced relative to controls. This was evident for clam larvae in both the lysed treatment at 1500 cells mL(-1) and in whole and lysed treatments at 3000 cells mL(-1), and for oyster larvae in the lysed treatment at 3000 cells mL(-1). After 96 h, both species exposed to the lysed cell treatment at 3000 cells mL(-1) had significantly smaller larvae compared to those in the control. Overall, lysed cells of K. brevis had a more pronounced effect on shell length, percent abnormality, and mortality in both clams and oysters than did whole cells. Given the fact that blooms of K. brevis overlap with the spawning periods of these two bivalves, and that cells of this naked dinoflagellate are readily lysed by wave action, these results suggest that exposure to K. brevis during the early life history stages of clams and oysters could adversely affect their population recruitment. Further, the presence of whole or lysed cells of K. brevis in hatcheries could have a major negative impact on production. Copyright © 2014 Elsevier B.V. All rights reserved.

Three-Dimensional Imaging of the Mouse Organ of Corti Cytoarchitecture for Mechanical Modeling

NASA Astrophysics Data System (ADS)

Puria, Sunil; Hartman, Byron; Kim, Jichul; Oghalai, John S.; Ricci, Anthony J.; Liberman, M. Charles

2011-11-01

Cochlear models typically use continuous anatomical descriptions and homogenized parameters based on two-dimensional images for describing the organ of Corti. To produce refined models based more closely on the actual cochlear cytoarchitecture, three-dimensional morphometric parameters of key mechanical structures are required. Towards this goal, we developed and compared three different imaging methods: (1) A fixed cochlear whole-mount preparation using the fluorescent dye Cellmask®, which is a molecule taken up by cell membranes and clearly delineates Deiters' cells, outer hair cells, and the phalangeal process, imaged using confocal microscopy; (2) An in situ fixed preparation with hair cells labeled using anti-prestin and supporting structures labeled using phalloidin, imaged using two-photon microscopy; and (3) A membrane-tomato (mT) mouse with fluorescent proteins expressed in all cell membranes, which enables two-photon imaging of an in situ live preparation with excellent visualization of the organ of Corti. Morphometric parameters including lengths, diameters, and angles, were extracted from 3D cellular surface reconstructions of the resulting images. Preliminary results indicate that the length of the phalangeal processes decreases from the first (inner most) to third (outer most) row of outer hair cells, and that their length also likely varies from base to apex and across species.

Negative regulation of ciliary length by ciliary male germ cell-associated kinase (Mak) is required for retinal photoreceptor survival.

PubMed

Omori, Yoshihiro; Chaya, Taro; Katoh, Kimiko; Kajimura, Naoko; Sato, Shigeru; Muraoka, Koichiro; Ueno, Shinji; Koyasu, Toshiyuki; Kondo, Mineo; Furukawa, Takahisa

2010-12-28

Cilia function as cell sensors in many organs, and their disorders are referred to as "ciliopathies." Although ciliary components and transport machinery have been well studied, regulatory mechanisms of ciliary formation and maintenance are poorly understood. Here we show that male germ cell-associated kinase (Mak) regulates retinal photoreceptor ciliary length and subcompartmentalization. Mak was localized both in the connecting cilia and outer-segment axonemes of photoreceptor cells. In the Mak-null retina, photoreceptors exhibit elongated cilia and progressive degeneration. We observed accumulation of intraflagellar transport 88 (IFT88) and IFT57, expansion of kinesin family member 3A (Kif3a), and acetylated α-tubulin signals in the Mak-null photoreceptor cilia. We found abnormal rhodopsin accumulation in the Mak-null photoreceptor cell bodies at postnatal day 14. In addition, overexpression of retinitis pigmentosa 1 (RP1), a microtubule-associated protein localized in outer-segment axonemes, induced ciliary elongation, and Mak coexpression rescued excessive ciliary elongation by RP1. The RP1 N-terminal portion induces ciliary elongation and increased intensity of acetylated α-tubulin labeling in the cells and is phosphorylated by Mak. These results suggest that Mak is essential for the regulation of ciliary length and is required for the long-term survival of photoreceptors.

Negative regulation of ciliary length by ciliary male germ cell-associated kinase (Mak) is required for retinal photoreceptor survival

PubMed Central

Omori, Yoshihiro; Chaya, Taro; Katoh, Kimiko; Kajimura, Naoko; Sato, Shigeru; Muraoka, Koichiro; Ueno, Shinji; Koyasu, Toshiyuki; Kondo, Mineo; Furukawa, Takahisa

2010-01-01

Cilia function as cell sensors in many organs, and their disorders are referred to as “ciliopathies.” Although ciliary components and transport machinery have been well studied, regulatory mechanisms of ciliary formation and maintenance are poorly understood. Here we show that male germ cell-associated kinase (Mak) regulates retinal photoreceptor ciliary length and subcompartmentalization. Mak was localized both in the connecting cilia and outer-segment axonemes of photoreceptor cells. In the Mak-null retina, photoreceptors exhibit elongated cilia and progressive degeneration. We observed accumulation of intraflagellar transport 88 (IFT88) and IFT57, expansion of kinesin family member 3A (Kif3a), and acetylated α-tubulin signals in the Mak-null photoreceptor cilia. We found abnormal rhodopsin accumulation in the Mak-null photoreceptor cell bodies at postnatal day 14. In addition, overexpression of retinitis pigmentosa 1 (RP1), a microtubule-associated protein localized in outer-segment axonemes, induced ciliary elongation, and Mak coexpression rescued excessive ciliary elongation by RP1. The RP1 N-terminal portion induces ciliary elongation and increased intensity of acetylated α-tubulin labeling in the cells and is phosphorylated by Mak. These results suggest that Mak is essential for the regulation of ciliary length and is required for the long-term survival of photoreceptors. PMID:21148103

Light absorption cell combining variable path and length pump

DOEpatents

Prather, William S.

1993-01-01

A device for use in making spectrophotometric measurements of fluid samples. In particular, the device is a measurement cell containing a movable and a fixed lens with a sample of the fluid therebetween and through which light shines. The cell is connected to a source of light and a spectrophotometer via optic fibers. Movement of the lens varies the path length and also pumps the fluid into and out of the cell. Unidirectional inlet and exit valves cooperate with the movable lens to assure a one-way flow of fluid through the cell. A linear stepper motor controls the movement of the lens and cycles it from a first position closer to the fixed lens and a second position farther from the fixed lens, preferably at least 10 times per minute for a nearly continuous stream of absorption spectrum data.

Extreme Telomere Length Dimorphism in the Tasmanian Devil and Related Marsupials Suggests Parental Control of Telomere Length

PubMed Central

Bender, Hannah S.; Murchison, Elizabeth P.; Pickett, Hilda A.; Deakin, Janine E.; Strong, Margaret A.; Conlan, Carly; McMillan, Daniel A.; Neumann, Axel A.; Greider, Carol W.; Hannon, Gregory J.; Reddel, Roger R.; Graves, Jennifer A. Marshall.

2012-01-01

Telomeres, specialised structures that protect chromosome ends, play a critical role in preserving chromosome integrity. Telomere dynamics in the Tasmanian devil (Sarcophilus harrisii) are of particular interest in light of the emergence of devil facial tumour disease (DFTD), a transmissible malignancy that causes rapid mortality and threatens the species with extinction. We used fluorescent in situ hybridisation to investigate telomere length in DFTD cells, in healthy Tasmanian devils and in four closely related marsupial species. Here we report that animals in the Order Dasyuromorphia have chromosomes characterised by striking telomere length dimorphism between homologues. Findings in sex chromosomes suggest that telomere length dimorphism may be regulated by events in the parental germlines. Long telomeres on the Y chromosome imply that telomere lengthening occurs during spermatogenesis, whereas telomere diminution occurs during oogenesis. Although found in several somatic cell tissue types, telomere length dimorphism was not found in DFTD cancer cells, which are characterised by uniformly short telomeres. This is, to our knowledge, the first report of naturally occurring telomere length dimorphism in any species and suggests a novel strategy of telomere length control. Comparative studies in five distantly related marsupials and a monotreme indicate that telomere dimorphism evolved at least 50 million years ago. PMID:23049977

Telomere length dynamics in human memory T cells specific for viruses causing acute or latent infections

PubMed Central

2013-01-01

Background Declining telomere length (TL) is associated with T cell senescence. While TL in naïve and memory T cells declines with increasing age, there is limited data on TL dynamics in virus-specific memory CD4+ T cells in healthy adults. We combined BrdU-labeling of virus-stimulated T cells followed with flow cytometry-fluorescent in situ hybridization for TL determination. We analyzed TL in T cells specific for several virus infections: non-recurring acute (vaccinia virus, VACV), recurring-acute (influenza A virus, IAV), and reactivating viruses (varicella-zoster virus, VZV, and cytomegalovirus, CMV) in 10 healthy subjects. Additionally, five subjects provided multiple blood samples separated by up to 10 years. Results VACV- and CMV-specific T cells had longer average TL than IAV-specific CD4+ T cells. Although most virus-specific cells were CD45RA-, we observed a minor population of BrdU+ CD45RA+ T cells characterized by long telomeres. Longitudinal analysis demonstrated a slow decline in average TL in virus-specific T cells. However, in one subject, VZV reactivation led to an increase in average TL in VZV-specific memory T cells, suggesting a conversion of longer TL cells from the naïve T cell repertoire. Conclusions TLs in memory CD4+ T cells in otherwise healthy adults are heterogeneous and follow distinct virus-specific kinetics. These findings suggests that the distribution of TL and the creation and maintenance of long TL memory T cells could be important for the persistence of long-lived T cell memory. PMID:23971624

Telomere length dynamics in human memory T cells specific for viruses causing acute or latent infections.

PubMed

O'Bryan, Joel M; Woda, Marcia; Co, Mary; Mathew, Anuja; Rothman, Alan L

2013-08-26

Declining telomere length (TL) is associated with T cell senescence. While TL in naïve and memory T cells declines with increasing age, there is limited data on TL dynamics in virus-specific memory CD4+ T cells in healthy adults. We combined BrdU-labeling of virus-stimulated T cells followed with flow cytometry-fluorescent in situ hybridization for TL determination. We analyzed TL in T cells specific for several virus infections: non-recurring acute (vaccinia virus, VACV), recurring-acute (influenza A virus, IAV), and reactivating viruses (varicella-zoster virus, VZV, and cytomegalovirus, CMV) in 10 healthy subjects. Additionally, five subjects provided multiple blood samples separated by up to 10 years. VACV- and CMV-specific T cells had longer average TL than IAV-specific CD4+ T cells. Although most virus-specific cells were CD45RA-, we observed a minor population of BrdU+ CD45RA+ T cells characterized by long telomeres. Longitudinal analysis demonstrated a slow decline in average TL in virus-specific T cells. However, in one subject, VZV reactivation led to an increase in average TL in VZV-specific memory T cells, suggesting a conversion of longer TL cells from the naïve T cell repertoire. TLs in memory CD4+ T cells in otherwise healthy adults are heterogeneous and follow distinct virus-specific kinetics. These findings suggests that the distribution of TL and the creation and maintenance of long TL memory T cells could be important for the persistence of long-lived T cell memory.

Persistence and Adaptation in Immunity: T Cells Balance the Extent and Thoroughness of Search

PubMed Central

Fricke, G. Matthew; Letendre, Kenneth A.; Moses, Melanie E.; Cannon, Judy L.

2016-01-01

Effective search strategies have evolved in many biological systems, including the immune system. T cells are key effectors of the immune response, required for clearance of pathogenic infection. T cell activation requires that T cells encounter antigen-bearing dendritic cells within lymph nodes, thus, T cell search patterns within lymph nodes may be a crucial determinant of how quickly a T cell immune response can be initiated. Previous work suggests that T cell motion in the lymph node is similar to a Brownian random walk, however, no detailed analysis has definitively shown whether T cell movement is consistent with Brownian motion. Here, we provide a precise description of T cell motility in lymph nodes and a computational model that demonstrates how motility impacts T cell search efficiency. We find that both Brownian and Lévy walks fail to capture the complexity of T cell motion. Instead, T cell movement is better described as a correlated random walk with a heavy-tailed distribution of step lengths. Using computer simulations, we identify three distinct factors that contribute to increasing T cell search efficiency: 1) a lognormal distribution of step lengths, 2) motion that is directionally persistent over short time scales, and 3) heterogeneity in movement patterns. Furthermore, we show that T cells move differently in specific frequently visited locations that we call “hotspots” within lymph nodes, suggesting that T cells change their movement in response to the lymph node environment. Our results show that like foraging animals, T cells adapt to environmental cues, suggesting that adaption is a fundamental feature of biological search. PMID:26990103

Dynamic FtsA and FtsZ localization and outer membrane alterations during polar growth and cell division in Agrobacterium tumefaciens

PubMed Central

Zupan, John R.; Cameron, Todd A.; Anderson-Furgeson, James; Zambryski, Patricia C.

2013-01-01

Growth and cell division in rod-shaped bacteria have been primarily studied in species that grow predominantly by peptidoglycan (PG) synthesis along the length of the cell. Rhizobiales species, however, predominantly grow by PG synthesis at a single pole. Here we characterize the dynamic localization of several Agrobacterium tumefaciens components during the cell cycle. First, the lipophilic dye FM 4-64 predominantly stains the outer membranes of old poles versus growing poles. In cells about to divide, however, both poles are equally labeled with FM 4-64, but the constriction site is not. Second, the cell-division protein FtsA alternates from unipolar foci in the shortest cells to unipolar and midcell localization in cells of intermediate length, to strictly midcell localization in the longest cells undergoing septation. Third, the cell division protein FtsZ localizes in a cell-cycle pattern similar to, but more complex than, FtsA. Finally, because PG synthesis is spatially and temporally regulated during the cell cycle, we treated cells with sublethal concentrations of carbenicillin (Cb) to assess the role of penicillin-binding proteins in growth and cell division. Cb-treated cells formed midcell circumferential bulges, suggesting that interrupted PG synthesis destabilizes the septum. Midcell bulges contained bands or foci of FtsA-GFP and FtsZ-GFP and no FM 4-64 label, as in untreated cells. There were no abnormal morphologies at the growth poles in Cb-treated cells, suggesting unipolar growth uses Cb-insensitive PG synthesis enzymes. PMID:23674672

Gigantism in a Bacterium, Epulopiscium fishelsoni, Correlates with Complex Patterns in Arrangement, Quantity, and Segregation of DNA

PubMed Central

Bresler, V.; Montgomery, W. L.; Fishelson, L.; Pollak, P. E.

1998-01-01

Epulopiscium fishelsoni, gut symbiont of the brown surgeonfish (Acanthurus nigrofuscus) in the Red Sea, attains a larger size than any other eubacterium, varies 10- to 20-fold in length (and >2,000-fold in volume), and undergoes a complex daily life cycle. In early morning, nucleoids contain highly condensed DNA in elongate, chromosome-like structures which are physically separated from the general cytoplasm. Cell division involves production of two (rarely three) nucleoids within a cell, deposition of cell walls around expanded nucleoids, and emergence of daughter cells from the parent cell. Fluorescence measurements of DNA, RNA, and other cell components indicate the following. DNA quantity is proportional to cell volume over cell lengths of ∼30 μm to >500 μm. For cells of a given size, nucleoids of cells with two nucleoids (binucleoid) contain approximately equal amounts of DNA. And each nucleoid of a binucleoid cell contains one-half the DNA of the single nucleoid in a uninucleoid cell of the same size. The life cycle involves approximately equal subdivision of DNA among daughter cells, formation of apical caps of condensed DNA from previously decondensed and diffusely distributed DNA, and “pinching” of DNA near the middle of the cell in the absence of new wall formation. Mechanisms underlying these patterns remain unclear, but formation of daughter nucleoids and cells occurs both during diurnal periods of host feeding and bacterial cell growth and during nocturnal periods of host inactivity when mean bacterial cell size declines. PMID:9791108

Consequences of Location-Dependent Organ of Corti Micro-Mechanics

PubMed Central

Liu, Yanju; Gracewski, Sheryl M.; Nam, Jong-Hoon

2015-01-01

The cochlea performs frequency analysis and amplification of sounds. The graded stiffness of the basilar membrane along the cochlear length underlies the frequency-location relationship of the mammalian cochlea. The somatic motility of outer hair cell is central for cochlear amplification. Despite two to three orders of magnitude change in the basilar membrane stiffness, the force capacity of the outer hair cell’s somatic motility, is nearly invariant over the cochlear length. It is puzzling how actuators with a constant force capacity can operate under such a wide stiffness range. We hypothesize that the organ of Corti sets the mechanical conditions so that the outer hair cell’s somatic motility effectively interacts with the media of traveling waves—the basilar membrane and the tectorial membrane. To test this hypothesis, a computational model of the gerbil cochlea was developed that incorporates organ of Corti structural mechanics, cochlear fluid dynamics, and hair cell electro-physiology. The model simulations showed that the micro-mechanical responses of the organ of Corti are different along the cochlear length. For example, the top surface of the organ of Corti vibrated more than the bottom surface at the basal (high frequency) location, but the amplitude ratio was reversed at the apical (low frequency) location. Unlike the basilar membrane stiffness varying by a factor of 1700 along the cochlear length, the stiffness of the organ of Corti complex felt by the outer hair cell remained between 1.5 and 0.4 times the outer hair cell stiffness. The Y-shaped structure in the organ of Corti formed by outer hair cell, Deiters cell and its phalange was the primary determinant of the elastic reactance imposed on the outer hair cells. The stiffness and geometry of the Deiters cell and its phalange affected cochlear amplification differently depending on the location. PMID:26317521

Tau Fibril Formation in Cultured Cells Compatible with a Mouse Model of Tauopathy.

PubMed

Matsumoto, Gen; Matsumoto, Kazuki; Kimura, Taeko; Suhara, Tetsuya; Higuchi, Makoto; Sahara, Naruhiko; Mori, Nozomu

2018-05-17

Neurofibrillary tangles composed of hyperphosphorylated tau protein are primarily neuropathological features of a number of neurodegenerative diseases collectively termed tauopathy. To understand the mechanisms underlying the cause of tauopathy, precise cellular and animal models are required. Recent data suggest that the transient introduction of exogenous tau can accelerate the development of tauopathy in the brains of non-transgenic and transgenic mice expressing wild-type human tau. However, the transmission mechanism leading to tauopathy is not fully understood. In this study, we developed cultured-cell models of tauopathy representing a human tauopathy. Neuro2a (N2a) cells containing propagative tau filaments were generated by introducing purified tau fibrils. These cell lines expressed full-length (2N4R) human tau and the green fluorescent protein (GFP)-fused repeat domain of tau with P301L mutation. Immunocytochemistry and super-resolution microscopic imaging revealed that tau inclusions exhibited filamentous morphology and were composed of both full-length and repeat domain fragment tau. Live-cell imaging analysis revealed that filamentous tau inclusions are transmitted to daughter cells, resulting in yeast-prion-like propagation. By a standard method of tau preparation, both full-length tau and repeat domain fragments were recovered in sarkosyl insoluble fraction. Hyperphosphorylation of full-length tau was confirmed by the immunoreactivity of phospho-Tau antibodies and mobility shifts by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). These properties were similar to the biochemical features of P301L mutated human tau in a mouse model of tauopathy. In addition, filamentous tau aggregates in cells barely co-localized with ubiquitins, suggesting that most tau aggregates were excluded from protein degradation systems, and thus propagated to daughter cells. The present cellular model of tauopathy will provide an advantage for dissecting the mechanisms of tau aggregation and degradation and be a powerful tool for drug screening to prevent tauopathy.

Hsp47 mediates Cx43-dependent skeletal growth and patterning in the regenerating fin.

PubMed

Bhadra, Joyita; Iovine, M Kathryn

2015-11-01

Skeletal morphogenesis describes how bones achieve their correct shape and size and appropriately position joints. We use the regenerating caudal fin of zebrafish to study this process. Our examination of the fin length mutant short fin (sof (b123)) has revealed that the gap junction protein Cx43 is involved in skeletal morphogenesis by promoting cell proliferation and inhibiting joint formation, thereby coordinating skeletal growth and patterning. Here we demonstrate that serpinh1b is molecularly and functionally downstream of cx43. The gene serpinh1b codes for a protein called Hsp47, a molecular chaperone responsible for proper folding of procollagen molecules. Knockdown of Hsp47 in regenerating fins recapitulates the sof (b123) phenotypes of reduced fin length, reduced segment length and reduced level of cell proliferation. Furthermore, Hsp47 knockdown affects the organization and localization of the collagen-based actinotrichia. Together, our findings reveal that serpinh1b acts in a cx43 dependent manner to regulate cell proliferation and joint formation. We conclude that disruption of the collagen-based extracellular matrix influences signaling events required for cell proliferation, as well as the patterning of skeletal precursor cells that influences segment length. Therefore, we suggest that Hsp47 function is necessary for skeletal growth and patterning during fin regeneration. Copyright © 2015 Elsevier B.V. All rights reserved.

Analysis of Nuclear Factor-κB (NF-κB) Essential Modulator (NEMO) Binding to Linear and Lysine-linked Ubiquitin Chains and Its Role in the Activation of NF-κB*

PubMed Central

Kensche, Tobias; Tokunaga, Fuminori; Ikeda, Fumiyo; Goto, Eiji; Iwai, Kazuhiro; Dikic, Ivan

2012-01-01

Nuclear factor-κB (NF-κB) essential modulator (NEMO), a component of the inhibitor of κB kinase (IKK) complex, controls NF-κB signaling by binding to ubiquitin chains. Structural studies of NEMO provided a rationale for the specific binding between the UBAN (ubiquitin binding in ABIN and NEMO) domain of NEMO and linear (Met-1-linked) di-ubiquitin chains. Full-length NEMO can also interact with Lys-11-, Lys-48-, and Lys-63-linked ubiquitin chains of varying length in cells. Here, we show that purified full-length NEMO binds preferentially to linear ubiquitin chains in competition with lysine-linked ubiquitin chains of defined length, including long Lys-63-linked deca-ubiquitins. Linear di-ubiquitins were sufficient to activate both the IKK complex in vitro and to trigger maximal NF-κB activation in cells. In TNFα-stimulated cells, NEMO chimeras engineered to bind exclusively to Lys-63-linked ubiquitin chains mediated partial NF-κB activation compared with cells expressing NEMO that binds to linear ubiquitin chains. We propose that NEMO functions as a high affinity receptor for linear ubiquitin chains and a low affinity receptor for long lysine-linked ubiquitin chains. This phenomenon could explain quantitatively distinct NF-κB activation patterns in response to numerous cell stimuli. PMID:22605335

Well-defined block copolymers for gene delivery to dendritic cells: probing the effect of polycation chain-length.

PubMed

Tang, Rupei; Palumbo, R Noelle; Nagarajan, Lakshmi; Krogstad, Emily; Wang, Chun

2010-03-03

The development of safe and efficient polymer carriers for DNA vaccine delivery requires mechanistic understanding of structure-function relationship of the polymer carriers and their interaction with antigen-presenting cells. Here we have synthesized a series of diblock copolymers with well-defined chain-length using atom transfer radical polymerization and characterized the influence of polycation chain-length on the physico-chemical properties of the polymer/DNA complexes as well as the interaction with dendritic cells. The copolymers consist of a hydrophilic poly(ethylene glycol) block and a cationic poly(aminoethyl methacrylate) (PAEM) block. The average degree of polymerization (DP) of the PAEM block was varied among 19, 39, and 75, with nearly uniform distribution. With increasing PAEM chain-length, polyplexes formed by the diblock copolymers and plasmid DNA had smaller average particle size and showed higher stability against electrostatic destabilization by salt and heparin. The polymers were not toxic to mouse dendritic cells (DCs) and only displayed chain-length-dependent toxicity at a high concentration (1mg/mL). In vitro gene transfection efficiency and polyplex uptake in DCs were also found to correlate with chain-length of the PAEM block with the longer polymer chain favoring transfection and cellular uptake. The polyplexes induced a modest up-regulation of surface markers for DC maturation that was not significantly dependent on PAEM chain-length. Finally, the polyplex prepared from the longest PAEM block (DP of 75) achieved an average of 20% enhancement over non-condensed anionic dextran in terms of uptake by DCs in the draining lymph nodes 24h after subcutaneous injection into mice. Insights gained from studying such structurally well-defined polymer carriers and their interaction with dendritic cells may contribute to improved design of practically useful DNA vaccine delivery systems. Copyright 2009 Elsevier B.V. All rights reserved.

Telomere dynamics in an immortal human cell line.

PubMed Central

Murnane, J P; Sabatier, L; Marder, B A; Morgan, W F

1994-01-01

The integration of transfected plasmid DNA at the telomere of chromosome 13 in an immortalized simian virus 40-transformed human cell line provided the first opportunity to study polymorphism in the number of telomeric repeat sequences on the end of a single chromosome. Three subclones of this cell line were selected for analysis: one with a long telomere on chromosome 13, one with a short telomere, and one with such extreme polymorphism that no distinct band was discernible. Further subcloning demonstrated that telomere polymorphism resulted from both gradual changes and rapid changes that sometimes involved many kilobases. The gradual changes were due to the shortening of telomeres at a rate similar to that reported for telomeres of somatic cells without telomerase, eventually resulting in the loss of nearly all of the telomere. However, telomeres were not generally lost completely, as shown by the absence of polymorphism in the subtelomeric plasmid sequences. Instead, telomeres that were less than a few hundred base pairs in length showed a rapid, highly heterogeneous increase in size. Rapid changes in telomere length also occurred on longer telomeres. The frequency of this type of change in telomere length varied among the subclones and correlated with chromosome fusion. Therefore, the rapid changes in telomere length appeared occasionally to result in the complete loss of telomeric repeat sequences. Rapid changes in telomere length have been associated with telomere loss and chromosome instability in yeast and could be responsible for the high rate of chromosome fusion observed in many human tumor cell lines. Images PMID:7957062

Programmable display of DNA-protein chimeras for controlling cell-hydrogel interactions via reversible intermolecular hybridization.

PubMed

Zhang, Zhaoyang; Li, Shihui; Chen, Niancao; Yang, Cheng; Wang, Yong

2013-04-08

Extensive studies have been recently carried out to achieve dynamic control of cell-material interactions primarily through physicochemical stimulation. The purpose of this study was to apply reversible intermolecular hybridization to program cell-hydrogel interactions in physiological conditions based on DNA-antibody chimeras and complementary oligonucleotides. The results showed that DNA oligonucleotides could be captured to and released from the immobilizing DNA-functionalized hydrogels with high specificity via DNA hybridization. Accordingly, DNA-antibody chimeras were captured to the hydrogels, successfully inducing specific cell attachment. The cell attachment to the hydrogels reached the plateau at approximately half an hour after the functionalized hydrogels and the cells were incubated together. The attached cells were rapidly released from the bound hydrogels when triggering complementary oligonucleotides were introduced to the system. However, the capability of the triggering complementary oligonucleotides in releasing cells was affected by the length of intermolecular hybridization. The length needed to be at least more than 20 base pairs in the current experimental setting. Notably, because the procedure of intermolecular hybridization did not involve any harsh condition, the released cells maintained the same viability as that of the cultured cells. The functionalized hydrogels also exhibited the potential to catch and release cells repeatedly. Therefore, this study demonstrates that it is promising to regulate cell-material interactions dynamically through the DNA-programmed display of DNA-protein chimeras.

Geometry and network connectivity govern the mechanics of stress fibers.

PubMed

Kassianidou, Elena; Brand, Christoph A; Schwarz, Ulrich S; Kumar, Sanjay

2017-03-07

Actomyosin stress fibers (SFs) play key roles in driving polarized motility and generating traction forces, yet little is known about how tension borne by an individual SF is governed by SF geometry and its connectivity to other cytoskeletal elements. We now address this question by combining single-cell micropatterning with subcellular laser ablation to probe the mechanics of single, geometrically defined SFs. The retraction length of geometrically isolated SFs after cutting depends strongly on SF length, demonstrating that longer SFs dissipate more energy upon incision. Furthermore, when cell geometry and adhesive spacing are fixed, cell-to-cell heterogeneities in SF dissipated elastic energy can be predicted from varying degrees of physical integration with the surrounding network. We apply genetic, pharmacological, and computational approaches to demonstrate a causal and quantitative relationship between SF connectivity and mechanics for patterned cells and show that similar relationships hold for nonpatterned cells allowed to form cell-cell contacts in monolayer culture. Remarkably, dissipation of a single SF within a monolayer induces cytoskeletal rearrangements in cells long distances away. Finally, stimulation of cell migration leads to characteristic changes in network connectivity that promote SF bundling at the cell rear. Our findings demonstrate that SFs influence and are influenced by the networks in which they reside. Such higher order network interactions contribute in unexpected ways to cell mechanics and motility.

Flagellum Density Regulates Proteus mirabilis Swarmer Cell Motility in Viscous Environments

PubMed Central

Tuson, Hannah H.; Copeland, Matthew F.; Carey, Sonia; Sacotte, Ryan

2013-01-01

Proteus mirabilis is an opportunistic pathogen that is frequently associated with urinary tract infections. In the lab, P. mirabilis cells become long and multinucleate and increase their number of flagella as they colonize agar surfaces during swarming. Swarming has been implicated in pathogenesis; however, it is unclear how energetically costly changes in P. mirabilis cell morphology translate into an advantage for adapting to environmental changes. We investigated two morphological changes that occur during swarming—increases in cell length and flagellum density—and discovered that an increase in the surface density of flagella enabled cells to translate rapidly through fluids of increasing viscosity; in contrast, cell length had a small effect on motility. We found that swarm cells had a surface density of flagella that was ∼5 times larger than that of vegetative cells and were motile in fluids with a viscosity that inhibits vegetative cell motility. To test the relationship between flagellum density and velocity, we overexpressed FlhD4C2, the master regulator of the flagellar operon, in vegetative cells of P. mirabilis and found that increased flagellum density produced an increase in cell velocity. Our results establish a relationship between P. mirabilis flagellum density and cell motility in viscous environments that may be relevant to its adaptation during the infection of mammalian urinary tracts and movement in contact with indwelling catheters. PMID:23144253

Full-length single-cell RNA-seq applied to a viral human cancer: applications to HPV expression and splicing analysis in HeLa S3 cells.

PubMed

Wu, Liang; Zhang, Xiaolong; Zhao, Zhikun; Wang, Ling; Li, Bo; Li, Guibo; Dean, Michael; Yu, Qichao; Wang, Yanhui; Lin, Xinxin; Rao, Weijian; Mei, Zhanlong; Li, Yang; Jiang, Runze; Yang, Huan; Li, Fuqiang; Xie, Guoyun; Xu, Liqin; Wu, Kui; Zhang, Jie; Chen, Jianghao; Wang, Ting; Kristiansen, Karsten; Zhang, Xiuqing; Li, Yingrui; Yang, Huanming; Wang, Jian; Hou, Yong; Xu, Xun

2015-01-01

Viral infection causes multiple forms of human cancer, and HPV infection is the primary factor in cervical carcinomas. Recent single-cell RNA-seq studies highlight the tumor heterogeneity present in most cancers, but virally induced tumors have not been studied. HeLa is a well characterized HPV+ cervical cancer cell line. We developed a new high throughput platform to prepare single-cell RNA on a nanoliter scale based on a customized microwell chip. Using this method, we successfully amplified full-length transcripts of 669 single HeLa S3 cells and 40 of them were randomly selected to perform single-cell RNA sequencing. Based on these data, we obtained a comprehensive understanding of the heterogeneity of HeLa S3 cells in gene expression, alternative splicing and fusions. Furthermore, we identified a high diversity of HPV-18 expression and splicing at the single-cell level. By co-expression analysis we identified 283 E6, E7 co-regulated genes, including CDC25, PCNA, PLK4, BUB1B and IRF1 known to interact with HPV viral proteins. Our results reveal the heterogeneity of a virus-infected cell line. It not only provides a transcriptome characterization of HeLa S3 cells at the single cell level, but is a demonstration of the power of single cell RNA-seq analysis of virally infected cells and cancers.

The sexually dimorphic adipose fin is an androgen target tissue in the brown trout (Salmo trutta fario).

PubMed

Hisar, Olcay; Sönmez, Adem Yavuz; Hisar, Şükriye Aras; Budak, Harun; Gültepe, Nejdet

2013-04-01

An investigation has been described on the relationship of body length, age and sex with adipose fin length and the number of androgen receptor (AR)-containing cells in the adipose fin as a secondary sexual characteristic for brown trout (Salmo trutta fario). Firstly, body and adipose fin lengths of 2- to 5-year-old brown trout were measured. Thereafter, these fish were killed by decapitation, then their sexes were determined, and adipose fins were excised. The cellular bases of AR binding activities in the adipose fins were analyzed with an antibody against human/rat AR peptide. Immunocytochemistry and western blotting techniques were performed with this antibody. Analysis of morphological measurements indicated that body length and age had a linear relationship with adipose fin length. The coefficients of determination for the body length and age were 0.92 and 0.85 in the male fish and 0.76 and 0.73 in the female fish against the adipose fin length, respectively. At 2 years of age, cells in the adipose fin did not exhibit AR immunoreactivity. However, AR-immunopositive cells were abundant in the adipose fin of 3- to 5-year-old fish. Moreover, the number of AR-immunopositive cells was significantly (P < 0.05) high in males and increased with age. These observations indicate that the adipose fin in the brown trout is a probable target for androgen action and that tissue function or development may to some extent be androgen dependent. In addition, it is likely that such an effect will be mediated by specific androgen receptors.

Therapeutic analysis of high-dose-rate {sup 192}Ir vaginal cuff brachytherapy for endometrial cancer using a cylindrical target volume model and varied cancer cell distributions

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

Zhang, Hualin, E-mail: hualin.zhang@northwestern.edu; Donnelly, Eric D.; Strauss, Jonathan B.

Purpose: To evaluate high-dose-rate (HDR) vaginal cuff brachytherapy (VCBT) in the treatment of endometrial cancer in a cylindrical target volume with either a varied or a constant cancer cell distributions using the linear quadratic (LQ) model. Methods: A Monte Carlo (MC) technique was used to calculate the 3D dose distribution of HDR VCBT over a variety of cylinder diameters and treatment lengths. A treatment planning system (TPS) was used to make plans for the various cylinder diameters, treatment lengths, and prescriptions using the clinical protocol. The dwell times obtained from the TPS were fed into MC. The LQ model wasmore » used to evaluate the therapeutic outcome of two brachytherapy regimens prescribed either at 0.5 cm depth (5.5 Gy × 4 fractions) or at the vaginal mucosal surface (8.8 Gy × 4 fractions) for the treatment of endometrial cancer. An experimentally determined endometrial cancer cell distribution, which showed a varied and resembled a half-Gaussian distribution, was used in radiobiology modeling. The equivalent uniform dose (EUD) to cancer cells was calculated for each treatment scenario. The therapeutic ratio (TR) was defined by comparing VCBT with a uniform dose radiotherapy plan in term of normal cell survival at the same level of cancer cell killing. Calculations of clinical impact were run twice assuming two different types of cancer cell density distributions in the cylindrical target volume: (1) a half-Gaussian or (2) a uniform distribution. Results: EUDs were weakly dependent on cylinder size, treatment length, and the prescription depth, but strongly dependent on the cancer cell distribution. TRs were strongly dependent on the cylinder size, treatment length, types of the cancer cell distributions, and the sensitivity of normal tissue. With a half-Gaussian distribution of cancer cells which populated at the vaginal mucosa the most, the EUDs were between 6.9 Gy × 4 and 7.8 Gy × 4, the TRs were in the range from (5.0){sup 4} to (13.4){sup 4} for the radiosensitive normal tissue depending on the cylinder size, treatment lengths, prescription depth, and dose as well. However, for a uniform cancer cell distribution, the EUDs were between 6.3 Gy × 4 and 7.1 Gy × 4, and the TRs were found to be between (1.4){sup 4} and (1.7){sup 4}. For the uniformly interspersed cancer and radio-resistant normal cells, the TRs were less than 1. The two VCBT prescription regimens were found to be equivalent in terms of EUDs and TRs. Conclusions: HDR VCBT strongly favors cylindrical target volume with the cancer cell distribution following its dosimetric trend. Assuming a half-Gaussian distribution of cancer cells, the HDR VCBT provides a considerable radiobiological advantage over the external beam radiotherapy (EBRT) in terms of sparing more normal tissues while maintaining the same level of cancer cell killing. But for the uniform cancer cell distribution and radio-resistant normal tissue, the radiobiology outcome of the HDR VCBT does not show an advantage over the EBRT. This study strongly suggests that radiation therapy design should consider the cancer cell distribution inside the target volume in addition to the shape of target.« less

Cleavage of the urokinase receptor (uPAR) on oral cancer cells: regulation by transforming growth factor - β1 (TGF-β1) and potential effects on migration and invasion.

PubMed

Magnussen, Synnove Norvoll; Hadler-Olsen, Elin; Costea, Daniela Elena; Berg, Eli; Jacobsen, Cristiane Cavalcanti; Mortensen, Bente; Salo, Tuula; Martinez-Zubiaurre, Inigo; Winberg, Jan-Olof; Uhlin-Hansen, Lars; Svineng, Gunbjorg

2017-05-19

Urokinase plasminogen activator (uPA) receptor (uPAR) is up-regulated at the invasive tumour front of human oral squamous cell carcinoma (OSCC), indicating a role for uPAR in tumour progression. We previously observed elevated expression of uPAR at the tumour-stroma interface in a mouse model for OSCC, which was associated with increased proteolytic activity. The tumour microenvironment regulated uPAR expression, as well as its glycosylation and cleavage. Both full-length- and cleaved uPAR (uPAR (II-III)) are involved in highly regulated processes such as cell signalling, proliferation, migration, stem cell mobilization and invasion. The aim of the current study was to analyse tumour associated factors and their effect on uPAR cleavage, and the potential implications for cell proliferation, migration and invasion. Mouse uPAR was stably overexpressed in the mouse OSCC cell line AT84. The ratio of full-length versus cleaved uPAR as analysed by Western blotting and its regulation was assessed by addition of different protease inhibitors and transforming growth factor - β1 (TGF-β1). The role of uPAR cleavage in cell proliferation and migration was analysed using real-time cell analysis and invasion was assessed using the myoma invasion model. We found that when uPAR was overexpressed a proportion of the receptor was cleaved, thus the cells presented both full-length uPAR and uPAR (II-III). Cleavage was mainly performed by serine proteases and urokinase plasminogen activator (uPA) in particular. When the OSCC cells were stimulated with TGF-β1, the production of the uPA inhibitor PAI-1 was increased, resulting in a reduction of uPAR cleavage. By inhibiting cleavage of uPAR, cell migration was reduced, and by inhibiting uPA activity, invasion was reduced. We could also show that medium containing soluble uPAR (suPAR), and cleaved soluble uPAR (suPAR (II-III)), induced migration in OSCC cells with low endogenous levels of uPAR. These results show that soluble factors in the tumour microenvironment, such as TGF-β1, PAI-1 and uPA, can influence the ratio of full length and uPAR (II-III) and thereby potentially effect cell migration and invasion. Resolving how uPAR cleavage is controlled is therefore vital for understanding how OSCC progresses and potentially provides new targets for therapy.

Efficient and Extensible Quasi-Explicit Modular Nonlinear Multiscale Battery Model: GH-MSMD

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

Kim, Gi-Heon; Smith, Kandler; Lawrence-Simon, Jake

Complex physics and long computation time hinder the adoption of computer aided engineering models in the design of large-format battery cells and systems. A modular, efficient battery simulation model -- the multiscale multidomain (MSMD) model -- was previously introduced to aid the scale-up of Li-ion material and electrode designs to complete cell and pack designs, capturing electrochemical interplay with 3-D electronic current pathways and thermal response. Here, this paper enhances the computational efficiency of the MSMD model using a separation of time-scales principle to decompose model field variables. The decomposition provides a quasi-explicit linkage between the multiple length-scale domains andmore » thus reduces time-consuming nested iteration when solving model equations across multiple domains. In addition to particle-, electrode- and cell-length scales treated in the previous work, the present formulation extends to bus bar- and multi-cell module-length scales. We provide example simulations for several variants of GH electrode-domain models.« less

Efficient and Extensible Quasi-Explicit Modular Nonlinear Multiscale Battery Model: GH-MSMD

DOE PAGES

Kim, Gi-Heon; Smith, Kandler; Lawrence-Simon, Jake; ...

2017-03-24

Complex physics and long computation time hinder the adoption of computer aided engineering models in the design of large-format battery cells and systems. A modular, efficient battery simulation model -- the multiscale multidomain (MSMD) model -- was previously introduced to aid the scale-up of Li-ion material and electrode designs to complete cell and pack designs, capturing electrochemical interplay with 3-D electronic current pathways and thermal response. Here, this paper enhances the computational efficiency of the MSMD model using a separation of time-scales principle to decompose model field variables. The decomposition provides a quasi-explicit linkage between the multiple length-scale domains andmore » thus reduces time-consuming nested iteration when solving model equations across multiple domains. In addition to particle-, electrode- and cell-length scales treated in the previous work, the present formulation extends to bus bar- and multi-cell module-length scales. We provide example simulations for several variants of GH electrode-domain models.« less

Diffusion length measurements of thin GaAs solar cells by means of energetic electrons

NASA Technical Reports Server (NTRS)

Vonross, O.

1980-01-01

A calculation of the short circuit current density (j sub sc) of a thin GaAs solar cell induced by fast electrons is presented. It is shown that in spite of the disparity in thickness between the N-type portion of the junction and the P-type portion of the junction, the measurement of the bulk diffusion length L sub p of the N-type part of the junction is seriously hampered due to the presence of a sizable contribution to the j sub sc from the P-type region of the junction. Corrections of up to 50% had to be made in order to interpret the data correctly. Since these corrections were not amenable to direct measurements it is concluded that the electron beam method for the determination of the bulk minority carrier diffusion length, which works so well for Si solar cells, is a poor method when applied to thin GaAs cells.

OmpA: A Flexible Clamp for Bacterial Cell Wall Attachment.

PubMed

Samsudin, Firdaus; Ortiz-Suarez, Maite L; Piggot, Thomas J; Bond, Peter J; Khalid, Syma

2016-12-06

The envelope of Gram-negative bacteria is highly complex, containing separate outer and inner membranes and an intervening periplasmic space encompassing a peptidoglycan (PGN) cell wall. The PGN scaffold is anchored non-covalently to the outer membrane via globular OmpA-like domains of various proteins. We report atomically detailed simulations of PGN bound to OmpA in three different states, including the isolated C-terminal domain (CTD), the full-length monomer, or the complete full-length dimeric form. Comparative analysis of dynamics of OmpA CTD from different bacteria helped to identify a conserved PGN-binding mode. The dynamics of full-length OmpA, embedded within a realistic representation of the outer membrane containing full-rough (Ra) lipopolysaccharide, phospholipids, and cardiolipin, suggested how the protein may provide flexible mechanical support to the cell wall. An accurate model of the heterogeneous bacterial cell envelope should facilitate future efforts to develop antibacterial agents. Copyright © 2016 Elsevier Ltd. All rights reserved.

Length of intact plasma membrane determines the diffusion properties of cellular water.

PubMed

Eida, Sato; Van Cauteren, Marc; Hotokezaka, Yuka; Katayama, Ikuo; Sasaki, Miho; Obara, Makoto; Okuaki, Tomoyuki; Sumi, Misa; Nakamura, Takashi

2016-01-11

Molecular diffusion in a boundary-free medium depends only on the molecular size, the temperature, and medium viscosity. However, the critical determinant of the molecular diffusion property in inhomogeneous biological tissues has not been identified. Here, using an in vitro system and a high-resolution MR imaging technique, we show that the length of the intact plasma membrane is a major determinant of water diffusion in a controlled cellular environment and that the cell perimeter length (CPL) is sufficient to estimate the apparent diffusion coefficient (ADC) of water in any cellular environment in our experimental system (ADC = -0.21 × CPL + 1.10). We used this finding to further explain the different diffusion kinetics of cells that are dying via apoptotic or non-apoptotic cell death pathways exhibiting characteristic changes in size, nuclear and cytoplasmic architectures, and membrane integrity. These results suggest that the ADC value can be used as a potential biomarker for cell death.

Length of intact plasma membrane determines the diffusion properties of cellular water

PubMed Central

Eida, Sato; Van Cauteren, Marc; Hotokezaka, Yuka; Katayama, Ikuo; Sasaki, Miho; Obara, Makoto; Okuaki, Tomoyuki; Sumi, Misa; Nakamura, Takashi

2016-01-01

Molecular diffusion in a boundary-free medium depends only on the molecular size, the temperature, and medium viscosity. However, the critical determinant of the molecular diffusion property in inhomogeneous biological tissues has not been identified. Here, using an in vitro system and a high-resolution MR imaging technique, we show that the length of the intact plasma membrane is a major determinant of water diffusion in a controlled cellular environment and that the cell perimeter length (CPL) is sufficient to estimate the apparent diffusion coefficient (ADC) of water in any cellular environment in our experimental system (ADC = −0.21 × CPL + 1.10). We used this finding to further explain the different diffusion kinetics of cells that are dying via apoptotic or non-apoptotic cell death pathways exhibiting characteristic changes in size, nuclear and cytoplasmic architectures, and membrane integrity. These results suggest that the ADC value can be used as a potential biomarker for cell death. PMID:26750342

Quantitative analysis of nanoscale intranuclear structural alterations in hippocampal cells in chronic alcoholism via transmission electron microscopy imaging.

PubMed

Sahay, Peeyush; Shukla, Pradeep K; Ghimire, Hemendra M; Almabadi, Huda M; Tripathi, Vibha; Mohanty, Samarendra K; Rao, Radhakrishna; Pradhan, Prabhakar

2017-03-01

Chronic alcoholism is known to alter the morphology of the hippocampus, an important region of cognitive function in the brain. Therefore, to understand the effect of chronic alcoholism on hippocampal neural cells, we employed a mouse model of chronic alcoholism and quantified intranuclear nanoscale structural alterations in these cells. Transmission electron microscopy (TEM) images of hippocampal neurons were obtained, and the degree of structural alteration in terms of mass density fluctuation was determined using the light-localization properties of optical media generated from TEM imaging. The results, which were obtained at length scales ranging from ~30 to 200 nm, show that 10-12 week-old mice fed a Lieber-DeCarli liquid (alcoholic) diet had a higher degree of structural alteration than control mice fed a normal diet without alcohol. The degree of structural alteration became significantly distinguishable at a sample length of ~100 nm, which is the typical length scale of the building blocks of cells, such as DNA, RNA, proteins and lipids. Interestingly, different degrees of structural alteration at such length scales suggest possible structural rearrangement of chromatin inside the nuclei in chronic alcoholism.

Quantitative analysis of nanoscale intranuclear structural alterations in hippocampal cells in chronic alcoholism via transmission electron microscopy imaging

NASA Astrophysics Data System (ADS)

Sahay, Peeyush; Shukla, Pradeep K.; Ghimire, Hemendra M.; Almabadi, Huda M.; Tripathi, Vibha; Mohanty, Samarendra K.; Rao, Radhakrishna; Pradhan, Prabhakar

2017-04-01

Chronic alcoholism is known to alter the morphology of the hippocampus, an important region of cognitive function in the brain. Therefore, to understand the effect of chronic alcoholism on hippocampal neural cells, we employed a mouse model of chronic alcoholism and quantified intranuclear nanoscale structural alterations in these cells. Transmission electron microscopy (TEM) images of hippocampal neurons were obtained, and the degree of structural alteration in terms of mass density fluctuation was determined using the light-localization properties of optical media generated from TEM imaging. The results, which were obtained at length scales ranging from ~30 to 200 nm, show that 10-12 week-old mice fed a Lieber-DeCarli liquid (alcoholic) diet had a higher degree of structural alteration than control mice fed a normal diet without alcohol. The degree of structural alteration became significantly distinguishable at a sample length of ~100 nm, which is the typical length scale of the building blocks of cells, such as DNA, RNA, proteins and lipids. Interestingly, different degrees of structural alteration at such length scales suggest possible structural rearrangement of chromatin inside the nuclei in chronic alcoholism.

Interrogating the Escherichia coli cell cycle by cell dimension perturbations

PubMed Central

Zheng, Hai; Ho, Po-Yi; Jiang, Meiling; Tang, Bin; Liu, Weirong; Li, Dengjin; Yu, Xuefeng; Kleckner, Nancy E.; Amir, Ariel; Liu, Chenli

2016-01-01

Bacteria tightly regulate and coordinate the various events in their cell cycles to duplicate themselves accurately and to control their cell sizes. Growth of Escherichia coli, in particular, follows a relation known as Schaechter’s growth law. This law says that the average cell volume scales exponentially with growth rate, with a scaling exponent equal to the time from initiation of a round of DNA replication to the cell division at which the corresponding sister chromosomes segregate. Here, we sought to test the robustness of the growth law to systematic perturbations in cell dimensions achieved by varying the expression levels of mreB and ftsZ. We found that decreasing the mreB level resulted in increased cell width, with little change in cell length, whereas decreasing the ftsZ level resulted in increased cell length. Furthermore, the time from replication termination to cell division increased with the perturbed dimension in both cases. Moreover, the growth law remained valid over a range of growth conditions and dimension perturbations. The growth law can be quantitatively interpreted as a consequence of a tight coupling of cell division to replication initiation. Thus, its robustness to perturbations in cell dimensions strongly supports models in which the timing of replication initiation governs that of cell division, and cell volume is the key phenomenological variable governing the timing of replication initiation. These conclusions are discussed in the context of our recently proposed “adder-per-origin” model, in which cells add a constant volume per origin between initiations and divide a constant time after initiation. PMID:27956612

Interrogating the Escherichia coli cell cycle by cell dimension perturbations.

PubMed

Zheng, Hai; Ho, Po-Yi; Jiang, Meiling; Tang, Bin; Liu, Weirong; Li, Dengjin; Yu, Xuefeng; Kleckner, Nancy E; Amir, Ariel; Liu, Chenli

2016-12-27

Bacteria tightly regulate and coordinate the various events in their cell cycles to duplicate themselves accurately and to control their cell sizes. Growth of Escherichia coli, in particular, follows a relation known as Schaechter's growth law. This law says that the average cell volume scales exponentially with growth rate, with a scaling exponent equal to the time from initiation of a round of DNA replication to the cell division at which the corresponding sister chromosomes segregate. Here, we sought to test the robustness of the growth law to systematic perturbations in cell dimensions achieved by varying the expression levels of mreB and ftsZ We found that decreasing the mreB level resulted in increased cell width, with little change in cell length, whereas decreasing the ftsZ level resulted in increased cell length. Furthermore, the time from replication termination to cell division increased with the perturbed dimension in both cases. Moreover, the growth law remained valid over a range of growth conditions and dimension perturbations. The growth law can be quantitatively interpreted as a consequence of a tight coupling of cell division to replication initiation. Thus, its robustness to perturbations in cell dimensions strongly supports models in which the timing of replication initiation governs that of cell division, and cell volume is the key phenomenological variable governing the timing of replication initiation. These conclusions are discussed in the context of our recently proposed "adder-per-origin" model, in which cells add a constant volume per origin between initiations and divide a constant time after initiation.

Telomere dynamics and homeostasis in a transmissible cancer.

PubMed

Ujvari, Beata; Pearse, Anne-Maree; Taylor, Robyn; Pyecroft, Stephen; Flanagan, Cassandra; Gombert, Sara; Papenfuss, Anthony T; Madsen, Thomas; Belov, Katherine

2012-01-01

Devil Facial Tumour Disease (DFTD) is a unique clonal cancer that threatens the world's largest carnivorous marsupial, the Tasmanian devil (Sarcophilus harrisii) with extinction. This transmissible cancer is passed between individual devils by cell implantation during social interactions. The tumour arose in a Schwann cell of a single devil over 15 years ago and since then has expanded clonally, without showing signs of replicative senescence; in stark contrast to a somatic cell that displays a finite capacity for replication, known as the "Hayflick limit". In the present study we investigate the role of telomere length, measured as Telomere Copy Number (TCN), and telomerase and shelterin gene expression, as well as telomerase activity in maintaining hyperproliferation of Devil Facial Tumour (DFT) cells. Our results show that DFT cells have short telomeres. DFTD TCN does not differ between geographic regions or between strains. However, TCN has increased over time. Unlimited cell proliferation is likely to have been achieved through the observed up-regulation of the catalytic subunit of telomerase (TERT) and concomitant activation of telomerase. Up-regulation of the central component of shelterin, the TRF1-intercating nuclear factor 2 (TINF2) provides DFT a mechanism for telomere length homeostasis. The higher expression of both TERT and TINF2 may also protect DFT cells from genomic instability and enhance tumour proliferation. DFT cells appear to monitor and regulate the length of individual telomeres: i.e. shorter telomeres are elongated by up-regulation of telomerase-related genes; longer telomeres are protected from further elongation by members of the shelterin complex, which may explain the lack of spatial and strain variation in DFT telomere copy number. The observed longitudinal increase in gene expression in DFT tissue samples and telomerase activity in DFT cell lines might indicate a selection for more stable tumours with higher proliferative potential.

Performance of OSC's initial Amtec generator design, and comparison with JPL's Europa Orbiter goals

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

Schock, A.; Noravian, H.; Or, C.

1998-07-01

The procedure for the analysis (with overpotential correction) of multitube AMTEC (Alkali Metal Thermal-to-Electrical Conversion) cells described in Paper IECEC 98-243 was applied to a wide range of multicell radioisotope space power systems. System design options consisting of one or two generators, each with 2, 3, or 4 stacked GPHS (General Purpose Heat Source) modules, identical to those used on previous NASA missions, were analyzed and performance-mapped. The initial generators analyzed by OSC had 8 AMTEC cells on each end of the heat source stack, with five beta-alumina solid electrolyte (BASE) tubes per cell. The heat source and converters inmore » the Orbital generator designs are embedded in a thermal insulation system consisting of Min-K fibrous insulation surrounded by graded-length molybdenum multifoils. Detailed analyses in previous Orbital studies found that such an insulation system could reduce extraneous heat losses to about 10%. For the above design options, the present paper presents the system mass and performance (i.e., the EOM system efficiency and power output and the BOM evaporator and clad temperatures) for a wide range of heat inputs and load voltages, and compares the results with JPL's preliminary goals for the Europa Orbiter mission to be launched in November 2003. The analytical results showed that the initial 16-cell generator designs resulted in either excessive evaporator and clad temperatures and/or insufficient power outputs to meet the JPL-specified mission goals. The computed performance of modified OSC generators with different numbers of AMTEC cells, cell diameters, cell lengths, cell materials, BASE tube lengths, and number of tubes per cell are described in Paper IECEC.98.245 in these proceedings.« less

Retinoic Acid Improves Morphology of Cultured Peritoneal Mesothelial Cells from Patients Undergoing Dialysis

PubMed Central

Retana, Carmen; Sanchez, Elsa I.; Gonzalez, Sirenia; Perez-Lopez, Alejandro; Cruz, Armando; Lagunas-Munoz, Jesus; Alfaro-Cruz, Carmen; Vital-Flores, Socorro; Reyes, José L.

2013-01-01

Patients undergoing continuous ambulatory peritoneal dialysis are classified according to their peritoneal permeability as low transporter (low solute permeability) or High transporter (high solute permeability). Factors that determine the differences in permeability between them have not been fully disclosed. We investigated morphological features of cultured human peritoneal mesothelial cells from low or high transporter patients and its response to All trans retinoic Acid (ATRA, vitamin A active metabolite), as compared to non-uremic human peritoneal mesothelial cells. Control cells were isolated from human omentum. High or low transporter cells were obtained from dialysis effluents. Cells were cultured in media containing ATRA (0, 50, 100 or 200 nM). We studied length and distribution of microvilli and cilia (scanning electron microscopy), epithelial (cytokeratin, claudin-1, ZO-1 and occludin) and mesenchymal (vimentin and α-smooth muscle actin) transition markers by immunofluorescence and Western blot, and transforming growth factor β1 expression by Western blot. Low and high transporter exhibited hypertrophic cells, reduction in claudin-1, occludin and ZO-1 expression, cytokeratin and vimentin disorganization and positive α-smooth muscle actin label. Vimentin, α-smooth muscle actin and transforming growth factor- β1 were overexpressed in low transporter. Ciliated cells were diminished in low and high transporters. Microvilli number and length were severely reduced in high transporter. ATRA reduced hypertrophic cells number in low transporter. It also improved cytokeratin and vimentin organization, decreased vimentin and α-smooth muscle actin expression, and increased claudin 1, occludin and ZO-1 expression, in low and high transporter. In low transporter, ATRA reduced transforming growth factor-β1 expression. ATRA augmented percentage of ciliated cells in low and high transporter. It also augmented cilia length in high transporter. Alterations in structure, epithelial mesenchymal markers and transforming growth factor-β1expression were differential between low and high transporter. Beneficial effects of ATRA were improved human peritoneal mesothelial cells morphology tending to normalize structures. PMID:24223992

Spiroplasma swim by a processive change in body helicity.

NASA Astrophysics Data System (ADS)

Shaevitz, Joshua

2006-03-01

Microscopic organisms must rely on very different strategies than their macroscopic counterparts to swim through liquid. To date, the best understood method for prokaryotic swimming employs the rotation of flagella. I will present data that Spiroplasma, tiny helical bacteria that infect plants and insects, use a very different approach. By measuring cell kinematics during free swimming, we find that propulsion is generated by the propagation of kink pairs down the length of the cell body. A processive change in the helicity of the body creates these waves and enables directional movement. Unlike the motion of other helical swimmers such as Spirochetes, Spiroplasma swimming velocity increases with increasing viscosity. In addition, cell morphological parameters such as helical pitch and cell length influence swimming velocity.

Thermal ablation therapeutics based on CNx multi-walled nanotubes

PubMed Central

Torti, Suzy V; Byrne, Fiona; Whelan, Orla; Levi, Nicole; Ucer, Burak; Schmid, Michael; Torti, Frank M; Akman, Steven; Liu, Jiwen; Ajayan, Pulickel M; Nalamasu, Omkaram; Carroll, David L

2007-01-01

We demonstrate that nitrogen doped, multi-walled carbon nanotubes (CNx-MWNT) result in photo-ablative destruction of kidney cancer cells when excited by near infrared (NIR) irradiation. Further, we show that effective heat transduction and cellular cytotoxicity depends on nanotube length: effective NIR coupling occurs at nanotube lengths that exceed half the wavelength of the stimulating radiation, as predicted in classical antenna theory. We also demonstrate that this radiation heats the nanotubes through induction processes, resulting in significant heat transfer to surrounding media and cell killing at extraordinarily small radiation doses. This cell death was attributed directly to photothermal effect generated within the culture, since neither the infrared irradiation itself nor the CNx-MWNT were toxic to the cells. PMID:18203437

Cavity ring-down spectroscopy in the liquid phase

NASA Astrophysics Data System (ADS)

Xu, Shucheng; Sha, Guohe; Xie, Jinchun

2002-02-01

A new application for cavity ring-down spectroscopic (CRDS) technique using a pulsed polarized light source has been developed in the absorption measurement of liquids for "colorless" organic compounds using both a single sample cell and double sample cells inserted in an optical cavity at Brewster angle. At present an experimental capability of measuring absorption coefficients as small as 2-5×10-7 cm-1 has been demonstrated by measurement of the absorption baselines. The first spectra for CRDS in the liquid phase, the C-H stretching fifth vibrational overtones of benzene in the pure liquid and hexane solution are obtained. The optical absorption length for liquids in both a single sample cell and double sample cells of 1 cm length is up to 900 cm due to multipass of light within an optical cavity. Compared to the thermal lens and optoacoustic spectroscopic techniques, the sensitivity for CRDS mainly depends on the optical absorption path of the sample (single passing path of the sample times multipass times), is not determined by the laser power and the length of the sample cell. The absolute absorption coefficient and band intensity for the sample are determined directly by the spectroscopy.

Telomere lengthening early in development.

PubMed

Liu, Lin; Bailey, Susan M; Okuka, Maja; Muñoz, Purificación; Li, Chao; Zhou, Lingjun; Wu, Chao; Czerwiec, Eva; Sandler, Laurel; Seyfang, Andreas; Blasco, Maria A; Keefe, David L

2007-12-01

Stem cells and cancer cells maintain telomere length mostly through telomerase. Telomerase activity is high in male germ line and stem cells, but is low or absent in mature oocytes and cleavage stage embryos, and then high again in blastocysts. How early embryos reset telomere length remains poorly understood. Here, we show that oocytes actually have shorter telomeres than somatic cells, but their telomeres lengthen remarkably during early cleavage development. Moreover, parthenogenetically activated oocytes also lengthen their telomeres, thus the capacity to elongate telomeres must reside within oocytes themselves. Notably, telomeres also elongate in the early cleavage embryos of telomerase-null mice, demonstrating that telomerase is unlikely to be responsible for the abrupt lengthening of telomeres in these cells. Coincident with telomere lengthening, extensive telomere sister-chromatid exchange (T-SCE) and colocalization of the DNA recombination proteins Rad50 and TRF1 were observed in early cleavage embryos. Both T-SCE and DNA recombination proteins decrease in blastocyst stage embryos, whereas telomerase activity increases and telomeres elongate only slowly. We suggest that telomeres lengthen during the early cleavage cycles following fertilization through a recombination-based mechanism, and that from the blastocyst stage onwards, telomerase only maintains the telomere length established by this alternative mechanism.

Cloning of calves from various somatic cell types of male and female adult, newborn and fetal cows.

PubMed

Kato, Y; Tani, T; Tsunoda, Y

2000-11-01

Twenty-four calves were cloned from six somatic cell types of female and male adult, newborn and fetal cows. The clones were derived from female cumulus (n = 3), oviduct (n = 2) and uterine (n = 2) cells, female and male skin cells (n = 10), and male ear (n = 5) and liver (n = 2) cells. On the basis of the number of cloned embryos transferred (n = 172) to surrogate cows, the overall rate of success was 14%, but based on the number of surrogate mothers that became pregnant (n = 50), the success rate was 48%. Cell nuclei from uterus, ear and liver cells, which have not been tested previously, developed into newborn calves after nuclear transfer into enucleated oocytes. To date, seven female and six male calves have survived: six of the females were from adult cells (cumulus (n = 3), oviduct (n = 2) and skin (n = 1) cells) and one was from newborn skin cells, whereas the male calves were derived from adult ear cells (n = 3), newborn liver and skin cells (n = 2), and fetal cells (n = 1). Clones derived from adult cells frequently aborted in the later stages of pregnancy and calves developing to term showed a higher number of abnormalities than did those derived from newborn or fetal cells. The telomeric DNA lengths in the ear cells of three male calves cloned from the ear cells of a bull aged 10 years were similar to those of the original bull. However, the telomeric DNA lengths from the white blood cells of the clones, although similar to those in an age-matched control, were shorter than those of the original bull, which indicates that telomeric shortening varies among tissues.

A nonlinear cochlear model with the outer hair cell piezoelectric activity

NASA Astrophysics Data System (ADS)

Jiang, Xiaoai; Grosh, Karl

2003-10-01

In this paper we present a simple cochlear model which captures the most important aspect of nonlinearity in the cochlea-the nonlinearity caused by the piezoelectric-like activity of outer hair cells and the variable conductance of the outer hair cell stereocilia. A one-dimensional long-wave model is built to simulate the dynamic response of the fluid-loaded basilar membrane. The basilar membrane is simulated as isolated linear oscillators along the cochlear length, and its motion is coupled with the fluid pressure and the nonlinear force produced by the outer hair cells. As the basilar membrane moves, the fluid shears stereocilia, and the resulting ion flow changes the transmembrane potential of the outer hair cells and subsequently their length, leading to further movement of the basilar membrane. The piezoelectric-like activity of the outer hair cell is simulated by a current source, and stereocilia motion is modeled as a varying conductance that changes as the basilar membrane moves. A solution in the time domain will be presented. [Work supported by NIH.

Herpesvirus papio: state and properties of intracellular viral DNA in baboon lymphoblastoid cell lines.

PubMed

Falk, L; Lindahl, T; Bjursell, G; Klein, G

1979-07-15

Herpesvirus papio (HVP) is an indigenous B-lymphotropic virus of baboons (Papio sp.) present in latent form in baboon lymphoblastoid cell lines. It shares cross-reacting viral capsid and early antigens with the Epstein-Barr virus (EBV), and HVP DNA and EBV DNA show partial sequence homology. EBV-specific complementary RNA was employed here as a probe to investigate the physical state of the HVP DNA component in baboon lymphoblastoid cells after fractionation of cellular DNA by density gradient centrifugation. Five virus-producing cultures contained both free and integrated HVP DNA sequences while one non-producing cell line had two or three viral genome equivalents per cell in an apparently integrated form. Further analysis of one virus-producing line showed that the free HVP DNA fraction was composed of both linear and circular viral DNA. Contour length measurements of HVP circular DNA molecules by electron microscopy revealed that they were similar in length to the EBV circular DNA present in human lymphoblastoid cells.

Characterizing Spatial Organization of Cell Surface Receptors in Human Breast Cancer with STORM

NASA Astrophysics Data System (ADS)

Lyall, Evan; Chapman, Matthew R.; Sohn, Lydia L.

2012-02-01

Regulation and control of complex biological functions are dependent upon spatial organization of biological structures at many different length scales. For instance Eph receptors and their ephrin ligands bind when opposing cells come into contact during development, resulting in spatial organizational changes on the nanometer scale that lead to changes on the macro scale, in a process known as organ morphogenesis. One technique able to probe this important spatial organization at both the nanometer and micrometer length scales, including at cell-cell junctions, is stochastic optical reconstruction microscopy (STORM). STORM is a technique that localizes individual fluorophores based on the centroids of their point spread functions and then reconstructs a composite image to produce super resolved structure. We have applied STORM to study spatial organization of the cell surface of human breast cancer cells, specifically the organization of tyrosine kinase receptors and chemokine receptors. A better characterization of spatial organization of breast cancer cell surface proteins is necessary to fully understand the tumorigenisis pathways in the most common malignancy in United States women.

Effects of spaceflight on the proliferation of jejunal mucosal cells

NASA Technical Reports Server (NTRS)

Phillips, Robert W.; Moeller, C. L.; Sawyer, Heywood R.; Smirnov, K. L.

1991-01-01

The purpose of this project was to test the hypothesis that the generalized, whole body decrease in synthetic activity due to microgravity conditions encountered during spaceflight would be demonstrable in cells and tissues characterized by a rapid rate of turnover. Jejunal mucosal cells were chosen as a model since these cells are among the most rapidly proliferating in the body. Accordingly, the percentage of mitotic cells present in the crypts of Lieberkuhn in each of 5 rats flown on the COSMOS 2044 mission were compared to the percentage of mitotic cells present in the crypts in rats included in each of 3 ground control groups (i.e., vivarium, synchronous and caudal-elevated). No significant difference (p greater than .05) was detected in mitotic indices between the flight and vivarium group. Although the ability of jejunal mucosal cells to divide by mitosis was not impaired in flight group, there was, however, a reduction in the length of villi and depth of crypts. The concommitant reduction in villus length and crypth depth in the flight group probably reflects changes in connective tissue components within the core of villi.

Quantitative proteomic analysis of human breast epithelial cells with differential telomere length

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

Yu, Li-Rong; Chan, King C.; Tahara, Hidetoshi

Telomeres play important functional roles in cell proliferation, cell cycle regulation, and genetic stability, in which telomere length is critical. In this study, quantitative proteome comparisons for the human breast epithelial cells with short and long telomeres (184-hTERT{sub L} vs. 184-hTERT{sub S} and 90P-hTERT{sub L} vs. 90P-hTERT{sub S}), resulting from transfection of the human telomerase reverse transcriptase (hTERT) gene, were performed using cleavable isotope-coded affinity tags. More than 2000 proteins were quantified in each comparative experiment, with approximately 77% of the proteins identified in both analyses. In the cells with long telomeres, significant and consistent alterations were observed in metabolismmore » (amino acid, nucleotide, and lipid metabolism), genetic information transmission (transcription and translation regulation, spliceosome and ribosome complexes), and cell signaling. Interestingly, the DNA excision repair pathway is enhanced, while integrin and its ligands are downregulated in the cells with long telomeres. These results may provide valuable information related to telomere functions.« less

Clonal expansion of genome-intact HIV-1 in functionally polarized Th1 CD4+ T cells

PubMed Central

Orlova-Fink, Nina; Einkauf, Kevin; Chowdhury, Fatema Z.; Sun, Xiaoming; Harrington, Sean; Kuo, Hsiao-Hsuan; Hua, Stephane; Chen, Hsiao-Rong; Ouyang, Zhengyu; Reddy, Kavidha; Dong, Krista; Ndung’u, Thumbi; Walker, Bruce D.; Rosenberg, Eric S.; Yu, Xu G.

2017-01-01

HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells. PMID:28628034

Clonal expansion of genome-intact HIV-1 in functionally polarized Th1 CD4+ T cells.

PubMed

Lee, Guinevere Q; Orlova-Fink, Nina; Einkauf, Kevin; Chowdhury, Fatema Z; Sun, Xiaoming; Harrington, Sean; Kuo, Hsiao-Hsuan; Hua, Stephane; Chen, Hsiao-Rong; Ouyang, Zhengyu; Reddy, Kavidha; Dong, Krista; Ndung'u, Thumbi; Walker, Bruce D; Rosenberg, Eric S; Yu, Xu G; Lichterfeld, Mathias

2017-06-30

HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells.

Effusion cytomorphology of small round cell tumors.

PubMed

Ikeda, Katsuhide; Tsuta, Koji

2016-01-01

Small round cell tumors (SRCTs) are a group of tumors composed of small, round, and uniform cells with high nuclear/cytoplasmic (N/C) ratios. The appearance of SRCT neoplastic cells in the effusion fluid is very rare. We reported the cytomorphological findings of SRCTs in effusion cytology, and performed statistical and mathematical analyses for a purpose to distinguish SRCTs. We analyzed the cytologic findings of effusion samples from 40 SRCT cases and measured the lengths of the nuclei, cytoplasms, and the cell cluster areas. The SRCT cases included 14 Ewing sarcoma (EWS)/primitive neuroectodermal tumor cases, 5 synovial sarcoma cases, 6 rhabdomyosarcoma cases, 9 small cell lung carcinoma (SCLC) cases, and 6 diffuse large B-cell lymphoma (DLBL) cases. Morphologically, there were no significant differences in the nuclear and cytoplasmic lengths in cases of EWS, synovial sarcoma, and rhabdomyosarcoma. The cytoplasmic lengths in cases of SCLC and DLBL were smaller than those of EWS, synovial sarcoma, and rhabdomyosarcoma. The nuclear density of the cluster in SCLC was higher than that in other SRCTs, and cases of DLBL showed a lack of anisokaryosis and anisocytosis. We believe that it might be possible to diagnose DLBL and SCLC from cytologic analysis of effusion samples but it is very difficult to use this method to distinguish EWS, synovial sarcoma, and rhabdomyosarcoma. Statistical and mathematical analyses indicated that nuclear density and dispersion of nuclear and cytoplasmic sizes are useful adjuncts to conventional cytologic diagnostic criteria, which are acquired from experience.

Geometry and network connectivity govern the mechanics of stress fibers

PubMed Central

Kassianidou, Elena; Brand, Christoph A.; Kumar, Sanjay

2017-01-01

Actomyosin stress fibers (SFs) play key roles in driving polarized motility and generating traction forces, yet little is known about how tension borne by an individual SF is governed by SF geometry and its connectivity to other cytoskeletal elements. We now address this question by combining single-cell micropatterning with subcellular laser ablation to probe the mechanics of single, geometrically defined SFs. The retraction length of geometrically isolated SFs after cutting depends strongly on SF length, demonstrating that longer SFs dissipate more energy upon incision. Furthermore, when cell geometry and adhesive spacing are fixed, cell-to-cell heterogeneities in SF dissipated elastic energy can be predicted from varying degrees of physical integration with the surrounding network. We apply genetic, pharmacological, and computational approaches to demonstrate a causal and quantitative relationship between SF connectivity and mechanics for patterned cells and show that similar relationships hold for nonpatterned cells allowed to form cell–cell contacts in monolayer culture. Remarkably, dissipation of a single SF within a monolayer induces cytoskeletal rearrangements in cells long distances away. Finally, stimulation of cell migration leads to characteristic changes in network connectivity that promote SF bundling at the cell rear. Our findings demonstrate that SFs influence and are influenced by the networks in which they reside. Such higher order network interactions contribute in unexpected ways to cell mechanics and motility. PMID:28213499

Lack of hormone binding in COS-7 cells expressing a mutated growth hormone receptor found in Laron dwarfism.

PubMed Central

Edery, M; Rozakis-Adcock, M; Goujon, L; Finidori, J; Lévi-Meyrueis, C; Paly, J; Djiane, J; Postel-Vinay, M C; Kelly, P A

1993-01-01

A single point mutation in the growth hormone (GH) receptor gene generating a Phe-->Ser substitution in the extracellular binding domain of the receptor has been identified in one family with Laron type dwarfism. The mutation was introduced by site-directed mutagenesis into cDNAs encoding the full-length rabbit GH receptor and the extracellular domain or binding protein (BP) of the human and rabbit GH receptor, and also in cDNAs encoding the full length and the extracellular domain of the related rabbit prolactin (PRL) receptor. All constructs were transiently expressed in COS-7 cells. Both wild type and mutant full-length rabbit GH and PRL receptors, as well as GH and prolactin BPs (wild type and mutant), were detected by Western blot in cell membranes and concentrated culture media, respectively. Immunofluorescence studies showed that wild type and mutant full-length GH receptors had the same cell surface and intracellular distribution and were expressed with comparable intensities. In contrast, all mutant forms (full-length receptors or BPs), completely lost their modify the synthesis ligand. These results clearly demonstrate that this point mutation (patients with Laron syndrome) does not modify the synthesis or the intracellular pathway of receptor proteins, but rather abolishes ability of the receptor or BP to bind GH and is thus responsible for the extreme GH resistance in these patients. Images PMID:8450064

Roles of telomeres and telomerase in cancer, and advances in telomerase-targeted therapies.

PubMed

Jafri, Mohammad A; Ansari, Shakeel A; Alqahtani, Mohammed H; Shay, Jerry W

2016-06-20

Telomeres maintain genomic integrity in normal cells, and their progressive shortening during successive cell divisions induces chromosomal instability. In the large majority of cancer cells, telomere length is maintained by telomerase. Thus, telomere length and telomerase activity are crucial for cancer initiation and the survival of tumors. Several pathways that regulate telomere length have been identified, and genome-scale studies have helped in mapping genes that are involved in telomere length control. Additionally, genomic screening for recurrent human telomerase gene hTERT promoter mutations and mutations in genes involved in the alternative lengthening of telomeres pathway, such as ATRX and DAXX, has elucidated how these genomic changes contribute to the activation of telomere maintenance mechanisms in cancer cells. Attempts have also been made to develop telomere length- and telomerase-based diagnostic tools and anticancer therapeutics. Recent efforts have revealed key aspects of telomerase assembly, intracellular trafficking and recruitment to telomeres for completing DNA synthesis, which may provide novel targets for the development of anticancer agents. Here, we summarize telomere organization and function and its role in oncogenesis. We also highlight genomic mutations that lead to reactivation of telomerase, and mechanisms of telomerase reconstitution and trafficking that shed light on its function in cancer initiation and tumor development. Additionally, recent advances in the clinical development of telomerase inhibitors, as well as potential novel targets, will be summarized.

Elevation effects in volcano applications of the COSPEC

USGS Publications Warehouse

Gerlach, T.M.

2003-01-01

Volcano applications commonly involve sizeable departures from the reference pressure and temperature of COSPEC calibration cells. Analysis shows that COSPEC SO2 column abundances and derived mass emission rates are independent of pressure and temperature, and thus unaffected by elevation effects related to deviations from calibration cell reference state. However, path-length concentrations are pressure and temperature dependent. Since COSPEC path-length concentration data assume the reference pressure and temperature of calibration cells, they can lead to large errors when used to calculate SO2 mixing ratios of volcanic plumes. Correction factors for COSPEC path-length concentrations become significant (c.10%) at elevations of about 1 km (e.g. Kilauea volcano) and rise rapidly to c.80% at 6 km (e.g. Cotopaxi volcano). Calculating SO2 mixing ratios for volcanic plumes directly from COSPEC path-length concentrations always gives low results. Corrections can substantially increase mixing ratios; for example, corrections increase SO2 ppm concentrations reported for the Mount St Helens, Colima, and Erebus plumes by 25-50%. Several arguments suggest it would be advantageous to calibrate COSPEC measurements in column abundance units rather than path-length concentration units.

Understanding the effect of alkyl chains of gemini cations on the physicochemical and cellular properties of polyurethane micelles.

PubMed

Pan, Zhicheng; Fang, Danxuan; Song, Yuanqing; Song, Nijia; Ding, Mingming; Li, Jiehua; Luo, Feng; Li, Jianshu; Tan, Hong; Fu, Qiang

2018-06-06

Cationic gemini quaternary ammonium (GQA) has been used as a cell internalization promoter to improve the permeability of the cell membrane and enhance the cellular uptake. However, the effect of the alkyl chain length on the cellular properties of nanocarriers has not been elucidated yet. In this study, we developed a series of polyurethane micelles containing GQAs with various alkyl chain lengths. The alteration of the gemini alkyl chain length was found to change the distribution of GQA surfactants in the micellar structure and affect the surface charge exposure, stability, and the protein absorption properties of nanocarriers. Moreover, we also clarified the role of the alkyl chain length in tumor cell internalization and macrophage uptake of polyurethane micelles. This work provides a new understanding on the effect of the GQA alkyl chain length on the physicochemical and biological properties of nanomedicines, and offers guidance on the rational design of effective drug delivery systems where the issue of functional group exposure at the micellar surface should be considered.

Accurate determination of lattice parameters based on Niggli reduced cell theory by using digitized electron diffraction micrograph.

PubMed

Yang, Yi; Cai, Canying; Lin, Jianguo; Gong, Lunjun; Yang, Qibin

2017-05-01

In this paper, we used Niggli reduced cell theory to determine lattice constants of a micro/nano crystal by using electron diffraction patterns. The Niggli reduced cell method enhanced the accuracy of lattice constant measurement obviously, because the lengths and the angles of lattice vectors of a primitive cell can be measured directly on the electron micrographs instead of a double tilt holder. With the aid of digitized algorithm and least square optimization by using three digitized micrographs, a valid reciprocal Niggli reduced cell number can be obtained. Thus a reciprocal and real Bravais lattices are acquired. The results of three examples, i.e., Mg 4 Zn 7 , an unknown phase (Precipitate phase in nickel-base superalloy) and Ba 4 Ti 13 O 30 showed that the maximum errors are 1.6% for lengths and are 0.3% for angles. Copyright © 2017 Elsevier Ltd. All rights reserved.

An EBIC equation for solar cells. [Electron Beam Induced Current

NASA Technical Reports Server (NTRS)

Luke, K. L.; Von Roos, O.

1983-01-01

When an electron beam of a scanning electron microscope (SEM) impinges on an N-P junction, the generation of electron-hole pairs by impact ionization causes a characteristic short circuit current I(sc) to flow. The I(sc), i.e., EBIC (electron beam induced current) depends strongly on the configuration used to investigate the cell's response. In this paper the case where the plane of the junction is perpendicular to the surface is considered. An EBIC equation amenable to numerical computations is derived as a function of cell thickness, source depth, surface recombination velocity, diffusion length, and distance of the junction to the beam-cell interaction point for a cell with an ohmic contact at its back surface. It is shown that the EBIC equation presented here is more general and easier to use than those previously reported. The effects of source depth, ohmic contact, and diffusion length on the normalized EBIC characteristic are discussed.

Multi-classification of cell deformation based on object alignment and run length statistic.

PubMed

Li, Heng; Liu, Zhiwen; An, Xing; Shi, Yonggang

2014-01-01

Cellular morphology is widely applied in digital pathology and is essential for improving our understanding of the basic physiological processes of organisms. One of the main issues of application is to develop efficient methods for cell deformation measurement. We propose an innovative indirect approach to analyze dynamic cell morphology in image sequences. The proposed approach considers both the cellular shape change and cytoplasm variation, and takes each frame in the image sequence into account. The cell deformation is measured by the minimum energy function of object alignment, which is invariant to object pose. Then an indirect analysis strategy is employed to overcome the limitation of gradual deformation by run length statistic. We demonstrate the power of the proposed approach with one application: multi-classification of cell deformation. Experimental results show that the proposed method is sensitive to the morphology variation and performs better than standard shape representation methods.

Light absorption cell combining variable path and length pump

DOEpatents

Prather, W.S.

1993-12-07

A device is described for use in making spectrophotometric measurements of fluid samples. In particular, the device is a measurement cell containing a movable and a fixed lens with a sample of the fluid there between and through which light shines. The cell is connected to a source of light and a spectrophotometer via optic fibers. Movement of the lens varies the path length and also pumps the fluid into and out of the cell. Unidirectional inlet and exit valves cooperate with the movable lens to assure a one-way flow of fluid through the cell. A linear stepper motor controls the movement of the lens and cycles it from a first position closer to the fixed lens and a second position farther from the fixed lens, preferably at least 10 times per minute for a nearly continuous stream of absorption spectrum data. 2 figures.

Low interleukin-2 concentration favors generation of early memory T cells over effector phenotypes during chimeric antigen receptor T-cell expansion.

PubMed

Kaartinen, Tanja; Luostarinen, Annu; Maliniemi, Pilvi; Keto, Joni; Arvas, Mikko; Belt, Heini; Koponen, Jonna; Loskog, Angelica; Mustjoki, Satu; Porkka, Kimmo; Ylä-Herttuala, Seppo; Korhonen, Matti

2017-06-01

Adoptive T-cell therapy offers new options for cancer treatment. Clinical results suggest that T-cell persistence, depending on T-cell memory, improves efficacy. The use of interleukin (IL)-2 for in vitro T-cell expansion is not straightforward because it drives effector T-cell differentiation but does not promote the formation of T-cell memory. We have developed a cost-effective expansion protocol for chimeric antigen receptor (CAR) T cells with an early memory phenotype. Lymphocytes were transduced with third-generation lentiviral vectors and expanded using CD3/CD28 microbeads. The effects of altering the IL-2 supplementation (0-300 IU/mL) and length of expansion (10-20 days) on the phenotype of the T-cell products were analyzed. High IL-2 levels led to a decrease in overall generation of early memory T cells by both decreasing central memory T cells and augmenting effectors. T memory stem cells (T SCM , CD95 + CD45RO - CD45RA + CD27 + ) were present variably during T-cell expansion. However, their presence was not IL-2 dependent but was linked to expansion kinetics. CD19-CAR T cells generated in these conditions displayed in vitro antileukemic activity. In summary, production of CAR T cells without any cytokine supplementation yielded the highest proportion of early memory T cells, provided a 10-fold cell expansion and the cells were functionally potent. The number of early memory T cells in a T-cell preparation can be increased by simply reducing the amount of IL-2 and limiting the length of T-cell expansion, providing cells with potentially higher in vivo performance. These findings are significant for robust and cost-effective T-cell manufacturing. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Mechanisms of nanoclay-enhanced plastic foaming processes: effects of nanoclay intercalation and exfoliation

NASA Astrophysics Data System (ADS)

Wong, Anson; Wijnands, Stephan F. L.; Kuboki, Takashi; Park, Chul B.

2013-08-01

The foaming behaviors of high-density polypropylene-nanoclay composites with intercalated and exfoliated nanoclay particles blown with carbon dioxide were examined via in situ observation of the foaming processes in a high-temperature/high-pressure view-cell. The intercalated nanoclay particles were 300-600 nm in length and 50-200 nm in thickness, while the exfoliated nanoclay particles were 100-200 nm in length and 1 nm in thickness. Contrary to common belief, it was discovered that intercalated nanoclay yielded higher cell density than exfoliated nanoclay despite its lower particle density. This was attributed to the higher tensile stresses generated around the larger and stiffer intercalated nanoclay particles, which led to increase in supersaturation level for cell nucleation. Also, the coupling agent used to exfoliate nanoclay would increase the affinity between polymer and surface of nanoclay particles. Consequently, the critical work needed for cell nucleation would be increased; pre-existing microvoids, which could act as seeds for cell nucleation, were also less likely to exist. Meanwhile, exfoliated nanoclay had better cell stabilization ability to prevent cell coalescence and cell coarsening. This investigation clarifies the roles of nanoclay in plastic foaming processes and provides guidance for the advancement of polymer nanocomposite foaming technology.

Construction of recombinant FGFR1 containing full-length gene and its potential application.

PubMed

Zhou, Yali; Luo, Wenjuan; Zheng, Lei; Li, Miao; Zhang, Yanmin

2010-07-01

FGFR1, one of the four fibroblast growth factor receptors, has been found to be over-expressed in many cancers. In this study, a full-length expression plasmid for FGFR1 was obtained by fragment amplification. The amplified PCR product was then digested and inserted into the pcDNA3.1(+) vector. A recombinant eukaryotic expression vector containing the complete CDS region of FGFR1 was successfully constructed. After it was transfected to Hek293 cell, the expression of the FGFR1 receptor in recombinant Hek293/FGFR1 was 18 times higher than that of Hek293 cell. The biological activities of high expression FGFR1 cell (Hek293/FGFR1) were verified by FCM, immunofluorescent, RT-PCR, western blot and cell cycle analysis. Then, Hek293/FGFR1 was used to screen taspine with cell membrane chromatography (CMC). Finally, we analyzed the effects of taspine on Hek293/FGFR1 cell and MCF-7 cell. In conclusion, Hek293/FGFR1 was successfully constructed. The results demonstrate that taspine can down-regulate phosphorylation of FGFR1 and ERK, and inhibit Hek293/FGFR1 and MCF-7 cell proliferation. Copyright 2010 Elsevier Inc. All rights reserved.

Telomerecat: A ploidy-agnostic method for estimating telomere length from whole genome sequencing data.

PubMed

Farmery, James H R; Smith, Mike L; Lynch, Andy G

2018-01-22

Telomere length is a risk factor in disease and the dynamics of telomere length are crucial to our understanding of cell replication and vitality. The proliferation of whole genome sequencing represents an unprecedented opportunity to glean new insights into telomere biology on a previously unimaginable scale. To this end, a number of approaches for estimating telomere length from whole-genome sequencing data have been proposed. Here we present Telomerecat, a novel approach to the estimation of telomere length. Previous methods have been dependent on the number of telomeres present in a cell being known, which may be problematic when analysing aneuploid cancer data and non-human samples. Telomerecat is designed to be agnostic to the number of telomeres present, making it suited for the purpose of estimating telomere length in cancer studies. Telomerecat also accounts for interstitial telomeric reads and presents a novel approach to dealing with sequencing errors. We show that Telomerecat performs well at telomere length estimation when compared to leading experimental and computational methods. Furthermore, we show that it detects expected patterns in longitudinal data, repeated measurements, and cross-species comparisons. We also apply the method to a cancer cell data, uncovering an interesting relationship with the underlying telomerase genotype.

System Design, Implementation, and Evaluation of the Optical Broadband Correlator

DTIC Science & Technology

1994-09-20

shear-mode TeO2 , Model No. N45075-6-20, manufactured by Newport Electro- Optic Systems with a length of 75 pjs, acoustic direction 1110], optical...optical aperture (or useful length) TOA of our cells are shown in Table 3. The Bragg cells are shear-mode TeO2 , Model No. N45075-6-20, manufactured by...focusing or integrating (Fourier transform) lens is a laser diode glass doublet Model 06LAI013/076, from Melles Griot. Its focal length is 145 nun at 830

Three-phase boundary length in solid-oxide fuel cells: A mathematical model

NASA Astrophysics Data System (ADS)

Janardhanan, Vinod M.; Heuveline, Vincent; Deutschmann, Olaf

A mathematical model to calculate the volume specific three-phase boundary length in the porous composite electrodes of solid-oxide fuel cell is presented. The model is exclusively based on geometrical considerations accounting for porosity, particle diameter, particle size distribution, and solids phase distribution. Results are presented for uniform particle size distribution as well as for non-uniform particle size distribution.

21 CFR 172.886 - Petroleum wax.

Code of Federal Regulations, 2014 CFR

2014-04-01

... hole bored in the center to closely fit the stem of the chromatographic tube. Heating jacket. Conical... joints. Spectrophotometric cells. Fused quartz cells, optical path length in the range of 5.000... the range 1.000 centimeter ±0.005 centimeter. With distilled water in the cells, determine any...

Use of back-scatter electron signals to visualise cell/nanowires interactions in vitro and in vivo; frustrated phagocytosis of long fibres in macrophages and compartmentalisation in mesothelial cells in vivo

PubMed Central

2012-01-01

Background Frustrated phagocytosis has been stated as an important factor in the initiation of an inflammatory response after fibre exposure. The length of fibrous structures has been linked to the potential of fibres to induce adverse health effects for at least 40 years. However, we only recently reported for the first time the threshold length for fibre-induced inflammation in the pleural space and we implicated frustrated phagocytosis in the pro-inflammatory effects of long fibres. This study extends the examination of the threshold value for frustrated phagocytosis using well-defined length classes of silver nanowires (AgNW) ranging from 3–28 μm and describes in detail the morphology of frustrated phagocytosis using a novel technique and also describes compartmentalisation of fibres in the pleural space. Methods A novel technique, backscatter scanning electron microscopy (BSE) was used to study frustrated phagocytosis since it provides high-contrast detection of nanowires, allowing clear discrimination between the nanofibres and other cellular features. A human monocyte-derived macrophage cell line THP-1 was used to investigate cell-nanowire interaction in vitro and the parietal pleura, the site of fibre retention after inhalation exposure was chosen to visualise the cell- fibre interaction in vivo after direct pleural installation of AgNWs. Results The length cut-off value for frustrated phagocytosis differs in vitro and in vivo. While in vitro frustrated phagocytosis could be observed with fibres ≥14 μm, in vivo studies showed incomplete uptake at a fibre length of ≥10 μm. Recently we showed that inflammation in the pleural space after intrapleural injection of the same nanofibre panel occurs at a length of ≥5 μm. This onset of inflammation does not correlate with the onset of frustrated phagocytosis as shown in this study, leading to the conclusion that intermediate length fibres fully enclosed within macrophages as well as frustrated phagocytosis are associated with a pro-inflammatory state in the pleural space. We further showed that fibres compartmentalise in the mesothelial cells at the parietal pleura as well as in inflammatory cells in the pleural space. Conclusion BSE is a useful way to clearly distinguish between fibres that are, or are not, membrane-bounded. Using this method we were able to show differences in the threshold length at which frustrated phagocytosis occurred between in vitro and in vivo models. Visualising nanowires in the pleura demonstrated at least 2 compartments – in leukocyte aggregations and in the mesothelium - which may have consequences for long term pathology in the pleural space including mesothelioma. PMID:22929371

Seeing and believing: recent advances in imaging cell-cell interactions

PubMed Central

Yap, Alpha S.; Michael, Magdalene; Parton, Robert G.

2015-01-01

Advances in cell and developmental biology have often been closely linked to advances in our ability to visualize structure and function at many length and time scales. In this review, we discuss how new imaging technologies and new reagents have provided novel insights into the biology of cadherin-based cell-cell junctions. We focus on three developments: the application of super-resolution optical technologies to characterize the nanoscale organization of cadherins at cell-cell contacts, new approaches to interrogate the mechanical forces that act upon junctions, and advances in electron microscopy which have the potential to transform our understanding of cell-cell junctions. PMID:26543555

Seeing and believing: recent advances in imaging cell-cell interactions.

PubMed

Yap, Alpha S; Michael, Magdalene; Parton, Robert G

2015-01-01

Advances in cell and developmental biology have often been closely linked to advances in our ability to visualize structure and function at many length and time scales. In this review, we discuss how new imaging technologies and new reagents have provided novel insights into the biology of cadherin-based cell-cell junctions. We focus on three developments: the application of super-resolution optical technologies to characterize the nanoscale organization of cadherins at cell-cell contacts, new approaches to interrogate the mechanical forces that act upon junctions, and advances in electron microscopy which have the potential to transform our understanding of cell-cell junctions.

High-temperature multipass cell for infrared spectroscopy of heated gases and vapors.

PubMed

Bartlome, R; Baer, M; Sigrist, M W

2007-01-01

In absorption spectroscopy, infrared spectra of heated gases or condensed samples in the vapor phase are usually recorded with a single pass heated gas cell. This device exhibits two orders of magnitude lower sensitivity than the high-temperature multipass cell presented in this article. Our device is a novel type of compact long path absorption cell that can withstand aggressive chemicals in addition to temperatures up to 723 K. The construction of the cell and its technical features are described in detail, paying special attention to the mechanisms that compensate for thermal expansion and that allow the user to vary the optical path length under any thermal or vacuum condition. The cell may be used with a laser source or implemented within a Fourier transform infrared spectrometer. Its design is compatible with optical arrangements using astigmatic mirrors or spherical mirrors in a Herriott configuration. Here we implement a homebuilt Herriott-type cell with a total optical path length of up to 35 m. In order to demonstrate the feasibility of the cell, methane and water vapor absorption lines showing dissimilar temperature effects on line intensity were recorded with the help of a mid-infrared laser source tunable between 3 and 4 microm. Emphasis is put on lines that are too weak to be recorded with a single pass cell.

Cell lineage patterns in the shoot meristem of the sunflower embryo in the dry seed

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

Jegla, D.E.; Sussex, I.M.

1989-01-01

We mapped the fate of cells in the shoot meristem of the dry-seed embryo of sunflower, Helianthus annuus L. cv. Peredovic, using irradiation-induced somatic sectors. We analyzed 249 chlorophyll-deficient or glabrous (hairless) sectors generated in 236 plants. Most sectors observed in the inflorescence extended into vegetative nodes. Thus cell lineages that ultimately gave rise to reproductive structures also contributed to vegetative structures. No single sector extended the entire length of the shoot. Thus the shoot is not derived from one or a few apical initials. Rather, the position, vertical extent, and width of the sectors at different levels of themore » shoot suggest that the shoot is derived from three to four circumferential populations of cells in each of three cell layers of the embryo meristem. Sectors had no common boundaries even in plants with two or three independent sectors, but varied in extent and overlapped along the length of the shoot. Thus individual cells in a single circumferential population behaved independently to contribute lineages of different vertical extents to the growing shoot. The predicted number of circumferential populations of cells as well as the apparent cell number in each population was consistent with the actual number of cells in the embryo meristem observed in histological sections.« less

Tunable Surface Repellency Maintains Stemness and Redox Capacity of Human Mesenchymal Stem Cells.

PubMed

Balikov, Daniel A; Crowder, Spencer W; Boire, Timothy C; Lee, Jung Bok; Gupta, Mukesh K; Fenix, Aidan M; Lewis, Holley N; Ambrose, Caitlyn M; Short, Philip A; Kim, Chang Soo; Burnette, Dylan T; Reilly, Matthew A; Murthy, N Sanjeeva; Kang, Mi-Lan; Kim, Won Shik; Sung, Hak-Joon

2017-07-12

Human bone marrow derived mesenchymal stem cells (hMSCs) hold great promise for regenerative medicine due to their multipotent differentiation capacity and immunomodulatory capabilities. Substantial research has elucidated mechanisms by which extracellular cues regulate hMSC fate decisions, but considerably less work has addressed how material properties can be leveraged to maintain undifferentiated stem cells. Here, we show that synthetic culture substrates designed to exhibit moderate cell-repellency promote high stemness and low oxidative stress-two indicators of naïve, healthy stem cells-in commercial and patient-derived hMSCs. Furthermore, the material-mediated effect on cell behavior can be tuned by altering the molar percentage (mol %) and/or chain length of poly(ethylene glycol) (PEG), the repellant block linked to hydrophobic poly(ε-caprolactone) (PCL) in the copolymer backbone. Nano- and angstrom-scale characterization of the cell-material interface reveals that PEG interrupts the adhesive PCL domains in a chain-length-dependent manner; this prevents hMSCs from forming mature focal adhesions and subsequently promotes cell-cell adhesions that require connexin-43. This study is the first to demonstrate that intrinsic properties of synthetic materials can be tuned to regulate the stemness and redox capacity of hMSCs and provides new insight for designing highly scalable, programmable culture platforms for clinical translation.

History of child maltreatment and telomere length in immune cell subsets: Associations with stress- and attachment-related hormones.

PubMed

Boeck, Christina; Krause, Sabrina; Karabatsiakis, Alexander; Schury, Katharina; Gündel, Harald; Waller, Christiane; Kolassa, Iris-Tatjana

2018-05-01

Experiencing maltreatment during childhood can have long-lasting consequences for both mental and physical health. Immune cell telomere length (TL) shortening might be one link between child maltreatment (CM) experiences and adverse health outcomes later in life. While the stress hormone cortisol has been associated with TL attrition, the attachment-related hormone oxytocin may promote resilience. In 15 mothers with and 15 age- and body mass index-matched mothers without CM, we assessed TL in peripheral blood mononuclear cells and selected immune cell subsets (monocytes, naive, and memory cytotoxic T cells) by quantitative fluorescence in situ hybridization, as well as peripheral cortisol and oxytocin levels. Memory cytotoxic T cells showed significantly shorter TL in association with CM, whereas TL in monocytes and naive cytotoxic T cells did not significantly differ between the two groups. Across both groups, cortisol was negatively associated with TL, while oxytocin was positively associated with TL in memory cytotoxic T cells. These results indicate that long-lived memory cytotoxic T cells are most affected by the increased biological stress state associated with CM. Keeping in mind the correlational and preliminary nature of the results, the data suggest that cortisol may have a damaging and oxytocin a protective function on TL.

Evaluation of medium-chain-length polyhydroxyalkanoate production by Pseudomonas putida LS46 using biodiesel by-product streams.

PubMed

Fu, Jilagamazhi; Sharma, Umesh; Sparling, Richard; Cicek, Nazim; Levin, David B

2014-07-01

Medium-chain-length polyhydroxyalkanoate (mcl-PHA) production by Pseudomonas putida LS46 was analyzed in shake-flask-based batch reactions, using pure chemical-grade glycerol (PG), biodiesel-derived "waste" glycerol (WG), and biodiesel-derived "waste" free fatty acids (WFA). Cell growth, substrate consumption, mcl-PHA accumulation within the cells, and the monomer composition of the synthesized biopolymers were monitored. The patterns of mcl-PHA synthesis in P. putida LS46 cells grown on PG and WG were similar but differed from that of cells grown with WFA. Polymer accumulation in glycerol-based cultures was stimulated by nitrogen limitation and plateaued after 48 h in both PG and WG cultures, with a total accumulation of 17.9% cell dry mass and 16.3% cell dry mass, respectively. In contrast, mcl-PHA synthesis was independent of nitrogen concentration in P. putida LS46 cells cultured with WFA, which accumulated to 29% cell dry mass. In all cases, the mcl-PHAs synthesized consisted primarily of 3-hydroxyoctanoate (C(8)) and 3-hydroxydecanoate (C(10)). WG and WFA supported similar or greater cell growth and mcl-PHA accumulation than PG under the experimental conditions used. These results suggest that biodiesel by-product streams could be used as low-cost carbon sources for sustainable mcl-PHA production.

Efficient, Glucose Responsive, and Islet-Specific Transgene Expression by a Modified Rat Insulin Promoter

PubMed Central

Chai, Renjie; Chen, Shuyuan; Ding, Jiahuan; Grayburn, Paul A

2009-01-01

This study was done to improve efficiency and islet specificity of the rat insulin promoter (RIP). Various rat insulin promoter lengths were prepared and tested in vitro to drive luciferase reporter gene expression in INS1-cells, alpha-cells, acinar cells, ductal cells, and fibroblasts. The CMV promoter was used as a positive control. In addition, the DsRed reporter gene was administered in vivo to rat pancreas by ultrasound-targeted microbubble destruction (UTMD). Confocal microscopy was used to detect the presence and distribution of DsRed within the pancreas after UTMD. A modified RIP3.1 promoter, which includes portions of the insulin gene after its transcription start site is 5-fold more active in INS-1 cells than the full length RIP promoter or the CMV promoter. RIP3.1 is regulated by glucose level and various islet transcription factors in vitro, and exhibits activity in alpha-cells, but not exocrine cells. In vivo delivery of RIP3.1-DsRed resulted in expression of DsRed protein in beta-cells, and to a lesser extent alpha cells under normal glucose conditions. No DsRed signal was present in exocrine pancreas under RIP3.1. A modified rat insulin promoter, RIP3.1, efficiently and specifically directs gene expression to endocrine pancreas. PMID:19727136

Novel anti-c-Mpl monoclonal antibodies identified multiple differentially glycosylated human c-Mpl proteins in megakaryocytic cells but not in human solid tumors.

PubMed

Zhan, Jinghui; Felder, Barbara; Ellison, Aaron R; Winters, Aaron; Salimi-Moosavi, Hossein; Scully, Sheila; Turk, James R; Wei, Ping

2013-06-01

Thrombopoietin and its cognate receptor, c-Mpl, are the primary molecular regulators of megakaryocytopoiesis and platelet production. To date the pattern of c-Mpl expression in human solid tumors and the distribution and biochemical properties of c-Mpl proteins in hematopoietic tissues are largely unknown. We have recently developed highly specific mouse monoclonal antibodies (MAb) against human c-Mpl. In this study we used these antibodies to demonstrate the presence of full-length and truncated human c-Mpl proteins in various megakaryocytic cell types, and their absence in over 100 solid tumor cell lines and in the 12 most common primary human tumor types. Quantitative assays showed a cell context-dependent distribution of full-length and truncated c-Mpl proteins. All forms of human c-Mpl protein were found to be modified with extensive N-linked glycosylation but different degrees of sialylation and O-linked glycosylation. Of note, different variants of full-length c-Mpl protein exhibiting differential glycosylation were expressed in erythromegakaryocytic leukemic cell lines and in platelets from healthy human donors. This work provides a comprehensive analysis of human c-Mpl mRNA and protein expression on normal and malignant hematopoietic and non-hematopoietic cells and demonstrates the multiple applications of several novel anti-c-Mpl antibodies.

Telomerase and mammalian ageing: a critical appraisal.

PubMed

Goyns, M H; Lavery, W L

2000-03-13

The telomeres that occur at the end of chromosomes are maintained by the activity of telomerase and are thought to be important protective factors in maintaining the integrity of chromosomes. It now appears that in vitro replicative senescence, which has been observed in cultured somatic cells, is due to a loss of telomere length in those cells, caused by inactivity of telomerase. This has led to the proposition that telomerase activity is an important determinant in organismal ageing. However, many cells in the body do not proliferate regularly and therefore will not lose telomere length. Cells that do proliferate frequently have now been shown to have active telomerase. Other cells, such as fibroblasts, that do not have telomerase activity but proliferate only occasionally may not reach the Hayflick limit during the lifetime of an animal. There is also no correlation between telomere length and the maximal lifespan exhibited by different species. Studies of telomerase knock-out mice have reported some aspects of accelerated ageing after three generations, but the relevance of these observations to normal ageing remains unconvincing. The role of telomerase in producing immortal tumour cells and the possibility that activation of telomerase is an important event in malignant transformation is similarly controversial and open to alternative interpretations. The significance of these and other observations, and how they define the role of telomerase in ageing, is discussed.

The clinical implication and molecular mechanism of preferential IL-4 production by modified glycolipid-stimulated NKT cells

PubMed Central

Oki, Shinji; Chiba, Asako; Yamamura, Takashi; Miyake, Sachiko

2004-01-01

OCH, a sphingosine-truncated analog of α-galactosylceramide (αGC), is a potential therapeutic reagent for a variety of Th1-mediated autoimmune diseases through its selective induction of Th2 cytokines from natural killer T (NKT) cells. We demonstrate here that the NKT cell production of IFN-γ is more susceptible to the sphingosine length of glycolipid ligand than that of IL-4 and that the length of the sphingosine chain determines the duration of NKT cell stimulation by CD1d-associated glycolipids. Furthermore, IFN-γ production by NKT cells requires longer T cell receptor stimulation than is required for IL-4 production by NKT cells stimulated either with immobilized mAb to CD3 or with immobilized “αGC-loaded” CD1d molecules. Interestingly, transcription of IFN-γ but not that of IL-4 was sensitive to cycloheximide treatment, indicating the intrinsic involvement of de novo protein synthesis for IFN-γ production by NKT cells. Finally, we determined c-Rel was preferentially transcribed in αGC-stimulated but not in OCH-stimulated NKT cells and was essential for IFN-γ production by activated NKT cells. Given the dominant immune regulation by the remarkable cytokine production of ligand-stimulated NKT cells in vivo, in comparison with that of (antigen-specific) T cells or NK cells, the current study confirms OCH as a likely therapeutic reagent for use against Th1-mediated autoimmune diseases and provides a novel clue for the design of drugs targeting NKT cells. PMID:15173890

Investigations on Two Co-C Fixed-Point Cells Prepared at INRIM and LNE-Cnam

NASA Astrophysics Data System (ADS)

Battuello, M.; Florio, M.; Sadli, M.; Bourson, F.

2011-08-01

INRIM and LNE-Cnam agreed to undertake a collaboration aimed to verify, through the use of metal-carbon eutectic fixed-point cells, methods and facilities used for defining the transition temperature of eutectic fixed points and manufacturing procedures of cells. To this purpose and as a first step of the cooperation, a Co-C cell manufactured at LNE-Cnam was measured at INRIM and compared with a local cell. The two cells were of different designs: the INRIM cell of 10 cm3 inner volume and the LNE-Cnam one of 3.9 cm3. The external dimensions of the two cells were noticeably different, namely, 40 mm in length and 24 mm in diameter for the LNE-Cnam cell 3Co4 and 110 mm in length and 42 mm in diameter for the INRIM cell. Consequently, the investigation of the effect of temperature distributions in the heating furnace was undertaken by implementing the cells inside single-zone and three-zone furnaces. The transition temperature of the cell was determined at the two institutes making use of different techniques: at INRIM radiation scales at 900 nm, 950 nm, and 1.6 μm were realized from In to Cu and then used to define T 90(Co-C) by extrapolation. At LNE-Cnam, a radiance comparator based on a grating monochromator was used for the extrapolation from the Cu fixed point. This paper presents a comparative description of the cells and the manufacturing methods and the results in terms of equivalence between the two cells and melting temperatures determined at INRIM and LNE-Cnam.

Cellular basis of neuroepithelial bending during mouse spinal neural tube closure

PubMed Central

McShane, Suzanne G.; Molè, Matteo A.; Savery, Dawn; Greene, Nicholas D. E; Tam, Patrick P.L.; Copp, Andrew J.

2015-01-01

Summary Bending of the neural plate at paired dorsolateral hinge points (DLHPs) is required for neural tube closure in the spinal region of the mouse embryo. As a step towards understanding the morphogenetic mechanism of DLHP development, we examined variations in neural plate cellular architecture and proliferation during closure. Neuroepithelial cells within the median hinge point (MHP) contain nuclei that are mainly basally located and undergo relatively slow proliferation, with a 7 h cell cycle length. In contrast, cells in the dorsolateral neuroepithelium, including the DLHP, exhibit nuclei distributed throughout the apico-basal axis and undergo rapid proliferation, with a 4 h cell cycle length. As the neural folds elevate, cell numbers increase to a greater extent in the dorsolateral neural plate that contacts the surface ectoderm, compared with the more ventromedial neural plate where cells contact paraxial mesoderm and notochord. This marked increase in dorsolateral cell number cannot be accounted for solely on the basis of enhanced cell proliferation in this region. We hypothesised that neuroepithelial cells may translocate in a ventral-to-dorsal direction as DLHP formation occurs, and this was confirmed by vital cell labelling in cultured embryos. The translocation of cells into the neural fold, together with its more rapid cell proliferation, leads to an increase in cell density dorsolaterally compared with the more ventromedial neural plate. These findings suggest a model in which DLHP formation may proceed through ‘buckling’ of the neuroepithelium at a dorso-ventral boundary marked by a change in cell-packing density. PMID:26079577

“Inclonals”

PubMed Central

Hakim, Rahely

2009-01-01

Full-length antibodies and antibodies that ferry a cargo to target cells are desired biopharmaceuticals. We describe the production of full-length IgGs and IgG-toxin fusion proteins in E. coli. In the presented examples of anti CD30 and anti EGF-receptor antibodies, the antibody heavy and light chains or toxin fusions thereof were expressed in separate bacterial cultures, where they accumulated as insoluble inclusion bodies. Following refolding and purification, high yields (up to 50 mg/L of shake flask culture) of highly purified (>90%) full-length antibodies and antibody-toxin fusions were obtained. The bacterially produced antibodies, named “Inclonals,” equaled the performance of the same IgGs that were produced using conventional mammalian cell culture in binding properties as well as in cell killing potency. The rapid and cost effective IgG production process and the high quality of the resultant product may make the bacterial production of full-length IgG and IgG-drug fusion proteins an attractive option for antibody production and a significant contribution to recombinant antibody technology. PMID:20065645

Emergent multicellular life cycles in filamentous bacteria owing to density-dependent population dynamics.

PubMed

Rossetti, Valentina; Filippini, Manuela; Svercel, Miroslav; Barbour, A D; Bagheri, Homayoun C

2011-12-07

Filamentous bacteria are the oldest and simplest known multicellular life forms. By using computer simulations and experiments that address cell division in a filamentous context, we investigate some of the ecological factors that can lead to the emergence of a multicellular life cycle in filamentous life forms. The model predicts that if cell division and death rates are dependent on the density of cells in a population, a predictable cycle between short and long filament lengths is produced. During exponential growth, there will be a predominance of multicellular filaments, while at carrying capacity, the population converges to a predominance of short filaments and single cells. Model predictions are experimentally tested and confirmed in cultures of heterotrophic and phototrophic bacterial species. Furthermore, by developing a formulation of generation time in bacterial populations, it is shown that changes in generation time can alter length distributions. The theory predicts that given the same population growth curve and fitness, species with longer generation times have longer filaments during comparable population growth phases. Characterization of the environmental dependence of morphological properties such as length, and the number of cells per filament, helps in understanding the pre-existing conditions for the evolution of developmental cycles in simple multicellular organisms. Moreover, the theoretical prediction that strains with the same fitness can exhibit different lengths at comparable growth phases has important implications. It demonstrates that differences in fitness attributed to morphology are not the sole explanation for the evolution of life cycles dominated by multicellularity.

Induced sensitivity of Bacillus subtilis colony morphology to mechanical media compression

PubMed Central

Polka, Jessica K.

2014-01-01

Bacteria from several taxa, including Kurthia zopfii, Myxococcus xanthus, and Bacillus mycoides, have been reported to align growth of their colonies to small features on the surface of solid media, including anisotropies created by compression. While the function of this phenomenon is unclear, it may help organisms navigate on solid phases, such as soil. The origin of this behavior is also unknown: it may be biological (that is, dependent on components that sense the environment and regulate growth accordingly) or merely physical. Here we show that B. subtilis, an organism that typically does not respond to media compression, can be induced to do so with two simple and synergistic perturbations: a mutation that maintains cells in the swarming (chained) state, and the addition of EDTA to the growth media, which further increases chain length. EDTA apparently increases chain length by inducing defects in cell separation, as the treatment has only marginal effects on the length of individual cells. These results lead us to three conclusions. First, the wealth of genetic tools available to B. subtilis will provide a new, tractable chassis for engineering compression sensitive organisms. Second, the sensitivity of colony morphology to media compression in Bacillus can be modulated by altering a simple physical property of rod-shaped cells. And third, colony morphology under compression holds promise as a rapid, simple, and low-cost way to screen for changes in the length of rod-shaped cells or chains thereof. PMID:25289183

Silibinin negatively contributes to primary cilia length via autophagy regulated by histone deacetylase 6 in confluent mouse embryo fibroblast 3T3-L1 cells.

PubMed

Xu, Qian; Liu, Wei; Liu, Xiaoling; Liu, Weiwei; Wang, Hongju; Yao, Guodong; Zang, Linghe; Hayashi, Toshihiko; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi

2016-09-01

Primary cilium is a cellular antenna, signalling as a sensory organelle. Numerous pathological manifestation is associated with change of its length. Although the interaction between autophagy and primary cilia has been suggested, the role of autophagy in primary cilia length is largely unknown. In this study the primary cilia were immunostained and observed by using confocal fluorescence microscopy, and we found that silibinin, a natural flavonoid, shortened the length of primary cilia, meanwhile it also induced autophagy in 3T3-L1 cells. This study was designed to investigate the significance of silibinin-induced autophagy in primary ciliary structure in confluent mouse embryo fibroblast 3T3-L1 cells. Either blocking the autophagic flux with pre-treatment with the autophagy inhibitor, 3-methyladenine (3-MA), or transfection of siRNA targeting LC3 inhibited the reduction of cilia length caused by silibinin exposure. Autophagy induced by silibinin decreased expressions of the cilia-associated proteins, such as IFT88, KIF3a and Ac-tubulin, while 3-MA restored it, indicating that autophagy induced by silibinin led to a reduction of primary cilia length. Histone deacetylase 6 (HDAC6), which was suggested as a mediator of autophagy, was up-regulated by silibinin in a time-dependent manner. In addition, 3T3-L1 cells treated with siRNA against HDAC6 had a reduced autophagic level and were protected from silibinin-induced cilia shortening. Taken together, we conclude that the HDAC6-mediated autophagy negatively regulates primary cilia length during silibinin treatment and has the potential to serve as a therapeutic target for primary cilia-associated ciliopathies. These findings thus provide new information about the potential link between autophagy and primary cilia.

Circadian Clock Synchronization of the Cell Cycle in Zebrafish Occurs through a Gating Mechanism Rather Than a Period-phase Locking Process.

PubMed

Laranjeiro, Ricardo; Tamai, T Katherine; Letton, William; Hamilton, Noémie; Whitmore, David

2018-04-01

Studies from a number of model systems have shown that the circadian clock controls expression of key cell cycle checkpoints, thus providing permissive or inhibitory windows in which specific cell cycle events can occur. However, a major question remains: Is the clock actually regulating the cell cycle through such a gating mechanism or, alternatively, is there a coupling process that controls the speed of cell cycle progression? Using our light-responsive zebrafish cell lines, we address this issue directly by synchronizing the cell cycle in culture simply by changing the entraining light-dark (LD) cycle in the incubator without the need for pharmacological intervention. Our results show that the cell cycle rapidly reentrains to a shifted LD cycle within 36 h, with changes in p21 expression and subsequent S phase timing occurring within the first few hours of resetting. Reentrainment of mitosis appears to lag S phase resetting by 1 circadian cycle. The range of entrainment of the zebrafish clock to differing LD cycles is large, from 16 to 32 hour periods. We exploited this feature to explore cell cycle entrainment at both the population and single cell levels. At the population level, cell cycle length is shortened or lengthened under corresponding T-cycles, suggesting that a 1:1 coupling mechanism is capable of either speeding up or slowing down the cell cycle. However, analysis at the single cell level reveals that this, in fact, is not true and that a gating mechanism is the fundamental method of timed cell cycle regulation in zebrafish. Cell cycle length at the single cell level is virtually unaltered with varying T-cycles.

Circadian Clock Synchronization of the Cell Cycle in Zebrafish Occurs through a Gating Mechanism Rather Than a Period-phase Locking Process

PubMed Central

Tamai, T. Katherine; Letton, William; Hamilton, Noémie; Whitmore, David

2018-01-01

Studies from a number of model systems have shown that the circadian clock controls expression of key cell cycle checkpoints, thus providing permissive or inhibitory windows in which specific cell cycle events can occur. However, a major question remains: Is the clock actually regulating the cell cycle through such a gating mechanism or, alternatively, is there a coupling process that controls the speed of cell cycle progression? Using our light-responsive zebrafish cell lines, we address this issue directly by synchronizing the cell cycle in culture simply by changing the entraining light-dark (LD) cycle in the incubator without the need for pharmacological intervention. Our results show that the cell cycle rapidly reentrains to a shifted LD cycle within 36 h, with changes in p21 expression and subsequent S phase timing occurring within the first few hours of resetting. Reentrainment of mitosis appears to lag S phase resetting by 1 circadian cycle. The range of entrainment of the zebrafish clock to differing LD cycles is large, from 16 to 32 hour periods. We exploited this feature to explore cell cycle entrainment at both the population and single cell levels. At the population level, cell cycle length is shortened or lengthened under corresponding T-cycles, suggesting that a 1:1 coupling mechanism is capable of either speeding up or slowing down the cell cycle. However, analysis at the single cell level reveals that this, in fact, is not true and that a gating mechanism is the fundamental method of timed cell cycle regulation in zebrafish. Cell cycle length at the single cell level is virtually unaltered with varying T-cycles. PMID:29444612

Effects of inorganic lead on the differentiation and growth of cultured hippocampal and neuroblastoma cells.

PubMed

Audesirk, T; Audesirk, G; Ferguson, C; Shugarts, D

1991-01-01

Lead exposure has devastating effects on the developing nervous system, and has been implicated in variety of behavioral and cognitive deficits as well as neural morphological abnormalities. Since lead impacts many calcium-dependent processes, one likely mechanism of lead toxicity is its disruption of calcium dependent processes, among which is neuronal differentiation. We investigated the effects of inorganic lead on survival and several parameters of differentiation of cultured neurons. Three different cell types were used: Rat hippocampal neurons (a primary CNS cell type), B50 rat neuroblastoma cells (a transformed CNS-derived cell line), and N1E-115 mouse neuroblastoma cells (a transformed peripherally-derived cell line). Lead concentrations ranged from low nM to 1 mM. Lead effects differed considerably among the three cell types, with B50 cells least affected. Lead effects were generally multimodal, with fewest effects observed at intermediate concentrations. Lead inhibited neurite initiation in hippocampal neurons, but stimulated initiation in N1E-115 cells. In those cells that differentiated, lead increased dendrite numbers in hippocampal neurons and neurite numbers in N1E-115 cells. Lead exposure increased both the length and the degree of branching of axons in hippocampal neurons and the length of neurites in N1E-115 cells. We hypothesize that lead impacts multiple regulatory processes that influence neuron survival and differentiation, and that its effects show differing dose-dependencies. The differing responses of the different cell types to lead suggests that differentiation may be regulated in different ways by the three types of cells. Alternatively, or additionally, the cell types may differ in their ability to compensate for, sequester, or expel lead.

RESISTANCE TO X-IRRADIATION BY EMBRYONIC CELLS OF THE LIMB-BUDS OF TADPOLES

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

Allen, B.M.; Ewell, L.M.

1959-01-01

Both total-body irradiation and shielding of the trunk were used to study the effects of x irradiation from 1000 to 30000 r upon the limb-buds of Bufo boreas and Hyla regilla tadpoles. The object was to test the view that the younger the cells the more sensitive they are to irradiation. The answer is negative. If there is any special susceptibility of these undifferentiated cells it should appear at levels far below the 30000 r maximum employed. A sharp distinction is made between the very susceptible mitotic cells and the resistant non-dividing embryonic cells that have been accumulated in suchmore » numbers that they may rapidly differentiate into the characteristic limb tissues under the stimulus of the thyroid hormone. Many irradiated ectoderm cells were changed to form bizarre excrescences but were not destroyed. Unicellular cuthneous gland cells continued to arise even after the heaviest irradiation. Irradiated tadpoles with hind limb-buds from 0.6 mm down to 0.2 mm length were unable to develop normal limbs. This capacity was propontional to the number of non-dividing embryonic cells stored at the time of irradiation. Irradiation of 5000, equal degree but the rapidity was greatest in the cases of higher dosage. Not only did these levels of irradiation fail to destroy the non-dividing embryonic cells but they did not effect their pre-deterrmined specificity nor modify their capacity for subsequent differentiation and growth. Exposure to a thyroxin solution caused the hind limb-buds without visible differentiation of cells to grow from a length of 0.8 or 0.9 mm or 1.0 mm at the time of irradiation to a length of as much as 5.0 mm in the course of 7 days. Development of thigh, shank, ankle, and toes was complete. Microscopic studies showed characteristic tissues such as cartilage, connective tissue, and muscle, developed to a comparable degree in control and irradiated specimens. (auth)« less

Regulation of Cilium Length and Intraflagellar Transport by the RCK-Kinases ICK and MOK in Renal Epithelial Cells

PubMed Central

Broekhuis, Joost R.; Verhey, Kristen J.; Jansen, Gert

2014-01-01

Primary cilia are important sensory organelles. They exist in a wide variety of lengths, which could reflect different cell-specific functions. How cilium length is regulated is unclear, but it probably involves intraflagellar transport (IFT), which transports protein complexes along the ciliary axoneme. Studies in various organisms have identified the small, conserved family of ros-cross hybridizing kinases (RCK) as regulators of cilium length. Here we show that Intestinal Cell Kinase (ICK) and MAPK/MAK/MRK overlapping kinase (MOK), two members of this family, localize to cilia of mouse renal epithelial (IMCD-3) cells and negatively regulate cilium length. To analyze the effects of ICK and MOK on the IFT machinery, we set up live imaging of five fluorescently tagged IFT proteins: KIF3B, a subunit of kinesin-II, the main anterograde IFT motor, complex A protein IFT43, complex B protein IFT20, BBSome protein BBS8 and homodimeric kinesin KIF17, whose function in mammalian cilia is unclear. Interestingly, all five proteins moved at ∼0.45 µm/s in anterograde and retrograde direction, suggesting they are all transported by the same machinery. Moreover, GFP tagged ICK and MOK moved at similar velocities as the IFT proteins, suggesting they are part of, or transported by the IFT machinery. Indeed, loss- or gain-of-function of ICK affected IFT speeds: knockdown increased anterograde velocities, whereas overexpression reduced retrograde speed. In contrast, MOK knockdown or overexpression did not affect IFT speeds. Finally, we found that the effects of ICK or MOK knockdown on cilium length and IFT are suppressed by rapamycin treatment, suggesting that these effects require the mTORC1 pathway. Our results confirm the importance of RCK kinases as regulators of cilium length and IFT. However, whereas some of our results suggest a direct correlation between cilium length and IFT speed, other results indicate that cilium length can be modulated independent of IFT speed. PMID:25243405

21 CFR 178.3620 - Mineral oil.

Code of Federal Regulations, 2014 CFR

2014-04-01

... cell in the range from 260-350 mµ, inclusive, compared to the solvent control. If the absorbance so... closely fit the stem of the chromatographic tube. Suction flask. 250-milliliter or 500-milliliter filter... evaporated. Spectrophotometric cells. Fused quartz cells, optical path length in the range of 5,000...

Optimization of differentiation time of mesenchymal-stem-cell to tenocyte under a cyclic stretching with a microgrooved culture membrane and selected measurement cells.

PubMed

Morita, Yasuyuki; Yamashita, Takahiro; Toku, Toku; Ju, Yang

2018-01-01

There is a need for efficient stem cell-to-tenocyte differentiation techniques for tendon tissue engineering. More than 1 week is required for tenogenic differentiation with chemical stimuli, including co-culturing. Research has begun to examine the utility of mechanical stimuli, which reduces the differentiation time to several days. However, the precise length of time required to differentiate human bone marrow-derived mesenchymal stem cells (hBMSCs) into tenocytes has not been clarified. Understanding the precise time required is important for future tissue engineering projects. Therefore, in this study, a method was developed to more precisely determine the length of time required to differentiate hBMSCs into tenocytes with cyclic stretching stimulus. First, it had to be determined how stretching stimulation affected the cells. Microgrooved culture membranes were used to suppress cell orientation behavior. Then, only cells oriented parallel to the microgrooves were selected and evaluated for protein synthesis levels for differentiation. The results revealed that growing cells on the microgrooved membrane and selecting optimally-oriented cells for measurement improved the accuracy of the differentiation evaluation, and that hBMSCs differentiated into tenocytes in approximately 10 h. The differentiation time corresponded to the time required for cellular cytoskeleton reorganization and cellular morphology alterations. This suggests that cells, when subjected to mechanical stimulus, secrete mRNAs and proteins for both cytoskeleton reorganization and differentiation.

Association between length of storage of red blood cell units and outcome of critically ill children: a prospective observational study

PubMed Central

2010-01-01

Introduction Transfusion is a common treatment in pediatric intensive care units (PICUs). Studies in adults suggest that prolonged storage of red blood cell units is associated with worse clinical outcome. No prospective study has been conducted in children. Our objectives were to assess the clinical impact of the length of storage of red blood cell units on clinical outcome of critically ill children. Methods Prospective, observational study conducted in 30 North American centers, in consecutive patients aged <18 years with a stay ≥ 48 hours in a PICU. The primary outcome measure was the incidence of multiple organ dysfunction syndrome after transfusion. The secondary outcomes were 28-day mortality and PICU length of stay. Odds ratios were adjusted for gender, age, number of organ dysfunctions at admission, total number of transfusions, and total dose of transfusion, using a multiple logistic regression model. Results The median length of storage was 14 days in 296 patients with documented length of storage. For patients receiving blood stored ≥ 14 days, the adjusted odds ratio for an increased incidence of multiple organ dysfunction syndrome was 1.87 (95% CI 1.04;3.27, P = 0.03). There was also a significant difference in the total PICU length of stay (adjusted median difference +3.7 days, P < 0.001) and no significant change in mortality. Conclusions In critically ill children, transfusion of red blood cell units stored for ≥ 14 days is independently associated with an increased occurrence of multiple organ dysfunction syndrome and prolonged PICU stay. PMID:20377853

Gold Nanoparticles as a Photothermal Agent in Cancer Therapy: The Thermal Ablation Characteristic Length.

PubMed

Grosges, Thomas; Barchiesi, Dominique

2018-05-31

In cancer therapy, the thermal ablation of diseased cells by embedded nanoparticles is one of the known therapies. It is based on the absorption of the energy of the illuminating laser by nanoparticles. The resulting heating of nanoparticles kills the cell where these photothermal agents are embedded. One of the main constraints of this therapy is preserving the surrounding healthy cells. Therefore, two parameters are of interest. The first one is the thermal ablation characteristic length, which corresponds to an action distance around the nanoparticles for which the temperature exceeds the ablation threshold. This critical geometric parameter is related to the expected conservation of the body temperature in the surroundings of the diseased cell. The second parameter is the temperature that should be reached to achieve active thermal agents. The temperature depends on the power of the illuminating laser, on the size of nanoparticles and on their physical properties. The purpose of this paper is to propose behavior laws under the constraints of both the body temperature at the boundary of the cell to preserve surrounding cells and an acceptable range of temperature in the target cell. The behavior laws are deduced from the finite element method, which is able to model aggregates of nanoparticles. We deduce sensitivities to the laser power and to the particle size. We show that the tuning of the temperature elevation and of the distance of action of a single nanoparticle is not significantly affected by variations of the particle size and of the laser power. Aggregates of nanoparticles are much more efficient, but represent a potential risk to the surrounding cells. Fortunately, by tuning the laser power, the thermal ablation characteristic length can be controlled.

Telomere elongation protects heart and lung tissue cells from fatal damage in rats exposed to severe hypoxia.

PubMed

Wang, Yaping; Zhao, Zhen; Zhu, Zhiyong; Li, Pingying; Li, Xiaolin; Xue, Xiaohong; Duo, Jie; Ma, Yingcai

2018-02-17

The effects of acute hypoxia at high altitude on the telomere length of the cells in the heart and lung tissues remain unclear. This study aimed to investigate the change in telomere length of rat heart and lung tissue cells in response to acute exposure to severe hypoxia and its role in hypoxia-induced damage to heart and lung tissues. Forty male Wistar rats (6-week old) were randomized into control group (n = 10) and hypoxia group (n = 30). Rats in control group were kept at an altitude of 1500 m, while rats in hypoxia group were exposed to simulated hypoxia with an altitude of 5000 m in a low-pressure oxygen chamber for 1, 3, and 7 days (n = 10). The left ventricular and right middle lobe tissues of each rat were collected for measurement of telomere length and reactive oxygen species (ROS) content, and the mRNA and protein levels of telomerase reverse transcriptase (TERT), hypoxia-inducible factor1α (HIF-1α), and hypoxia-inducible factor1α (HIF-2α). Increased exposure to hypoxia damaged rat heart and lung tissue cells and increased ROS production and telomere length. The mRNA and protein levels of TERT and HIF-1α were significantly higher in rats exposed to hypoxia and increased with prolonged exposure; mRNA and protein levels of HIF-2α increased only in rats exposed to hypoxia for 7 days. TERT was positively correlated with telomere length and the levels of HIF-1α but not HIF-2α. Acute exposure to severe hypoxia causes damage to heart and lung tissues due to the production of ROS but promotes telomere length and adaptive response by upregulating TERT and HIF-1α, which protect heart and lung tissue cells from fatal damage.

Diffusing-wave spectroscopy in a standard dynamic light scattering setup

NASA Astrophysics Data System (ADS)

Fahimi, Zahra; Aangenendt, Frank J.; Voudouris, Panayiotis; Mattsson, Johan; Wyss, Hans M.

2017-12-01

Diffusing-wave spectroscopy (DWS) extends dynamic light scattering measurements to samples with strong multiple scattering. DWS treats the transport of photons through turbid samples as a diffusion process, thereby making it possible to extract the dynamics of scatterers from measured correlation functions. The analysis of DWS data requires knowledge of the path length distribution of photons traveling through the sample. While for flat sample cells this path length distribution can be readily calculated and expressed in analytical form; no such expression is available for cylindrical sample cells. DWS measurements have therefore typically relied on dedicated setups that use flat sample cells. Here we show how DWS measurements, in particular DWS-based microrheology measurements, can be performed in standard dynamic light scattering setups that use cylindrical sample cells. To do so we perform simple random-walk simulations that yield numerical predictions of the path length distribution as a function of both the transport mean free path and the detection angle. This information is used in experiments to extract the mean-square displacement of tracer particles in the material, as well as the corresponding frequency-dependent viscoelastic response. An important advantage of our approach is that by performing measurements at different detection angles, the average path length through the sample can be varied. For measurements performed on a single sample cell, this gives access to a wider range of length and time scales than obtained in a conventional DWS setup. Such angle-dependent measurements also offer an important consistency check, as for all detection angles the DWS analysis should yield the same tracer dynamics, even though the respective path length distributions are very different. We validate our approach by performing measurements both on aqueous suspensions of tracer particles and on solidlike gelatin samples, for which we find our DWS-based microrheology data to be in good agreement with rheological measurements performed on the same samples.

Far off-resonance laser frequency stabilization using multipass cells in Faraday rotation spectroscopy.

PubMed

Quan, Wei; Li, Yang; Li, Rujie; Shang, Huining; Fang, Zishan; Qin, Jie; Wan, Shuangai

2016-04-01

We propose a far off-resonance laser frequency stabilization method by using multipass cells in Rb Faraday rotation spectroscopy. Based on the detuning equation, if multipass cells with several meters optical path length are used in the conventional Faraday spectroscopy, the detuning of the lock point can be extended much further from the alkali metal resonance. A plate beam splitter was used to generate two different Faraday signals at the same time. The transmitted optical path length was L=50  mm and the reflected optical path length was 2L=100  mm. When the optical path length doubled, the detuning of the lock points moved further away from the atomic resonance. The temperature dependence of the detuning of the lock point was also analyzed. A temperature-insensitive lock point was found near resonance when the cell temperature was between 110°C and 130°C. We achieved an rms fluctuation of 0.9 MHz/23 h at a detuning of 0.5 GHz. A frequency drift of 16 MHz/h at a detuning of -5.6  GHz and 4 MHz/h at a detuning of -5.2  GHz were also obtained for the transmitted and reflected light Faraday signal.

Transverse mode analysis of optofluidic intracavity spectroscopy of canine hemangiosarcoma

NASA Astrophysics Data System (ADS)

Wang, Weina; Thamm, Douglas H.; Kisker, David W.; Lear, Kevin L.

2010-02-01

The label-free technique of optofluidic intracavity spectroscopy (OFIS) uses the optical transmission spectrum of a cell in a microfluidic optical resonator to distinguish cancerous and non-cancerous cells. Based on their distinctive characteristic transmission spectra, canine hemangiosarcoma (HSA) cancer cells and normal peripheral blood mononuclear cells (PBMCs) have been differentiated using the OFIS technique with high statistical significance (p<10- 6). 95% sensitivity and 98% specificity were achieved simultaneously. A cell lens model explains trends in the transverse mode pattern in the transmission spectra of HSA cells and allows extraction of cell focal length.

Single-cell transcriptome conservation in cryopreserved cells and tissues.

PubMed

Guillaumet-Adkins, Amy; Rodríguez-Esteban, Gustavo; Mereu, Elisabetta; Mendez-Lago, Maria; Jaitin, Diego A; Villanueva, Alberto; Vidal, August; Martinez-Marti, Alex; Felip, Enriqueta; Vivancos, Ana; Keren-Shaul, Hadas; Heath, Simon; Gut, Marta; Amit, Ido; Gut, Ivo; Heyn, Holger

2017-03-01

A variety of single-cell RNA preparation procedures have been described. So far, protocols require fresh material, which hinders complex study designs. We describe a sample preservation method that maintains transcripts in viable single cells, allowing one to disconnect time and place of sampling from subsequent processing steps. We sequence single-cell transcriptomes from >1000 fresh and cryopreserved cells using 3'-end and full-length RNA preparation methods. Our results confirm that the conservation process did not alter transcriptional profiles. This substantially broadens the scope of applications in single-cell transcriptomics and could lead to a paradigm shift in future study designs.

Tunneling Nanotubes are Novel Cellular Structures That Communicate Signals Between Trabecular Meshwork Cells.

PubMed

Keller, Kate E; Bradley, John M; Sun, Ying Ying; Yang, Yong-Feng; Acott, Ted S

2017-10-01

The actin cytoskeleton of trabecular meshwork (TM) cells plays a role in regulating aqueous humor outflow. Many studies have investigated stress fibers, but F-actin also assembles into other supramolecular structures including filopodia. Recently, specialized filopodia called tunneling nanotubes (TNTs) have been described, which communicate molecular signals and organelles directly between cells. Here, we investigate TNT formation by TM cells. Human TM cells were labeled separately with the fluorescent dyes, DiO and DiD, or with mitochondrial dye. Fixed or live TM cells were imaged using confocal microscopy. Image analysis software was used to track fluorescent vesicles and count the number and length of filopodia. The number of fluorescently labeled vesicles transferred between cells was counted in response to specific inhibitors of the actin cytoskeleton. Human TM tissue was stained with phalloidin. Live-cell confocal imaging of cultured TM cells showed transfer of fluorescently labeled vesicles and mitochondria via TNTs. In TM tissue, a long (160 μm) actin-rich cell process bridged an intertrabecular space and did not adhere to the substratum. Treatment of TM cells with CK-666, an Arp2/3 inhibitor, significantly decreased the number and length of filopodia, decreased transfer of fluorescently labeled vesicles and induced thick stress fibers compared to vehicle control. Conversely, inhibiting stress fibers using Y27632 increased transfer of vesicles and induced long cell processes. Identification of TNTs provides a means by which TM cells can directly communicate with each other over long distances. This may be particularly important to overcome limitations of diffusion-based signaling in the aqueous humor fluid environment.

Irradiation With Carbon Ion Beams Induces Apoptosis, Autophagy, and Cellular Senescence in a Human Glioma-Derived Cell Line

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

Jinno-Oue, Atsushi; Shimizu, Nobuaki; 21st Century Center of Excellence Program for Biomedical Research Using Accelerator Technology, Maebashi, Gunma

2010-01-15

Purpose: We examined biological responses of human glioma cells to irradiation with carbon ion beams (C-ions). Methods and Materials: A human glioma-derived cell line, NP-2, was irradiated with C-ions. Apoptotic cell nuclei were stained with Hoechst 33342. Induction of autophagy was examined either by staining cells with monodansylcadaverine (MDC) or by Western blotting to detect conversion of microtuble-associated protein light chain 3 (MAP-LC3) (LC3-I) to the membrane-bound form (LC3-II). Cellular senescence markers including induction of senescence-associated beta-galactosidase (SA-beta-gal) were examined. The mean telomere length of irradiated cells was determined by Southern blot hybridization. Expression of tumor suppressor p53 and cyclin/cyclin-dependentmore » kinase inhibitor p21{sup WAF1/CIP1} in the irradiated cells was analyzed by Western blotting. Results: When NP-2 cells were irradiated with C-ions at 6 Gy, the major population of the cells died of apoptosis and autophagy. The residual fraction of attached cells (<1% of initially irradiated cells) could not form a colony: however, they showed a morphological phenotype consistent with cellular senescence, that is, enlarged and flattened appearance. The senescent nature of these attached cells was further indicated by staining for SA-beta-gal. The mean telomere length was not changed after irradiation with C-ions. Phosphorylation of p53 at serine 15 as well as the expression of p21{sup WAF1/CIP1} was induced in NP-2 cells after irradiation. Furthermore, we found that irradiation with C-ions induced cellular senescence in a human glioma cell line lacking functional p53. Conclusions: Irradiation with C-ions induced apoptosis, autophagy, and cellular senescence in human glioma cells.« less

Preparation, cryopreservation, and growth of cells prepared from the green turtle (Chelonia mydas)

USGS Publications Warehouse

Moore, Melody K.; Work, Thierry M.; Balazs, George H.; Docherty, Douglas E.

1997-01-01

Techniques are described for preparing, preserving, and growing cell cultures from 30 to 40-day old green turtle embryos (2.0-3.0 cm length) including cells derived from skeletal muscle, liver, heart, kidney, eye, lung, and brain. Acceptable growth of all cells occurred in all standard cell culture media tested, with optimum growth temperature near 30??C. These cell cultures will be used in the study of sea turtle viral diseases including fibropapillomatosis, which is currently epidemic in some green turtle populations.

Transfected connexin45 alters gap junction permeability in cells expressing endogenous connexin43

PubMed Central

1995-01-01

Many cells express multiple connexins, the gap junction proteins that interconnect the cytosol of adjacent cells. Connexin43 (Cx43) channels allow intercellular transfer of Lucifer Yellow (LY, MW = 443 D), while connexin45 (Cx45) channels do not. We transfected full-length or truncated chicken Cx45 into a rat osteosarcoma cell line ROS-17/2.8, which expresses endogenous Cx43. Both forms of Cx45 were expressed at high levels and colocalized with Cx43 at plasma membrane junctions. Cells transfected with full-length Cx45 (ROS/Cx45) and cells transfected with Cx45 missing the 37 carboxyl-terminal amino acids (ROS/Cx45tr) showed 30-60% of the gap junctional conductance exhibited by ROS cells. Intercellular transfer of three negatively charged fluorescent reporter molecules was examined. In ROS cells, microinjected LY was transferred to an average of 11.2 cells/injected cell, while dye transfer between ROS/Cx45 cells was reduced to 3.9 transfer between ROS/Cx45 cells was reduced to 3.9 cells. In contrast, ROS/Cx45tr cells transferred LY to > 20 cells. Transfer of calcein (MW = 623 D) was also reduced by approximately 50% in ROS/Cx45 cells, but passage of hydroxycoumarin carboxylic acid (HCCA; MW = 206 D) was only reduced by 35% as compared to ROS cells. Thus, introduction of Cx45 altered intercellular coupling between cells expressing Cx43, most likely the result of direct interaction between Cx43 and Cx45. Transfection of Cx45tr and Cx45 had different effects in ROS cells, consistent with a role of the carboxyl-terminal domain of Cx45 in determining gap junction permeability or interactions between connexins. These data suggest that coexpression of multiple connexins may enable cells to achieve forms of intercellular communication that cannot be attained by expression of a single connexin. PMID:7642714

Thin n-i-p silicon solar cell

NASA Technical Reports Server (NTRS)

Meulenberg, A., Jr.; Allison, J. F.; Arndt, R. A.

1980-01-01

A space solar cell concept which combines high cell output with low diffusion length damage coefficients is presented for the purpose of reducing solar cell susceptibility to degradation from the radiation environment. High resistivity n-i-p silicon solar cells ranging from upward of 83 micron-cm were exposed to AM0 ultraviolet illumination. It is shown that high resistivity cells act as extrinsic devices under dark conditions and as intrinsic devices under AM0 illumination. Resistive losses in thin n-i-p cells are found to be comparable to those in low resistivity cells. Present voltage limitations appear to be due to generation and recombination in the diffused regions.

Stress and strain in the contractile and cytoskeletal filaments of airway smooth muscle.

PubMed

Deng, Linhong; Bosse, Ynuk; Brown, Nathan; Chin, Leslie Y M; Connolly, Sarah C; Fairbank, Nigel J; King, Greg G; Maksym, Geoffrey N; Paré, Peter D; Seow, Chun Y; Stephen, Newman L

2009-10-01

Stress and strain are omnipresent in the lung due to constant lung volume fluctuation associated with respiration, and they modulate the phenotype and function of all cells residing in the airways including the airway smooth muscle (ASM) cell. There is ample evidence that the ASM cell is very sensitive to its physical environment, and can alter its structure and/or function accordingly, resulting in either desired or undesired consequences. The forces that are either conferred to the ASM cell due to external stretching or generated inside the cell must be borne and transmitted inside the cytoskeleton (CSK). Thus, maintaining appropriate levels of stress and strain within the CSK is essential for maintaining normal function. Despite the importance, the mechanisms regulating/dysregulating ASM cytoskeletal filaments in response to stress and strain remained poorly understood until only recently. For example, it is now understood that ASM length and force are dynamically regulated, and both can adapt over a wide range of length, rendering ASM one of the most malleable living tissues. The malleability reflects the CSK's dynamic mechanical properties and plasticity, both of which strongly interact with the loading on the CSK, and all together ultimately determines airway narrowing in pathology. Here we review the latest advances in our understanding of stress and strain in ASM cells, including the organization of contractile and cytoskeletal filaments, range and adaptation of functional length, structural and functional changes of the cell in response to mechanical perturbation, ASM tone as a mediator of strain-induced responses, and the novel glassy dynamic behaviors of the CSK in relation to asthma pathophysiology.

Cutting Edge: Inflammasome Activation in Primary Human Macrophages Is Dependent on Flagellin

PubMed Central

Kortmann, Jens; Brubaker, Sky W.

2015-01-01

Murine NLR family, apoptosis inhibitory protein (Naip)1, Naip2, and Naip5/6 are host sensors that detect the cytosolic presence of needle and rod proteins from bacterial type III secretion systems and flagellin, respectively. Previous studies using human-derived macrophage-like cell lines indicate that human macrophages sense the cytosolic needle protein, but not bacterial flagellin. In this study, we show that primary human macrophages readily sense cytosolic flagellin. Infection of primary human macrophages with Salmonella elicits robust cell death and IL-1β secretion that is dependent on flagellin. We show that flagellin detection requires a full-length isoform of human Naip. This full-length Naip isoform is robustly expressed in primary macrophages from healthy human donors, but it is drastically reduced in monocytic tumor cells, THP-1, and U937, rendering them insensitive to cytosolic flagellin. However, ectopic expression of full-length Naip rescues the ability of U937 cells to sense flagellin. In conclusion, human Naip functions to activate the inflammasome in response to flagellin, similar to murine Naip5/6. PMID:26109648

An algorithm for the design and tuning of RF accelerating structures with variable cell lengths

NASA Astrophysics Data System (ADS)

Lal, Shankar; Pant, K. K.

2018-05-01

An algorithm is proposed for the design of a π mode standing wave buncher structure with variable cell lengths. It employs a two-parameter, multi-step approach for the design of the structure with desired resonant frequency and field flatness. The algorithm, along with analytical scaling laws for the design of the RF power coupling slot, makes it possible to accurately design the structure employing a freely available electromagnetic code like SUPERFISH. To compensate for machining errors, a tuning method has been devised to achieve desired RF parameters for the structure, which has been qualified by the successful tuning of a 7-cell buncher to π mode frequency of 2856 MHz with field flatness <3% and RF coupling coefficient close to unity. The proposed design algorithm and tuning method have demonstrated the feasibility of developing an S-band accelerating structure for desired RF parameters with a relatively relaxed machining tolerance of ∼ 25 μm. This paper discusses the algorithm for the design and tuning of an RF accelerating structure with variable cell lengths.

Mutant fatty acid desaturase

DOEpatents

Shanklin, John; Cahoon, Edgar B.

2004-02-03

The present invention relates to a method for producing mutants of a fatty acid desaturase having a substantially increased activity towards fatty acid substrates with chains containing fewer than 18 carbons relative to an unmutagenized precursor desaturase having an 18 carbon atom chain length substrate specificity. The method involves inducing one or more mutations in the nucleic acid sequence encoding the precursor desaturase, transforming the mutated sequence into an unsaturated fatty acid auxotroph cell such as MH13 E. coli, culturing the cells in the absence of supplemental unsaturated fatty acids, thereby selecting for recipient cells which have received and which express a mutant fatty acid desaturase with an elevated specificity for fatty acid substrates having chain lengths of less than 18 carbon atoms. A variety of mutants having 16 or fewer carbon atom chain length substrate specificities are produced by this method. Mutant desaturases produced by this method can be introduced via expression vectors into prokaryotic and eukaryotic cells and can also be used in the production of transgenic plants which may be used to produce specific fatty acid products.

COMMUNICATION: Alternative splicing and genomic stability

NASA Astrophysics Data System (ADS)

Cahill, Kevin

2004-06-01

Alternative splicing allows an organism to make different proteins in different cells at different times, all from the same gene. In a cell that uses alternative splicing, the total length of all the exons is much shorter than in a cell that encodes the same set of proteins without alternative splicing. This economical use of exons makes genes more stable during reproduction and development because a genome with a shorter exon length is more resistant to harmful mutations. Genomic stability may be the reason why higher vertebrates splice alternatively. For a broad class of alternatively spliced genes, a formula is given for the increase in their stability.

Coarse-Grained Models for Protein-Cell Membrane Interactions

PubMed Central

Bradley, Ryan; Radhakrishnan, Ravi

2015-01-01

The physiological properties of biological soft matter are the product of collective interactions, which span many time and length scales. Recent computational modeling efforts have helped illuminate experiments that characterize the ways in which proteins modulate membrane physics. Linking these models across time and length scales in a multiscale model explains how atomistic information propagates to larger scales. This paper reviews continuum modeling and coarse-grained molecular dynamics methods, which connect atomistic simulations and single-molecule experiments with the observed microscopic or mesoscale properties of soft-matter systems essential to our understanding of cells, particularly those involved in sculpting and remodeling cell membranes. PMID:26613047

Reduction of the immunostainable length of the hippocampal dentate granule cells' primary cilia in 3xAD-transgenic mice producing human A{beta}{sub 1-42} and tau

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

Chakravarthy, Balu, E-mail: Balu.Chakravarthy@nrc-cnrc.gc.ca; Gaudet, Chantal; Menard, Michel

2012-10-12

Highlights: Black-Right-Pointing-Pointer A{beta} and tau-induced neurofibrillary tangles play a key role in Alzheimer's disease. Black-Right-Pointing-Pointer A{beta}{sub 1-42} and mutant tau protein together reduce the primary cilium length. Black-Right-Pointing-Pointer This shortening likely reduces cilium-dependent neurogenesis and memory function. Black-Right-Pointing-Pointer This provides a model of an A{beta}/tau targeting of a neuronal signaling organelle. -- Abstract: The hippocampal dentate gyrus is one of the two sites of continuous neurogenesis in adult rodents and humans. Virtually all dentate granule cells have a single immobile cilium with a microtubule spine or axoneme covered with a specialized cell membrane loaded with receptors such as the somatostatinmore » receptor 3 (SSTR3), and the p75 neurotrophin receptor (p75{sup NTR}). The signals from these receptors have been reported to stimulate neuroprogenitor proliferation and the post-mitotic maturation of newborn granule cells into functioning granule cells. We have found that in 6-24-months-old triple transgenic Alzheimer's disease model mice (3xTg-AD) producing both A{beta}{sub 1-42} and the mutant human tau protein tau{sub P301L,} the dentate granule cells still had immunostainable SSTR3- and p75{sup NTR}-bearing cilia but they were only half the length of the immunostained cilia in the corresponding wild-type mice. However, the immunostainable length of the granule cell cilia was not reduced either in 2xTg-AD mice accumulating large amounts of A{beta}{sub 1-42} or in mice accumulating only a mutant human tau protein. Thus it appears that a combination of A{beta}{sub 1-42} and tau protein accumulation affects the levels of functionally important receptors in 3xTg-AD mice. These observations raise the important possibility that structural and functional changes in granule cell cilia might have a role in AD.« less

Mechanisms of mechanical strain memory in airway smooth muscle.

PubMed

Kim, Hak Rim; Hai, Chi-Ming

2005-10-01

We evaluated the hypothesis that mechanical deformation of airway smooth muscle induces structural remodeling of airway smooth muscle cells, thereby modulating mechanical performance in subsequent contractions. This hypothesis implied that past experience of mechanical deformation was retained (or "memorized") as structural changes in airway smooth muscle cells, which modulated the cell's subsequent contractile responses. We termed this phenomenon mechanical strain memory. Preshortening has been found to induce attenuation of both force and isotonic shortening velocity in cholinergic receptor-activated airway smooth muscle. Rapid stretching of cholinergic receptor-activated airway smooth muscle from an initial length to a final length resulted in post-stretch force and myosin light chain phosphorylation that correlated significantly with initial length. Thus post-stretch muscle strips appeared to retain memory of the initial length prior to rapid stretch (mechanical strain memory). Cytoskeletal recruitment of actin- and integrin-binding proteins and Erk 1/2 MAPK appeared to be important mechanisms of mechanical strain memory. Sinusoidal length oscillation led to force attenuation during oscillation and in subsequent contractions in intact airway smooth muscle, and p38 MAPK appeared to be an important mechanism. In contrast, application of local mechanical strain to cultured airway smooth muscle cells induced local actin polymerization and cytoskeletal stiffening. It is conceivable that deep inspiration-induced bronchoprotection may be a manifestation of mechanical strain memory such that mechanical deformation from past breathing cycles modulated the mechanical performance of airway smooth muscle in subsequent cycles in a continuous and dynamic manner.

Stimulus-dependent modulation of spike burst length in cat striate cortical cells.

PubMed

DeBusk, B C; DeBruyn, E J; Snider, R K; Kabara, J F; Bonds, A B

1997-07-01

Burst activity, defined by groups of two or more spikes with intervals of < or = 8 ms, was analyzed in responses to drifting sinewave gratings elicited from striate cortical neurons in anesthetized cats. Bursting varied broadly across a population of 507 simple and complex cells. Half of this population had > or = 42% of their spikes contained in bursts. The fraction of spikes in bursts did not vary as a function of average firing rate and was stationary over time. Peaks in the interspike interval histograms were found at both 3-5 ms and 10-30 ms. In many cells the locations of these peaks were independent of firing rate, indicating a quantized control of firing behavior at two different time scales. The activity at the shorter time scale most likely results from intrinsic properties of the cell membrane, and that at the longer scale from recurrent network excitation. Burst frequency (bursts per s) and burst length (spikes per burst) both depended on firing rate. Burst frequency was essentially linear with firing rate, whereas burst length was a nonlinear function of firing rate and was also governed by stimulus orientation. At a given firing rate, burst length was greater for optimal orientations than for nonoptimal orientations. No organized orientation dependence was seen in bursts from lateral geniculate nucleus cells. Activation of cortical contrast gain control at low response amplitudes resulted in no burst length modulation, but burst shortening at optimal orientations was found in responses characterized by supersaturation. At a given firing rate, cortical burst length was shortened by microinjection of gamma-aminobutyric acid (GABA), and bursts became longer in the presence of N-methyl-bicuculline, a GABA(A) receptor blocker. These results are consistent with a model in which responses are reduced at nonoptimal orientations, at least in part, by burst shortening that is mediated by GABA. A similar mechanism contributes to response supersaturation at high contrasts via recruitment of inhibitory responses that are tuned to adjacent orientations. Burst length modulation can serve as a form of coding by supporting dynamic, stimulus-dependent reorganization of the effectiveness of individual network connections.

Diffused junction p(+)-n solar cells in bulk GaAs. II - Device characterization and modelling

NASA Technical Reports Server (NTRS)

Keeney, R.; Sundaram, L. M. G.; Rode, H.; Bhat, I.; Ghandhi, S. K.; Borrego, J. M.

1984-01-01

The photovoltaic characteristics of p(+)-n junction solar cells fabricated on bulk GaAs by an open tube diffusion technique are presented in detail. Quantum efficiency measurements were analyzed and compared to computer simulations of the cell structure in order to determine material parameters such as diffusion length, surface recombination velocity and junction depth. From the results obtained it is projected that proper optimization of the cell parameters can increase the efficiency of the cells to close to 20 percent.

Fidelity of DNA Replication in Normal and Malignant Human Breast Cells

DTIC Science & Technology

1998-07-01

synthesome has been extensively demonstrated to carry out full length DNA replication in vitro, and to accurately depict the DNA replication process as it...occurs in the intact cell. By examining the fidelity of the DNA replication process carried out by the DNA synthesome from a number of breast cell types...we have demonstrated for the first time, that the cellular DNA replication machinery of malignant human breast cells is significantly more error-prone than that of non- malignant human breast cells.

Cell response to quasi-monochromatic light with different coherence

NASA Astrophysics Data System (ADS)

Budagovsky, A. V.; Solovykh, N. V.; Budagovskaya, O. N.; Budagovsky, I. A.

2015-04-01

The problem of the light coherence effect on the magnitude of the photoinduced cell response is discussed. The origins of ambiguous interpretation of the known experimental results are considered. Using the biological models, essentially differing in anatomy, morphology and biological functions (acrospires of radish, blackberry microsprouts cultivated in vitro, plum pollen), the effect of statistical properties of quasi-monochromatic light (λmax = 633 nm) on the magnitude of the photoinduced cell response is shown. It is found that for relatively low spatial coherence, the cell functional activity changes insignificantly. The maximal enhancement of growing processes (stimulating effect) is observed when the coherence length Lcoh and the correlation radius rcor are greater than the cell size, i.e., the entire cell fits into the field coherence volume. In this case, the representative indicators (germination of seeds and pollen, the spears length) exceeds those of non-irradiated objects by 1.7 - 3.9 times. For more correct assessment of the effect of light statistical properties on photocontrol processes, it is proposed to replace the qualitative description (coherent - incoherent) with the quantitative one, using the determination of spatial and temporal correlation functions and comparing them with the characteristic dimensions of the biological structures, e.g., the cell size.

Experiment K-7-17: Effects of Spaceflight on the Proliferation of Jejunal Mucosal Cells

NASA Technical Reports Server (NTRS)

Phillips, R. W.; Moeller, C. L.; Sawyer, H. R.; Smirnov, K. L.

1994-01-01

The purpose of this project was to test the hypothesis that the generalized, whole body decrease in synthetic activity due to microgravity conditions encountered during spaceflight would be demonstrable in cells and tissues characterized by a rapid rate of turnover. Jejunal mucosal cells were chosen as a model since these cells are among the most rapidly proliferating in the body. Accordingly, the percentage of mitotic cells present in the crypts of Lieberkuhn in each of 5 rats flown on the COSMOS 2044 mission were compared to the percentage of mitotic cells present in the crypts in rats included in each of 3 ground control groups (i.e., vivarium, synchronous and caudal-elevated). No significant difference (p greater than .05) was detected in mitotic indices between the flight and vivarium group. Although the ability of jejunal mucosal cells to divide by mitosis was not impaired in flight group, there was, however, a reduction in the length of villi and depth of crypts. The concommitant reduction in villus length and crypth depth in the flight group probably reflects changes in connective tissue components within the core of villi.

Imaging and Scattering Measurements for Diesel Spray Combustion: Optical Development and Phenomenological Studies

DTIC Science & Technology

2012-09-30

the CS2 was contained in a rectangular colorimeter cell with a custom built Teflon cap to alleviate the evaporation of the hazardous chemical...6: A comparison of the image quality between the older colorimeter cell (a) and the new containment cell (b). 2.5 Autocorrelation-Based Pulse Length

Failure analysis of fuel cell electrodes using three-dimensional multi-length scale X-ray computed tomography

NASA Astrophysics Data System (ADS)

Pokhrel, A.; El Hannach, M.; Orfino, F. P.; Dutta, M.; Kjeang, E.

2016-10-01

X-ray computed tomography (XCT), a non-destructive technique, is proposed for three-dimensional, multi-length scale characterization of complex failure modes in fuel cell electrodes. Comparative tomography data sets are acquired for a conditioned beginning of life (BOL) and a degraded end of life (EOL) membrane electrode assembly subjected to cathode degradation by voltage cycling. Micro length scale analysis shows a five-fold increase in crack size and 57% thickness reduction in the EOL cathode catalyst layer, indicating widespread action of carbon corrosion. Complementary nano length scale analysis shows a significant reduction in porosity, increased pore size, and dramatically reduced effective diffusivity within the remaining porous structure of the catalyst layer at EOL. Collapsing of the structure is evident from the combination of thinning and reduced porosity, as uniquely determined by the multi-length scale approach. Additionally, a novel image processing based technique developed for nano scale segregation of pore, ionomer, and Pt/C dominated voxels shows an increase in ionomer volume fraction, Pt/C agglomerates, and severe carbon corrosion at the catalyst layer/membrane interface at EOL. In summary, XCT based multi-length scale analysis enables detailed information needed for comprehensive understanding of the complex failure modes observed in fuel cell electrodes.

Actin kinetics shapes cortical network structure and mechanics

PubMed Central

Fritzsche, Marco; Erlenkämper, Christoph; Moeendarbary, Emad; Charras, Guillaume; Kruse, Karsten

2016-01-01

The actin cortex of animal cells is the main determinant of cellular mechanics. The continuous turnover of cortical actin filaments enables cells to quickly respond to stimuli. Recent work has shown that most of the cortical actin is generated by only two actin nucleators, the Arp2/3 complex and the formin Diaph1. However, our understanding of their interplay, their kinetics, and the length distribution of the filaments that they nucleate within living cells is poor. Such knowledge is necessary for a thorough comprehension of cellular processes and cell mechanics from basic polymer physics principles. We determined cortical assembly rates in living cells by using single-molecule fluorescence imaging in combination with stochastic simulations. We find that formin-nucleated filaments are, on average, 10 times longer than Arp2/3-nucleated filaments. Although formin-generated filaments represent less than 10% of all actin filaments, mechanical measurements indicate that they are important determinants of cortical elasticity. Tuning the activity of actin nucleators to alter filament length distribution may thus be a mechanism allowing cells to adjust their macroscopic mechanical properties to their physiological needs. PMID:27152338

Actin kinetics shapes cortical network structure and mechanics.

PubMed

Fritzsche, Marco; Erlenkämper, Christoph; Moeendarbary, Emad; Charras, Guillaume; Kruse, Karsten

2016-04-01

The actin cortex of animal cells is the main determinant of cellular mechanics. The continuous turnover of cortical actin filaments enables cells to quickly respond to stimuli. Recent work has shown that most of the cortical actin is generated by only two actin nucleators, the Arp2/3 complex and the formin Diaph1. However, our understanding of their interplay, their kinetics, and the length distribution of the filaments that they nucleate within living cells is poor. Such knowledge is necessary for a thorough comprehension of cellular processes and cell mechanics from basic polymer physics principles. We determined cortical assembly rates in living cells by using single-molecule fluorescence imaging in combination with stochastic simulations. We find that formin-nucleated filaments are, on average, 10 times longer than Arp2/3-nucleated filaments. Although formin-generated filaments represent less than 10% of all actin filaments, mechanical measurements indicate that they are important determinants of cortical elasticity. Tuning the activity of actin nucleators to alter filament length distribution may thus be a mechanism allowing cells to adjust their macroscopic mechanical properties to their physiological needs.

Quantitative fluorescence in situ hybridization measurement of telomere length in skin with/without sun exposure or actinic keratosis.

PubMed

Ikeda, Hiroyuki; Aida, Junko; Hatamochi, Atsushi; Hamasaki, Yoichiro; Izumiyama-Shimomura, Naotaka; Nakamura, Ken-Ichi; Ishikawa, Naoshi; Poon, Steven S; Fujiwara, Mutsunori; Tomita, Ken-Ichiro; Hiraishi, Naoki; Kuroiwa, Mie; Matsuura, Masaaki; Sanada, Yukihiro; Kawano, Youichi; Arai, Tomio; Takubo, Kaiyo

2014-03-01

Chromosomal and genomic instability due to telomere dysfunction is known to play an important role in carcinogenesis. To study telomere shortening in the epidermis surrounding actinic keratosis, we measured telomere lengths of basal, parabasal, and suprabasal cells in epidermis with actinic keratosis (actinic keratosis group, n = 18) and without actinic keratosis (sun-protected, n = 15, and sun-exposed, n = 13 groups) and in actinic keratosis itself as well as in dermal fibroblasts in the 3 groups, using quantitative fluorescence in situ hybridization. Among the 3 cell types, telomeres of basal cells were not always the longest, suggesting that tissue stem cells are not necessarily located among basal cells. Telomeres of basal cells in the sun-exposed group were shorter than those in the sun-protected group. Telomeres in the background of actinic keratosis and in actinic keratosis itself and those of fibroblasts in actinic keratosis were significantly shorter than those in the controls. Our findings demonstrate that sun exposure induces telomere shortening and that actinic keratosis arises from epidermis with shorter telomeres despite the absence of any histologic atypia. © 2014.

Enhancing light absorption within the carrier transport length in quantum junction solar cells.

PubMed

Fu, Yulan; Hara, Yukihiro; Miller, Christopher W; Lopez, Rene

2015-09-10

Colloidal quantum dot (CQD) solar cells have attracted tremendous attention because of their tunable absorption spectrum window and potentially low processing cost. Recently reported quantum junction solar cells represent a promising approach to building a rectifying photovoltaic device that employs CQD layers on each side of the p-n junction. However, the ultimate efficiency of CQD solar cells is still highly limited by their high trap state density in both p- and n-type CQDs. By modeling photonic structures to enhance the light absorption within the carrier transport length and by ensuring that the carrier generation and collection efficiencies were both augmented, our work shows that overall device current density could be improved. We utilized a two-dimensional numerical model to calculate the characteristics of patterned CQD solar cells based on a simple grating structure. Our calculation predicts a short circuit current density as high as 31  mA/cm², a value nearly 1.5 times larger than that of the conventional flat design, showing the great potential value of patterned quantum junction solar cells.

A Semi-Automatic Method for Image Analysis of Edge Dynamics in Living Cells

PubMed Central

Huang, Lawrence; Helmke, Brian P.

2011-01-01

Spatial asymmetry of actin edge ruffling contributes to the process of cell polarization and directional migration, but mechanisms by which external cues control actin polymerization near cell edges remain unclear. We designed a quantitative image analysis strategy to measure the spatiotemporal distribution of actin edge ruffling. Time-lapse images of endothelial cells (ECs) expressing mRFP-actin were segmented using an active contour method. In intensity line profiles oriented normal to the cell edge, peak detection identified the angular distribution of polymerized actin within 1 µm of the cell edge, which was localized to lamellipodia and edge ruffles. Edge features associated with filopodia and peripheral stress fibers were removed. Circular statistical analysis enabled detection of cell polarity, indicated by a unimodal distribution of edge ruffles. To demonstrate the approach, we detected a rapid, nondirectional increase in edge ruffling in serum-stimulated ECs and a change in constitutive ruffling orientation in quiescent, nonpolarized ECs. Error analysis using simulated test images demonstrate robustness of the method to variations in image noise levels, edge ruffle arc length, and edge intensity gradient. These quantitative measurements of edge ruffling dynamics enable investigation at the cellular length scale of the underlying molecular mechanisms regulating actin assembly and cell polarization. PMID:21643526

Mycobacterium tuberculosis GroEL2 Modulates Dendritic Cell Responses.

PubMed

Georgieva, Maria; Sia, Jonathan Kevin; Bizzell, Erica; Madan-Lala, Ranjna; Rengarajan, Jyothi

2018-02-01

Mycobacterium tuberculosis successfully subverts the host immune response to promote disease progression. In addition to its known intracellular niche in macrophages, M. tuberculosis interferes with the functions of dendritic cells (DCs), which are the primary antigen-presenting cells of the immune system. We previously showed that M. tuberculosis dampens proinflammatory responses and impairs DC functions through the cell envelope-associated serine protease Hip1. Here we present data showing that M. tuberculosis GroEL2, a substrate of Hip1, modulates DC functions. The full-length GroEL2 protein elicited robust proinflammatory responses from DCs and promoted DC maturation and antigen presentation to T cells. In contrast, the cleaved form of GroEL2, which predominates in M. tuberculosis , was poorly immunostimulatory and was unable to promote DC maturation and antigen presentation. Moreover, DCs exposed to full-length, but not cleaved, GroEL2 induced strong antigen-specific gamma interferon (IFN-γ), interleukin-2 (IL-2), and IL-17A cytokine responses from CD4 + T cells. Moreover, the expression of cleaved GroEL2 in the hip1 mutant restored the robust T cell responses to wild-type levels, suggesting that proteolytic cleavage of GroEL2 allows M. tuberculosis to prevent optimal DC-T cell cross talk during M. tuberculosis infection. Copyright © 2018 American Society for Microbiology.

Slot-coupled CW standing wave accelerating cavity

DOEpatents

Wang, Shaoheng; Rimmer, Robert; Wang, Haipeng

2017-05-16

A slot-coupled CW standing wave multi-cell accelerating cavity. To achieve high efficiency graded beta acceleration, each cell in the multi-cell cavity may include different cell lengths. Alternatively, to achieve high efficiency with acceleration for particles with beta equal to 1, each cell in the multi-cell cavity may include the same cell design. Coupling between the cells is achieved with a plurality of axially aligned kidney-shaped slots on the wall between cells. The slot-coupling method makes the design very compact. The shape of the cell, including the slots and the cone, are optimized to maximize the power efficiency and minimize the peak power density on the surface. The slots are non-resonant, thereby enabling shorter slots and less power loss.

Coupling between apical tension and basal adhesion allow epithelia to collectively sense and respond to substrate topography over long distances.

PubMed

Broaders, Kyle E; Cerchiari, Alec E; Gartner, Zev J

2015-12-01

Epithelial sheets fold into complex topographies that contribute to their function in vivo. Cells can sense and respond to substrate topography in their immediate vicinity by modulating their interfacial mechanics, but the extent to which these mechanical properties contribute to their ability to sense substrate topography across length scales larger than a single cell has not been explored in detail. To study the relationship between the interfacial mechanics of single cells and their collective behavior as tissues, we grew cell-sheets on substrates engraved with surface features spanning macroscopic length-scales. We found that many epithelial cell-types sense and respond to substrate topography, even when it is locally nearly planar. Cells clear or detach from regions of local negative curvature, but not from regions with positive or no curvature. We investigated this phenomenon using a finite element model where substrate topography is coupled to epithelial response through a balance of tissue contractility and adhesive forces. The model correctly predicts the focal sites of cell-clearing and epithelial detachment. Furthermore, the model predicts that local tissue response to substrate curvature is a function of the surrounding topography of the substrate across long distances. Analysis of cell-cell and cell-substrate contact angles suggests a relationship between these single-cell interfacial properties, epithelial interfacial properties, and collective epithelial response to substrate topography. Finally, we show that contact angles change upon activation of oncogenes or inhibition of cell-contractility, and that these changes correlate with collective epithelial response. Our results demonstrate that in mechanically integrated epithelial sheets, cell contractility can be transmitted through multiple cells and focused by substrate topography to affect a behavioral response at distant sites.

Fatty acids from membrane lipids become incorporated into lipid bodies during Myxococcus xanthus differentiation.

PubMed

Bhat, Swapna; Boynton, Tye O; Pham, Dan; Shimkets, Lawrence J

2014-01-01

Myxococcus xanthus responds to amino acid limitation by producing fruiting bodies containing dormant spores. During development, cells produce triacylglycerides in lipid bodies that become consumed during spore maturation. As the cells are starved to induce development, the production of triglycerides represents a counterintuitive metabolic switch. In this paper, lipid bodies were quantified in wild-type strain DK1622 and 33 developmental mutants at the cellular level by measuring the cross sectional area of the cell stained with the lipophilic dye Nile red. We provide five lines of evidence that triacylglycerides are derived from membrane phospholipids as cells shorten in length and then differentiate into myxospores. First, in wild type cells, lipid bodies appear early in development and their size increases concurrent with an 87% decline in membrane surface area. Second, developmental mutants blocked at different stages of shortening and differentiation accumulated lipid bodies proportionate with their cell length with a Pearson's correlation coefficient of 0.76. Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten. Fourth, genes for fatty acid synthesis are down-regulated while genes for fatty acid degradation are up regulated. Finally, direct movement of fatty acids from membrane lipids in growing cells to lipid bodies in developing cells was observed by pulse labeling cells with palmitate. Recycling of lipids released by Programmed Cell Death appears not to be necessary for lipid body production as a fadL mutant was defective in fatty acid uptake but proficient in lipid body production. The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation. MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes.

Development of software and modification of Q-FISH protocol for estimation of individual telomere length in immunopathology.

PubMed

Barkovskaya, M Sh; Bogomolov, A G; Knauer, N Yu; Rubtsov, N B; Kozlov, V A

2017-04-01

Telomere length is an important indicator of proliferative cell history and potential. Decreasing telomere length in the cells of an immune system can indicate immune aging in immune-mediated and chronic inflammatory diseases. Quantitative fluorescent in situ hybridization (Q-FISH) of a labeled (C 3 TA[Formula: see text] peptide nucleic acid probe onto fixed metaphase cells followed by digital image microscopy allows the evaluation of telomere length in the arms of individual chromosomes. Computer-assisted analysis of microscopic images can provide quantitative information on the number of telomeric repeats in individual telomeres. We developed new software to estimate telomere length. The MeTeLen software contains new options that can be used to solve some Q-FISH and microscopy problems, including correction of irregular light effects and elimination of background fluorescence. The identification and description of chromosomes and chromosome regions are essential to the Q-FISH technique. To improve the quality of cytogenetic analysis after Q-FISH, we optimized the temperature and time of DNA-denaturation to get better DAPI-banding of metaphase chromosomes. MeTeLen was tested by comparing telomere length estimations for sister chromatids, background fluorescence estimations, and correction of nonuniform light effects. The application of the developed software for analysis of telomere length in patients with rheumatoid arthritis was demonstrated.

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 migration or cytokinesis and proliferation? – Revisiting the “go or grow” hypothesis in cancer cells in vitro

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

Garay, Tamás; Juhász, Éva; Molnár, Eszter

The mortality of patients with solid tumors is mostly due to metastasis that relies on the interplay between migration and proliferation. The “go or grow” hypothesis postulates that migration and proliferation spatiotemporally excludes each other. We evaluated this hypothesis on 35 cell lines (12 mesothelioma, 13 melanoma and 10 lung cancer) on both the individual cell and population levels. Following three-day-long videomicroscopy, migration, proliferation and cytokinesis-length were quantified. We found a significantly higher migration in mesothelioma cells compared to melanoma and lung cancer while tumor types did not differ in mean proliferation or duration of cytokinesis. Strikingly, we found inmore » melanoma and lung cancer a significant positive correlation between mean proliferation and migration. Furthermore, non-dividing melanoma and lung cancer cells displayed slower migration. In contrast, in mesothelioma there were no such correlations. Interestingly, negative correlation was found between cytokinesis-length and migration in melanoma. FAK activation was higher in melanoma cells with high motility. We demonstrate that the cancer cells studied do not defer proliferation for migration. Of note, tumor cells from various organ systems may differently regulate migration and proliferation. Furthermore, our data is in line with the observation of pathologists that highly proliferative tumors are often highly invasive. - Highlights: • We investigated the “go or grow” hypothesis in human cancer cells in vitro. • Proliferation and migration positively correlate in melanoma and lung cancer cells. • Duration of cytokinesis and migration shows inverse correlation. • Increased FAK activation is present in highly motile melanoma cells.« less

Telomere maintenance in laser capture microdissection-purified Barrett's adenocarcinoma cells and effect of telomerase inhibition in vivo.

PubMed

Shammas, Masood A; Qazi, Aamer; Batchu, Ramesh B; Bertheau, Robert C; Wong, Jason Y Y; Rao, Manjula Y; Prasad, Madhu; Chanda, Diptiman; Ponnazhagan, Selvarangan; Anderson, Kenneth C; Steffes, Christopher P; Munshi, Nikhil C; De Vivo, Immaculata; Beer, David G; Gryaznov, Sergei; Weaver, Donald W; Goyal, Raj K

2008-08-01

The aims of this study were to investigate telomere function in normal and Barrett's esophageal adenocarcinoma (BEAC) cells purified by laser capture microdissection and to evaluate the effect of telomerase inhibition in cancer cells in vitro and in vivo. Epithelial cells were purified from surgically resected esophagi. Telomerase activity was measured by modified telomeric repeat amplification protocol and telomere length was determined by real-time PCR assay. To evaluate the effect of telomerase inhibition, adenocarcinoma cell lines were continuously treated with a specific telomerase inhibitor (GRN163L) and live cell number was determined weekly. Apoptosis was evaluated by Annexin labeling and senescence by beta-galactosidase staining. For in vivo studies, severe combined immunodeficient mice were s.c. inoculated with adenocarcinoma cells and following appearance of palpable tumors, injected i.p. with saline or GRN163L. Telomerase activity was significantly elevated whereas telomeres were shorter in BEAC cells relative to normal esophageal epithelial cells. The treatment of adenocarcinoma cells with telomerase inhibitor, GRN163L, led to loss of telomerase activity, reduction in telomere length, and growth arrest through induction of both the senescence and apoptosis. GRN163L-induced cell death could also be expedited by addition of the chemotherapeutic agents doxorubicin and ritonavir. Finally, the treatment with GRN163L led to a significant reduction in tumor volume in a subcutaneous tumor model. We show that telomerase activity is significantly elevated whereas telomeres are shorter in BEAC and suppression of telomerase inhibits proliferation of adenocarcinoma cells both in vitro and in vivo.

Activity-dependent self-regulation of viscous length scales in biological systems

NASA Astrophysics Data System (ADS)

Nandi, Saroj Kumar

2018-05-01

The cellular cortex, which is a highly viscous thin cytoplasmic layer just below the cell membrane, controls the cell's mechanical properties, which can be characterized by a hydrodynamic length scale ℓ . Cells actively regulate ℓ via the activity of force-generating molecules, such as myosin II. Here we develop a general theory for such systems through a coarse-grained hydrodynamic approach including activity in the static description of the system providing an experimentally accessible parameter and elucidate the detailed mechanism of how a living system can actively self-regulate its hydrodynamic length scale, controlling the rigidity of the system. Remarkably, we find that ℓ , as a function of activity, behaves universally and roughly inversely proportional to the activity of the system. Our theory rationalizes a number of experimental findings on diverse systems, and comparison of our theory with existing experimental data shows good agreement.

Photoperiod length paces the temporal orchestration of cell cycle and carbon-nitrogen metabolism in Crocosphaera watsonii.

PubMed

Dron, Anthony; Rabouille, Sophie; Claquin, Pascal; Talec, Amélie; Raimbault, Virginie; Sciandra, Antoine

2013-12-01

We analysed the effect of photoperiod length (PPL) (16:8 and 8:16 h of light-dark regime, named long and short PPL, respectively) on the temporal orchestration of the two antagonistic, carbon and nitrogen acquisitions in the unicellular, diazotrophic cyanobacterium Crocosphaera watsonii strain WH8501 growing diazotrophically. Carbon and nitrogen metabolism were monitored at high frequency, and their patterns were compared with the cell cycle progression. The oxygen-sensitive N2 fixation process occurred mainly during the dark period, where photosynthesis cannot take place, inducing a light-dark cycle of cellular C : N ratio. Examination of circadian patterns in the cell cycle revealed that cell division occurred during the midlight period, (8 h and 4 h into the light in the long and short PPL conditions, respectively), thus timely separated from the energy-intensive diazotrophic process. Results consistently show a nearly 5 h time lag between the end of cell division and the onset of N2 fixation. Shorter PPLs affected DNA compaction of C. watsonii cells and also led to a decrease in the cell division rate. Therefore, PPL paces the growth of C. watsonii: a long PPL enhances cell division while a short PPL favours somatic growth (biomass production) with higher carbon and nitrogen cell contents. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

Highly efficient single-junction GaAs thin-film solar cell on flexible substrate.

PubMed

Moon, Sunghyun; Kim, Kangho; Kim, Youngjo; Heo, Junseok; Lee, Jaejin

2016-07-20

There has been much interest in developing a thin-film solar cell because it is lightweight and flexible. The GaAs thin-film solar cell is a top contender in the thin-film solar cell market in that it has a high power conversion efficiency (PCE) compared to that of other thin-film solar cells. There are two common structures for the GaAs solar cell: n (emitter)-on-p (base) and p-on-n. The former performs better due to its high collection efficiency because the electron diffusion length of the p-type base region is much longer than the hole diffusion length of the n-type base region. However, it has been limited to fabricate highly efficient n-on-p single-junction GaAs thin film solar cell on a flexible substrate due to technical obstacles. We investigated a simple and fast epitaxial lift-off (ELO) method that uses a stress originating from a Cr/Au bilayer on a 125-μm-thick flexible substrate. A metal combination of AuBe/Pt/Au is employed as a new p-type ohmic contact with which an n-on-p single-junction GaAs thin-film solar cell on flexible substrate was successfully fabricated. The PCE of the fabricated single-junction GaAs thin-film solar cells reached 22.08% under air mass 1.5 global illumination.

Intact Arabidopsis RPB1 functions in stem cell niches maintenance and cell cycling control.

PubMed

Zhang, Qian-Qian; Li, Ying; Fu, Zhao-Ying; Liu, Xun-Biao; Yuan, Kai; Fang, Ying; Liu, Yan; Li, Gang; Zhang, Xian-Sheng; Chong, Kang; Ge, Lei

2018-05-12

Plant meristem activity depends on accurate execution of transcriptional networks required for establishing optimum functioning of stem cell niches. An Arabidopsis mutant card1-1 (constitutive auxin response with DR5:GFP) that encodes a truncated RPB1 (RNA Polymerase II's largest subunit) with shortened C-terminal domain (CTD) was identified. Phosphorylation of the CTD repeats of RPB1 is coupled to transcription in eukaryotes. Here we uncover that the truncated CTD of RPB1 disturbed cell cycling and enlarged the size of shoot and root meristem. The defects in patterning of root stem cell niche in card1-1 indicates that intact CTD of RPB1 is necessary for fine-tuning the specific expression of genes responsible for cell-fate determination. The gene-edited plants with different CTD length of RPB1, created by CRISPR-CAS9 technology, confirmed that both the full length and the DK-rich tail of RPB1's CTD play roles in the accurate transcription of CYCB1;1 encoding a cell-cycle marker protein in root meristem and hence participate in maintaining root meristem size. Our experiment proves that the intact RPB1 CTD is necessary for stem cell niche maintenance, which is mediated by transcriptional regulation of cell cycling genes. © 2018 The Authors The Plant Journal © 2018 John Wiley & Sons Ltd.

Ontogeny of serotonin-immunoreactive cells in the gut epithelium of the cloudy dogfish, Scyliorhinus torazame, with reference to coexistence of serotonin and neuropeptide Y.

PubMed

Chiba, A

1998-09-01

The ontogeny of serotonin (5-HT)-immunoreactive (IR) cells in the gut epithelium of an oviparous elasmobranch, Scyliorhinus torazame, was examined immunohistochemically. 5-HT-IR cells first appeared in the proximal part of the vitellointestinal duct (VID) and in the anterior part of the midgut of the embryo (30 mm in total length). At the 40-mm stage, the cells slightly increased in number and spread to the posterior part of the midgut, but no labeled cells were found in the foregut or hindgut. By the late embryonic (74- and 80-mm) stages, 5-HT-IR cells were markedly increased in number in the spiral intestine and stomach, whereas they were few in the VID and rectum. During these stages, the density of the cells in the inner yolk sac, the derivative of the VID, tended to be increased. This tendency seemed to be consistent in the posthatching juveniles at the 95-mm stage. In juveniles, 125 mm in length and 1.7 months after hatching, the cells further increased in number in the spiral intestine, reaching their adult value. Double immunostaining by the use of anti-5-HT and -neuropeptide Y (NPY) antisera demonstrated that some of the 5-HT-IR cells were also positive for NPY. Copyright 1998 Academic Press.

Improved OSC Amtec generator design to meet goals of JPL's candidate Europa Orbiter mission

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

Schock, A.; Noravian, H.; Or, C.

1998-07-01

The preceding paper (Paper IECEC.98.244) described OSC's initial designs of AMTEC (Alkali Metal Thermal-to-Electrical Conversion) power systems, consisting of one or two generators, each with 2, 3, or 4 General Purpose Heat Source (GPHS) modules and with 16 refractory AMTEC cells containing 5 Beta Alumina Solid Electrolyte (BASE) tubes; and presented the effect of heat input and voltage output on the generator's BOM evaporator and clad temperatures and on its EOM system efficiency and power output. Comparison of the computed results with JPL's goals for the Europa Orbiter mission showed that all of the initial 16-cell design options yielded eithermore » excessive evaporator and clad temperatures or insufficient EOM power to satisfy the JPL-specified mission goals. The present paper describes modified OSC generator designs with different numbers of AMTEC cells, cell diameters, cell lengths, cell materials, BASE tube lengths, and number of tubes per cell. These efforts succeeded in identifying generator designs with only half the number of AMTEC cells which -- for the same assumptions -- can produce EOM power outputs substantially in excess of JPL's goals for NASA's Europa Orbiter mission while operating well below the prescribed BOM limits on evaporator and clad temperature; and revealed that lowering the emissivity of the generator's housing to raise the cells' condenser temperatures can achieve substantial additional performance improvement. Finally, the paper culminates in programmatic recommendations.« less

Developmental kinetics of pig embryos by parthenogenetic activation or by handmade cloning.

PubMed

Li, J; Li, R; Liu, Y; Villemoes, K; Purup, S; Callesen, H

2013-10-01

The developmental kinetics of pig embryos produced by parthenogenetic activation without (PAZF) or with (PAZI) zona pellucida or by handmade cloning (HMC) was compared by time-lapse videography. After cumulus cell removal, the matured oocytes were either left zona intact (PAZI) or were made zona free by pronase digestion (PAZF) before they were activated (PA). Other matured oocytes were used for HMC based on foetal fibroblast cells. On Day 0 (day of PA or reconstruction), the embryos were cultured for 7 days in vitro in our time-lapse system. Pictures were taken every 30 min, and afterwards, each cell cycle was identified for each embryo to be analysed. Results showed that the PA embryos (both PAZF and PAZI) had shorter first cell cycle compared with HMC (17.4. 17.8 vs 23.6 h), but had a longer time length from four cell to morula stages (57.9, 53.8 vs 44.9 h). However, at the second cell cycle, PAZF embryos needed shorter time, while PAZI embryos had similar time length as HMC embryos, and both were longer than PAZF (23.4, 24.8 vs 14.6 h). Both PAZF and PAZI embryos used similar time to reach the blastocyst stage, and this was later than HMC embryos. In addition, when all of these embryos were grouped into viable (developed to blastocysts) and non-viable (not developed to blastocysts), the only difference in the time length was observed on the first cell cycle (18.6 vs 24.5 h), but not on the later cell cycles. In conclusion, our results not only give detailed information regarding the time schedule of in vitro-handled pig embryos, but also indicate that the first cell cycle could be used as a selecting marker for embryo viability. However, to evaluate the effect of the produced techniques, the whole time schedule of the pre-implantation developmental kinetics should be observed. © 2013 Blackwell Verlag GmbH.

Telomere Dynamics and Homeostasis in a Transmissible Cancer

PubMed Central

Ujvari, Beata; Pearse, Anne-Maree; Taylor, Robyn; Pyecroft, Stephen; Flanagan, Cassandra; Gombert, Sara; Papenfuss, Anthony T.; Madsen, Thomas; Belov, Katherine

2012-01-01

Background Devil Facial Tumour Disease (DFTD) is a unique clonal cancer that threatens the world's largest carnivorous marsupial, the Tasmanian devil (Sarcophilus harrisii) with extinction. This transmissible cancer is passed between individual devils by cell implantation during social interactions. The tumour arose in a Schwann cell of a single devil over 15 years ago and since then has expanded clonally, without showing signs of replicative senescence; in stark contrast to a somatic cell that displays a finite capacity for replication, known as the “Hayflick limit”. Methodology/Principal Findings In the present study we investigate the role of telomere length, measured as Telomere Copy Number (TCN), and telomerase and shelterin gene expression, as well as telomerase activity in maintaining hyperproliferation of Devil Facial Tumour (DFT) cells. Our results show that DFT cells have short telomeres. DFTD TCN does not differ between geographic regions or between strains. However, TCN has increased over time. Unlimited cell proliferation is likely to have been achieved through the observed up-regulation of the catalytic subunit of telomerase (TERT) and concomitant activation of telomerase. Up-regulation of the central component of shelterin, the TRF1-intercating nuclear factor 2 (TINF2) provides DFT a mechanism for telomere length homeostasis. The higher expression of both TERT and TINF2 may also protect DFT cells from genomic instability and enhance tumour proliferation. Conclusions/Significance DFT cells appear to monitor and regulate the length of individual telomeres: i.e. shorter telomeres are elongated by up-regulation of telomerase-related genes; longer telomeres are protected from further elongation by members of the shelterin complex, which may explain the lack of spatial and strain variation in DFT telomere copy number. The observed longitudinal increase in gene expression in DFT tissue samples and telomerase activity in DFT cell lines might indicate a selection for more stable tumours with higher proliferative potential. PMID:22952882

Description of a new planktonic mixotrophic dinoflagellate Paragymnodinium shiwhaense n. gen., n. sp. from the coastal waters off Western Korea: morphology, pigments, and ribosomal DNA gene sequence.

PubMed

Kang, Nam Seon; Jeong, Hae Jin; Moestrup, Øjvind; Shin, Woongghi; Nam, Seung Won; Park, Jae Yeon; De Salas, Miguel F; Kim, Ki Woo; Noh, Jae Hoon

2010-01-01

The mixotrophic dinoflagellate Paragymnodinium shiwhaense n. gen., n. sp. is described from living cells and from cells prepared by light, scanning electron, and transmission electron microscopy. In addition, sequences of the small subunit (SSU) and large subunit (LSU) rDNA and photosynthetic pigments are reported. The episome is conical, while the hyposome is hemispherical. Cells are covered with polygonal amphiesmal vesicles arranged in 16 rows and containing a very thin plate-like component. There is neither an apical groove nor apical line of narrow plates. Instead, there is a sulcal extension-like furrow. The cingulum is as wide as 0.2-0.3 x cell length and displaced by 0.2-0.3 x cell length. Cell length and width of live cells fed Amphidinium carterae were 8.4-19.3 and 6.1-16.0 microm, respectively. Paragymnodinium shiwhaense does not have a nuclear envelope chamber nor a nuclear fibrous connective (NFC). Cells contain chloroplasts, nematocysts, trichocysts, and peduncle, though eyespots, pyrenoids, and pusules are absent. The main accessory pigment is peridinin. The sequence of the SSU rDNA of this dinoflagellate (GenBank AM408889) is 4% different from that of Gymnodinium aureolum, Lepidodinium viride, and Gymnodinium catenatum, the three closest species, while the LSU rDNA was 17-18% different from that of G. catenatum, Lepidodinium chlorophorum, and Gymnodinium nolleri. The phylogenetic trees show that this dinoflagellate belongs within the Gymnodinium sensu stricto clade. However, in contrast to Gymnodinium spp., cells lack nuclear envelope chambers, NFC, and an apical groove. Unlike Polykrikos spp., which have a taeniocyst-nematocyst complex, P. shiwhaense has nematocysts without taeniocysts. In addition, P. shiwhaense does not have ocelloids in contrast to Warnowia spp. and Nematodinium spp. Therefore, based on morphological and molecular analyses, we suggest that this taxon is a new species, also within a new genus.

Alpha-latrotoxin induces exocytosis by inhibition of voltage-dependent K+ channels and by stimulation of L-type Ca2+ channels via latrophilin in beta-cells.

PubMed

Lajus, Sophie; Vacher, Pierre; Huber, Denise; Dubois, Mathilde; Benassy, Marie-Noëlle; Ushkaryov, Yuri; Lang, Jochen

2006-03-03

The spider venom alpha-latrotoxin (alpha-LTX) induces massive exocytosis after binding to surface receptors, and its mechanism is not fully understood. We have investigated its action using toxin-sensitive MIN6 beta-cells, which express endogenously the alpha-LTX receptor latrophilin (LPH), and toxin-insensitive HIT-T15 beta-cells, which lack endogenous LPH. alpha-LTX evoked insulin exocytosis in HIT-T15 cells only upon expression of full-length LPH but not of LPH truncated after the first transmembrane domain (LPH-TD1). In HIT-T15 cells expressing full-length LPH and in native MIN6 cells, alpha-LTX first induced membrane depolarization by inhibition of repolarizing K(+) channels followed by the appearance of Ca(2+) transients. In a second phase, the toxin induced a large inward current and a prominent increase in intracellular calcium ([Ca(2+)](i)) reflecting pore formation. Upon expression of LPH-TD1 in HIT-T15 cells just this second phase was observed. Moreover, the mutated toxin LTX(N4C), which is devoid of pore formation, only evoked oscillations of membrane potential by reversible inhibition of iberiotoxin-sensitive K(+) channels via phospholipase C, activated L-type Ca(2+) channels independently from its effect on membrane potential, and induced an inositol 1,4,5-trisphosphate receptor-dependent release of intracellular calcium in MIN6 cells. The combined effects evoked transient increases in [Ca(2+)](i) in these cells, which were sensitive to inhibitors of phospholipase C, protein kinase C, or L-type Ca(2+) channels. The latter agents also reduced toxin-induced insulin exocytosis. In conclusion, alpha-LTX induces signaling distinct from pore formation via full-length LPH and phospholipase C to regulate physiologically important K(+) and Ca(2+) channels as novel targets of its secretory activity.

An in silico investigation into the causes of telomere length heterogeneity and its implications for the Hayflick limit.

PubMed

Golubev, A; Khrustalev, S; Butov, A

2003-11-21

In telomerase-negative cell populations the mean telomere length (TL) decreases with increasing population doubling number (PD). A critically small TL is believed to stop cell proliferation at a cell-, age- and species-specific PD thus defining the Hayflick limit. However, positively skewed TL distributions are broad compared to differences between initial and final mean TL and strongly overlap at middle and late PD, which is inconsistent with a limiting role of TL. We used computer-assisted modelling to define what set of premises may account for the above. Our model incorporates the following concepts. DNA end replication problem: telomeres loose 1 shortening unit (SU) upon each cell division. Free radical-caused TL decrease: telomeres experience random events resulting in the loss of a random SU number within a remaining TL. Stochasticity of gene expression and cell differentiation: cells experience random events inducing mitoses or committing cells to proliferation arrest, the latter option requiring a specified number of mitoses to be passed. Cells whose TL reaches 1SU cannot divide. The proliferation kinetics of such virtual cells conforms to the transition probability model of cell cycle. When no committing events occur and at realistic SU estimates of the initial TL, maximal PD values far exceed the Hayflick limit observed in normal cells and are consistent with the crisis stage entered by transformed cells that have surpassed the Hayflick limit. At intermediate PD, symmetrical TL distributions are yielded. Upon introduction of committing events making the ratio of the rates of proliferating and committing events (P/C) range from 1.10 to 1.25, TL distributions at intermediate PD become positively skewed, and virtual cell clones show bimodal size distributions. At P/C as high as 1.25 the majority of virtual cells at maximal PD contain telomeres with TL>1SU. A 10% increase in P/C within the 1.10-1.25 range produces a two-fold increase in the maximal PD, which can reach values of up to 25 observed in rodent and some human cells. Increasing the number of committed mitoses from 0 to 10 can increases PD to about 50 observed in human fibroblasts. Introduction of the random TL breakage makes the shapes of TL distributions quite dissimilar from those observed in real cells. Telomere length decrease is a correlate of cell proliferation that cannot alone account for the Hayflick limit, which primarily depends on parameters of cell population kinetics. Free radical damage influences the Hayflick limit not through TL but rather by affecting the ratio of the rates of events that commit cells to mitoses or to proliferation arrest.

Different Effects of sgRNA Length on CRISPR-mediated Gene Knockout Efficiency.

PubMed

Zhang, Jian-Ping; Li, Xiao-Lan; Neises, Amanda; Chen, Wanqiu; Hu, Lin-Ping; Ji, Guang-Zhen; Yu, Jun-Yao; Xu, Jing; Yuan, Wei-Ping; Cheng, Tao; Zhang, Xiao-Bing

2016-06-24

CRISPR-Cas9 is a powerful genome editing technology, yet with off-target effects. Truncated sgRNAs (17nt) have been found to decrease off-target cleavage without affecting on-target disruption in 293T cells. However, the potency of 17nt sgRNAs relative to the full-length 20nt sgRNAs in stem cells, such as human mesenchymal stem cells (MSCs) and induced pluripotent stem cells (iPSCs), has not been assessed. Using a GFP reporter system, we found that both 17nt and 20nt sgRNAs expressed by lentiviral vectors induce ~95% knockout (KO) in 293T cells, whereas the KO efficiencies are significantly lower in iPSCs (60-70%) and MSCs (65-75%). Furthermore, we observed a decrease of 10-20 percentage points in KO efficiency with 17nt sgRNAs compared to full-length sgRNAs in both iPSCs and MSCs. Off-target cleavage was observed in 17nt sgRNAs with 1-2nt but not 3-4nt mismatches; whereas 20nt sgRNAs with up to 5nt mismatches can still induce off-target mutations. Of interest, we occasionally observed off-target effects induced by the 17nt but not the 20nt sgRNAs. These results indicate the importance of balancing on-target gene cleavage potency with off-target effects: when efficacy is a major concern such as genome editing in stem cells, the use of 20nt sgRNAs is preferable.

Antagonistic Actions of HLH/bHLH Proteins Are Involved in Grain Length and Weight in Rice

PubMed Central

Heang, Dany; Sassa, Hidenori

2012-01-01

Grain size is a major yield component in rice, and partly controlled by the sizes of the lemma and palea. Molecular mechanisms controlling the sizes of these organs largely remain unknown. In this study, we show that an antagonistic pair of basic helix-loop-helix (bHLH) proteins is involved in determining rice grain length by controlling cell length in the lemma/palea. Overexpression of an atypical bHLH, named POSITIVE REGULATOR OF GRAIN LENGTH 1 (PGL1), in lemma/palea increased grain length and weight in transgenic rice. PGL1 is an atypical non-DNA-binding bHLH and assumed to function as an inhibitor of a typical DNA-binding bHLH through heterodimerization. We identified the interaction partner of PGL1 and named it ANTAGONIST OF PGL1 (APG). PGL1 and APG interacted in vivo and localized in the nucleus. As expected, silencing of APG produced the same phenotype as overexpression of PGL1, suggesting antagonistic roles for the two genes. Transcription of two known grain-length-related genes, GS3 and SRS3, was largely unaffected in the PGL1-overexpressing and APG-silenced plants. Observation of the inner epidermal cells of lemma revealed that are caused by increased cell length. PGL1-APG represents a new grain length and weight-controlling pathway in which APG is a negative regulator whose function is inhibited by PGL1. PMID:22363621

Architectural protein subclasses shape 3-D organization of genomes during lineage commitment

PubMed Central

Phillips-Cremins, Jennifer E.; Sauria, Michael E. G.; Sanyal, Amartya; Gerasimova, Tatiana I.; Lajoie, Bryan R.; Bell, Joshua S. K.; Ong, Chin-Tong; Hookway, Tracy A.; Guo, Changying; Sun, Yuhua; Bland, Michael J.; Wagstaff, William; Dalton, Stephen; McDevitt, Todd C.; Sen, Ranjan; Dekker, Job; Taylor, James; Corces, Victor G.

2013-01-01

Summary Understanding the topological configurations of chromatin may reveal valuable insights into how the genome and epigenome act in concert to control cell fate during development. Here we generate high-resolution architecture maps across seven genomic loci in embryonic stem cells and neural progenitor cells. We observe a hierarchy of 3-D interactions that undergo marked reorganization at the sub-Mb scale during differentiation. Distinct combinations of CTCF, Mediator, and cohesin show widespread enrichment in looping interactions at different length scales. CTCF/cohesin anchor long-range constitutive interactions that form the topological basis for invariant sub-domains. Conversely, Mediator/cohesin together with pioneer factors bridge shortrange enhancer-promoter interactions within and between larger sub-domains. Knockdown of Smc1 or Med12 in ES cells results in disruption of spatial architecture and down-regulation of genes found in cohesin-mediated interactions. We conclude that cell type-specific chromatin organization occurs at the sub-Mb scale and that architectural proteins shape the genome in hierarchical length scales. PMID:23706625

Multi-Material Front Contact for 19% Thin Film Solar Cells.

PubMed

van Deelen, Joop; Tezsevin, Yasemin; Barink, Marco

2016-02-06

The trade-off between transmittance and conductivity of the front contact material poses a bottleneck for thin film solar panels. Normally, the front contact material is a metal oxide and the optimal cell configuration and panel efficiency were determined for various band gap materials, representing Cu(In,Ga)Se₂ (CIGS), CdTe and high band gap perovskites. Supplementing the metal oxide with a metallic copper grid improves the performance of the front contact and aims to increase the efficiency. Various front contact designs with and without a metallic finger grid were calculated with a variation of the transparent conductive oxide (TCO) sheet resistance, scribing area, cell length, and finger dimensions. In addition, the contact resistance and illumination power were also assessed and the optimal thin film solar panel design was determined. Adding a metallic finger grid on a TCO gives a higher solar cell efficiency and this also enables longer cell lengths. However, contact resistance between the metal and the TCO material can reduce the efficiency benefit somewhat.

Computational Fluid Dynamics-Population Balance Model Simulation of Effects of Cell Design and Operating Parameters on Gas-Liquid Two-Phase Flows and Bubble Distribution Characteristics in Aluminum Electrolysis Cells

NASA Astrophysics Data System (ADS)

Zhan, Shuiqing; Wang, Junfeng; Wang, Zhentao; Yang, Jianhong

2018-02-01

The effects of different cell design and operating parameters on the gas-liquid two-phase flows and bubble distribution characteristics under the anode bottom regions in aluminum electrolysis cells were analyzed using a three-dimensional computational fluid dynamics-population balance model. These parameters include inter-anode channel width, anode-cathode distance (ACD), anode width and length, current density, and electrolyte depth. The simulations results show that the inter-anode channel width has no significant effect on the gas volume fraction, electrolyte velocity, and bubble size. With increasing ACD, the above values decrease and more uniform bubbles can be obtained. Different effects of the anode width and length can be concluded in different cell regions. With increasing current density, the gas volume fraction and electrolyte velocity increase, but the bubble size keeps nearly the same. Increasing electrolyte depth decreased the gas volume fraction and bubble size in particular areas and the electrolyte velocity increased.

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

Liu Xiaohong; Zhang Shuhui; Lin Jing

The role of the hepatitis B virus X protein (HBx) in hepatocarcinogenesis remains controversial. To investigate the biological impact of hepatitis B virus x gene (HBx) mutation on hepatoma cells, plasmids expressing the full-length HBx or HBx deletion mutants were constructed. The biological activities in these transfectants were analyzed by a series of assays. Results showed that HBx3'-20 and HBx3'-40 amino acid deletion mutants exhibited an increase in cellular proliferation, focus formation, tumorigenicity, and invasive growth and metastasis through promotion of the cell cycle from G0/G1 to the S phase, when compared with the full-length HBx. In contrast, HBx3'-30 aminomore » acid deletion mutant repressed cell proliferation by blocking in G1 phase. The expression of P53, p21{sup WAF1}, p14{sup ARF}, and MDM2 proteins was regulated by expression of HBx mutants. In conclusions, HBx variants showed different effects and functions on cell proliferation and invasion by regulation of the cell cycle progression and its associated proteins expression.« less

Length dependence of force generation exhibit similarities between rat cardiac myocytes and skeletal muscle fibres.

PubMed

Hanft, Laurin M; McDonald, Kerry S

2010-08-01

According to the Frank-Starling relationship, increased ventricular volume increases cardiac output, which helps match cardiac output to peripheral circulatory demand. The cellular basis for this relationship is in large part the myofilament length-tension relationship. Length-tension relationships in maximally calcium activated preparations are relatively shallow and similar between cardiac myocytes and skeletal muscle fibres. During twitch activations length-tension relationships become steeper in both cardiac and skeletal muscle; however, it remains unclear whether length dependence of tension differs between striated muscle cell types during submaximal activations. The purpose of this study was to compare sarcomere length-tension relationships and the sarcomere length dependence of force development between rat skinned left ventricular cardiac myocytes and fast-twitch and slow-twitch skeletal muscle fibres. Muscle cell preparations were calcium activated to yield 50% maximal force, after which isometric force and rate constants (k(tr)) of force development were measured over a range of sarcomere lengths. Myofilament length-tension relationships were considerably steeper in fast-twitch fibres compared to slow-twitch fibres. Interestingly, cardiac myocyte preparations exhibited two populations of length-tension relationships, one steeper than fast-twitch fibres and the other similar to slow-twitch fibres. Moreover, myocytes with shallow length-tension relationships were converted to steeper length-tension relationships by protein kinase A (PKA)-induced myofilament phosphorylation. Sarcomere length-k(tr) relationships were distinct between all three cell types and exhibited patterns markedly different from Ca(2+) activation-dependent k(tr) relationships. Overall, these findings indicate cardiac myocytes exhibit varied length-tension relationships and sarcomere length appears a dominant modulator of force development rates. Importantly, cardiac myocyte length-tension relationships appear able to switch between slow-twitch-like and fast-twitch-like by PKA-mediated myofibrillar phosphorylation, which implicates a novel means for controlling Frank-Starling relationships.

Effects of CASP5 gene overexpression on angiogenesis of HMEC-1 cells.

PubMed

Li, Haiyan; Li, Yuzhen; Cai, Limin; Bai, Bingxue; Wang, Yanhua

2015-01-01

The efficacy of gene overexpression of CASP5, a caspase family member, in angiogenesis in vitro and its mechanisms were clarified. Human full-length CASP5 gene was delivered into human microvascular endothelial HMEC-1 cells by recombinant lentivirus. The infection was estimated by green fluorescent protein. MTT method was used to analyze the efficacy of gene overexpression in cell proliferation ability, and Matrigel was used to estimate its effects in angiogenesis ability of cells. Meanwhile, Western blot was used to analyze the effects of CASP5 gene overexpression on the expression levels of angpt-1, angpt-2, Tie2 and VEGF-1 in the cells, which were signaling pathway factors related to angiogenesis. Recombinant lentivirus containing human full-length CASP5 gene was packed and purified successfully, with virus titer of 1×10(8) TU/ml. The recombinant lentivirus was used to infect HMEC-1 cells with MOI of 1, leading to a cell infection rate of 100%. There were no significant effects of CASP5 gene overexpression on both cell proliferation ability and the expression level of angpt-1. Meanwhile, expressions of angpt-2 and VEGF-1 were both enhanced, while Tie2 expression was inhibited. Results indicated that CASP5 gene overexpression promoted angiogenesis of HMEC-1 cells. CASP5 gene overexpression significantly promoted angiogenesis ability of HMEC-1 cells, which was probably achieved by inhibiting angpt-1/Tie2 and promoting VEGF-1 signal pathway.

Microarray analysis of human milk cells: persistent high expression of osteopontin during the lactation period

PubMed Central

NAGATOMO, T; OHGA, S; TAKADA, H; NOMURA, A; HIKINO, S; IMURA, M; OHSHIMA, K; HARA, T

2004-01-01

To continue the search for immunological roles of breast milk, cDNA microarray analysis on cytokines and growth factors was performed for human milk cells. Among the 240 cytokine-related genes, osteopontin (OPN) gene ranked top of the expression. Real-time PCR revealed that the OPN mRNA levels in colostrum cells were approximately 100 times higher than those in PHA-stimulated peripheral blood mononuclear cells (PBMNCs), and 10 000 times higher than those in PB CD14+ cells. The median levels of OPN mRNA in early milk or mature milk cells were more than three times higher than those in colostrum cells. Western blot analysis of human milk showed appreciable expression of full-length and short form proteins of OPN. The concentrations of full-length OPN in early milk or mature milk whey continued to be higher than those in colostrum whey and plasma as assessed by ELISA. The early milk (3–7 days postpartum) contained the highest concentrations of OPN protein, while the late mature milk cells (1 years postpartum) had the highest expression of OPN mRNA of all the lactating periods. The results of immunohistochemical and immunocytochemical staining indicated that OPN-producing epithelial cells and macrophages are found in actively lactating mammary glands. These results suggest that the persistently and extraordinarily high expression of OPN in human milk cells plays a potential role in the immunological development of breast-fed infants. PMID:15373904

Biomimetic surface patterning for long-term transmembrane access

PubMed Central

VanDersarl, Jules J.; Renaud, Philippe

2016-01-01

Here we present a planar patch clamp chip based on biomimetic cell membrane fusion. This architecture uses nanometer length-scale surface patterning to replicate the structure and function of membrane proteins, creating a gigaohm seal between the cell and a planar electrode array. The seal is generated passively during cell spreading, without the application of a vacuum to the cell surface. This interface can enable cell-attached and whole-cell recordings that are stable to 72 hours, and generates no visible damage to the cell. The electrodes can be very small (<5 μm) and closely packed, offering a high density platform for cellular measurement. PMID:27577519

Biomimetic surface patterning for long-term transmembrane access.

PubMed

VanDersarl, Jules J; Renaud, Philippe

2016-08-31

Here we present a planar patch clamp chip based on biomimetic cell membrane fusion. This architecture uses nanometer length-scale surface patterning to replicate the structure and function of membrane proteins, creating a gigaohm seal between the cell and a planar electrode array. The seal is generated passively during cell spreading, without the application of a vacuum to the cell surface. This interface can enable cell-attached and whole-cell recordings that are stable to 72 hours, and generates no visible damage to the cell. The electrodes can be very small (<5 μm) and closely packed, offering a high density platform for cellular measurement.

Leukocyte telomere length in relation to risk of lung adenocarcinoma incidence: Findings from the Singapore Chinese Health Study.

PubMed

Yuan, Jian-Min; Beckman, Kenneth B; Wang, Renwei; Bull, Caroline; Adams-Haduch, Jennifer; Huang, Joyce Y; Jin, Aizhen; Opresko, Patricia; Newman, Anne B; Zheng, Yun-Ling; Fenech, Michael; Koh, Woon-Puay

2018-06-01

Telomeres are crucial in the maintenance of chromosome integrity and genomic stability. Critically short telomeres can trigger programed cell death while cells with longer telomeres may have increased likelihood of replicative errors, resulting in genetic mutations and chromosomal alterations, and ultimately promoting oncogenesis. Data on telomere length and lung cancer risk from large prospective cohort studies are spare. Relative telomere length in peripheral blood leukocytes was quantified using a validated monochrome multiplex quantitative polymerase chain reaction (qPCR) method in 26,540 participants of the Singapore Chinese Health Study. After a follow-up of 12 years, 654 participants developed lung cancer including 288 adenocarcinoma, 113 squamous cell carcinoma and 253 other/unknown histological type. The Cox proportional hazard regression was used to estimate hazard ratio (HR) and 95% confidence interval (CI). HR of lung adenocarcinoma for individuals in the highest comparing the lowest 20 percentile of telomere length was 2.84 (95% CI 1.94-4.14, p trend  < 0.0001). This positive association was present in never smokers (p trend  < 0.0001), ever smokers (p trend  = 0.0010), men (p trend  = 0.0003), women (p trend  < 0.0001), and in shorter (p trend  = 0.0002) and longer (p trend  = 0.0001) duration of follow-up. There was no association between telomere length and risk of squamous cell carcinoma or other histological type of lung cancer in all or subgroups of individuals. The agreement of results from this prospective cohort study with those of previous prospective studies and Mendelian randomization studies suggest a possible etiological role of telomere length in the development of lung adenocarcinoma. © 2018 UICC.

A novel T cell receptor single-chain signaling complex mediates antigen-specific T cell activity and tumor control

PubMed Central

Stone, Jennifer D.; Harris, Daniel T.; Soto, Carolina M.; Chervin, Adam S.; Aggen, David H.; Roy, Edward J.; Kranz, David M.

2014-01-01

Adoptive transfer of genetically modified T cells to treat cancer has shown promise in several clinical trials. Two main strategies have been applied to redirect T cells against cancer: 1) introduction of a full-length T cell receptor (TCR) specific for a tumor-associated peptide-MHC, or 2) introduction of a chimeric antigen receptor (CAR), including an antibody fragment specific for a tumor cell surface antigen, linked intracellularly to T cell signaling domains. Each strategy has advantages and disadvantages for clinical applications. Here, we present data on the in vitro and in vivo effectiveness of a single-chain signaling receptor incorporating a TCR variable fragment as the targeting element (referred to as TCR-SCS). This receptor contained a single-chain TCR (Vβ-linker-Vα) from a high-affinity TCR called m33, linked to the intracellular signaling domains of CD28 and CD3ζ. This format avoided mispairing with endogenous TCR chains, and mediated specific T cell activity when expressed in either CD4 or CD8 T cells. TCR-SCS-transduced CD8-negative cells showed an intriguing sensitivity, compared to full-length TCRs, to higher densities of less stable pepMHC targets. T cells that expressed this peptide-specific receptor persisted in vivo, and exhibited polyfunctional responses. Growth of metastatic antigen-positive tumors was significantly inhibited by T cells that expressed this receptor, and tumor cells that escaped were antigen loss variants. TCR-SCS receptors represent an alternative targeting receptor strategy that combines the advantages of single-chain expression, avoidance of TCR chain mispairing, and targeting of intracellular antigens presented in complex with MHC proteins. PMID:25082071

In Situ Spatiotemporal Mapping of Flow Fields around Seeded Stem Cells at the Subcellular Length Scale

PubMed Central

Song, Min Jae; Dean, David; Knothe Tate, Melissa L.

2010-01-01

A major hurdle to understanding and exploiting interactions between the stem cell and its environment is the lack of a tool for precise delivery of mechanical cues concomitant to observing sub-cellular adaptation of structure. These studies demonstrate the use of microscale particle image velocimetry (μ-PIV) for in situ spatiotemporal mapping of flow fields around mesenchymal stem cells, i.e. murine embryonic multipotent cell line C3H10T1/2, at the subcellular length scale, providing a tool for real time observation and analysis of stem cell adaptation to the prevailing mechanical milieu. In the absence of cells, computational fluid dynamics (CFD) predicts flow regimes within 12% of μ-PIV measures, achieving the technical specifications of the chamber and the flow rates necessary to deliver target shear stresses at a particular height from the base of the flow chamber. However, our μ-PIV studies show that the presence of cells per se as well as the density at which cells are seeded significantly influences local flow fields. Furthermore, for any given cell or cell seeding density, flow regimes vary significantly along the vertical profile of the cell. Hence, the mechanical milieu of the stem cell exposed to shape changing shear stresses, induced by fluid drag, varies with respect to proximity of surrounding cells as well as with respect to apical height. The current study addresses a previously unmet need to predict and observe both flow regimes as well as mechanoadaptation of cells in flow chambers designed to deliver precisely controlled mechanical signals to live cells. An understanding of interactions and adaptation in response to forces at the interface between the surface of the cell and its immediate local environment may be key for de novo engineering of functional tissues from stem cell templates as well as for unraveling the mechanisms underlying multiscale development, growth and adaptation of organisms. PMID:20862249

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule.

PubMed

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-10-29

Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding alphavbeta5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain.

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule

PubMed Central

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-01-01

Background Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. Results L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding αvβ5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Conclusion Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain. PMID:19874583

Loss of ACTH expression in cultured human corticotroph macroadenoma cells is consistent with loss of the POMC gene signal sequence.

PubMed

Rees, D A; Hepburn, P J; McNicol, A M; Francis, K; Jasani, B; Lewis, M D; Farrell, W E; Lewis, B M; Scanlon, M F; Ham, J

2002-03-28

The proopiomelanocortin (POMC) gene is highly expressed in the pituitary gland where the resulting mRNA of 1200 base pairs (bp) gives rise to a full-length protein sequence. In peripheral tissues however both shorter and longer POMC variants have been described, these include for example placental tissue which contain 800 (truncated at the 5' end) and 1500 as well as the 1200 bp transcripts. The importance of the 800 bp transcript is unclear as the lack of a signal sequence renders the molecule to be non-functional. This transcript has not been previously demonstrated in the pituitary gland. In this report we show evidence of a 5' truncated POMC gene in human pituitary corticotroph macroadenoma cells (JE) maintained in primary culture for >1 year. The original tumour tissue and the derived cells during early passage (up to passage 4-5) immunostained for ACTH and in situ hybridisation confirmed the presence of the POMC gene in the cultured cells. These cells also secreted 15-40 pg/10(5) cells/24 h ACTH. In addition, as expected RT-PCR demonstrated the presence of all three POMC gene exons and is thus indicative of a full-length POMC gene. In late culture passages (passages 8-15) JE cells ceased to express ACTH and cell growth became very slow due presumably to cells reaching their Hayflick limit. ACTH immunostaining in these cells was undetectable and ACTH secretion was also at the detection limits of the assay and no greater than 10 pg/10(5) cells/24 h. ACTH precursor molecules were also undetectable. RT-PCR for the POMC gene in these late passage cells showed that only exon 3 was detectable, in contrast to early passage cells where all three exons were present. In summary we isolated in culture, human pituitary cells that possessed initially all three exons of the POMC gene and immunostained for ACTH. On further passaging these cells showed a loss of exons 1 and 2 in the POMC gene and a loss of ACTH immunostaining and secretion. We would like to suggest that the loss of ACTH peptide expression in these late passage cells is in part due to the loss of the POMC signal sequence. An alternative explanation for our findings is that there were originally two populations of corticotrophs in the cultures, one of which possessed the full-length POMC gene and the other only the 5' truncated POMC transcript and it is these latter cells which survived in culture. In either scenario this is the first report of the 5' truncated POMC gene occurring in pituitary cells.

Effect of vitamin E administration on the elevated oxygen stress and the telomeric and subtelomeric status in Alzheimer's disease.

PubMed

Guan, Jing-Zhi; Guan, Wei-Ping; Maeda, Toyoki; Makino, Naoki

2012-01-01

Oxidative stress (OS) may be involved in the neurodegenerative process in Alzheimer's disease (AD). Telomeres, the repeated sequences that cap chromosome ends, undergo shortening with each cell division, are sensitive to OS, and serve as markers of a cell's replicative history. Telomere length shortening has been reported to relate to OS with aging process and aging-associated diseases, but the telomeric changes were not always identical, especially in change of telomere length distribution and subtelomeric methylation. The involvement of an OS-associated telomere change in the pathogenesis of AD has been discussed for decades, and the telomere length and telomerase activity were analyzed. However, other telomeric factors, such as the telomere distribution and subtelomeric methylation status, have not yet been analyzed. The subtelomeric methylation status as well as the telomere length were studied in AD with an antioxidant vitamin in terms of OS. We measured urinary 8-iso-PGF2α, a lipid-peroxidation product as an OS marker, and methylated and non-methylated telomere lengths in the peripheral blood mononuclear cells by Southern blotting in AD patients before and after vitamin E treatment. The level of urinary 8-iso-PGF2α was found to have increased in AD. Middle-ranged telomeres (4.4-9.4 kb) increased and the shortest telomeres (<4.4 kb) decreased in AD patients. Telomeres were more methylated in both long telomeres and in short telomeres in AD compared with the control. The oral administration of the antioxidant vitamin E in 400 mg/day for 6 months in AD patients partly reversed AD-associated alterations in OS marker levels. AD patients showed an elevated OS marker level, and vitamin E lowered the OS level. In comparison with controls, AD patients showed shorter telomere lengths. Cells with short and long telomeres bore relatively hypermethylated subtelomeres in AD patients. Aging-associated accumulation of cells bearing short telomeres was not observed in AD. These results imply that long telomeres with hypomethylation tend to shorten faster, and cells bearing short telomeres with hypomethylation tend to more easily enter into a senescent state under elevated OS stress in AD. However, no significant effect on the altered telomeric profiles in AD patients could be detected after a 6-month administration of vitamin E. Copyright © 2011 S. Karger AG, Basel.

Introduction to basic solar cell measurements

NASA Technical Reports Server (NTRS)

Brandhorst, H. W., Jr.

1976-01-01

The basic approaches to solar cell performance and diagnostic measurements are described. The light sources, equipment for I-V curve measurement, and the test conditions and procedures for performance measurement are detailed. Solar cell diagnostic tools discussed include analysis of I-V curves, series resistance and reverse saturation current determination, spectral response/quantum yield measurement, and diffusion length/lifetime determination.

Fabrication and photovoltaic properties of ZnO nanorods/perovskite solar cells

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

Shirahata, Yasuhiro; Tanaike, Kohei; Akiyama, Tsuyoshi

2016-02-01

ZnO nanorods/perovskite solar cells with different lengths of ZnO nanorods were fabricated. The ZnO nanorods were prepared by chemical bath deposition and directly confirmed to be hexagon-shaped nanorods. The lengths of the ZnO nanorads were controlled by deposition condition of ZnO seed layer. Photovoltaic properties of the ZnO nanorods/CH{sub 3}NH{sub 3}PbI{sub 3} solar cells were investigated by measuring current density-voltage characteristics and incident photon to current conversion efficiency. The highest conversion efficiency was obtained in ZnO nanorods/CH{sub 3}NH{sub 3}PbI{sub 3} with the longest ZnO nanorods.

Application of the SEM to the measurement of solar cell parameters

NASA Technical Reports Server (NTRS)

Weizer, V. G.; Andrews, C. W.

1977-01-01

Techniques are described which make use of the SEM to measure the minority carrier diffusion length and the metallurgical junction depth in silicon solar cells. The former technique permits the measurement of the true bulk diffusion length through the application of highly doped field layers to the back surfaces of the cells being investigated. It is shown that the secondary emission contrast observed in the SEM on a reverse-biased diode can depict the location of the metallurgical junction if the diode has been prepared with the proper beveled geometry. The SEM provides the required contrast and the option of high magnification, permitting the measurement of extremely shallow junction depths.

Lyme disease and relapsing fever Borrelia elongate through zones of peptidoglycan synthesis that mark division sites of daughter cells.

PubMed

Jutras, Brandon Lyon; Scott, Molly; Parry, Bradley; Biboy, Jacob; Gray, Joe; Vollmer, Waldemar; Jacobs-Wagner, Christine

2016-08-16

Agents that cause Lyme disease, relapsing fever, leptospirosis, and syphilis belong to the phylum Spirochaetae-a unique lineage of bacteria most known for their long, spiral morphology. Despite the relevance to human health, little is known about the most fundamental aspects of spirochete growth. Here, using quantitative microscopy to track peptidoglycan cell-wall synthesis, we found that the Lyme disease spirochete Borrelia burgdorferi displays a complex pattern of growth. B. burgdorferi elongates from discrete zones that are both spatially and temporally regulated. In addition, some peptidoglycan incorporation occurs along the cell body, with the notable exception of a large region at the poles. Newborn cells inherit a highly active zone of peptidoglycan synthesis at midcell that contributes to elongation for most of the cell cycle. Concomitant with the initiation of nucleoid separation and cell constriction, second and third zones of elongation are established at the 1/4 and 3/4 cellular positions, marking future sites of division for the subsequent generation. Positioning of elongation zones along the cell is robust to cell length variations and is relatively precise over long distances (>30 µm), suggesting that cells ‟sense" relative, as opposed to absolute, cell length to establish zones of peptidoglycan synthesis. The transition from one to three zones of peptidoglycan growth during the cell cycle is also observed in relapsing fever Borrelia. However, this mode of growth does not extend to representative species from other spirochetal genera, suggesting that this distinctive growth mode represents an evolutionary divide in the spirochete phylum.

Radiosensitization of head/neck sqaumous cell carcinoma by adenovirus-mediated expression of the Nbs1 protein

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

Rhee, Juong G.; Marlene and Stewart Greenebaum Cancer Center, University of Maryland School of Medicine, Baltimore, MD; Li, Daqing

2007-01-01

Purpose: Local failure and toxicity to adjacent critical structures is a significant problem in radiation therapy of cancers of the head and neck. We are developing a gene therapy based method of sensitizing head/neck squamous cell carcinoma (HNSCC) to radiation treatment. As patients with the rare hereditary disorder, Nijmegen breakage syndrome, show radiation sensitivity we hypothesized that tumor-specific disruption of the function of the Nbs1 protein would lead to enhanced cellular sensitivity to ionizing radiation. Experimental Procedures: We constructed two recombinant adenoviruses by cloning the full-length Nbs1 cDNA as well as the C-terminal 300 amino acids of Nbs1 into anmore » adenovirus backbone under the control of a CMV promoter. The resulting adenoviruses were used to infect HNSCC cell line JHU011. These cells were evaluated for expression of the viral based constructs and assayed for clonogenic survival following radiation exposure. Results: Exposure of cells expressing Nbs1-300 to ionizing radiation resulted in a small reduction in survival relative to cells infected with control virus. Surprisingly, expression of full-length Nbs1 protein resulted in markedly enhanced sensitivity to ionizing radiation. Furthermore, the use of a fractionated radiation scheme following virus infection demonstrates that expression of full-length Nbs1 protein results in significant reduction in cell survival. Conclusions: These results provide a proof of principle that disruption of Nbs1 function may provide a means of enhancing the radiosensitivity of head and neck tumors. Additionally, this work highlights the Mre11 complex as an attractive target for development of radiation sensitizers.« less

Treatment of Tourniquet-Induced Ischemia Reperfusion Injury with Muscle Progenitor Cells

DTIC Science & Technology

2011-09-01

application. Muscle mass, isometric contractile properties, and selected histologic properties were evaluated at 2 wk after ischemia. Results. IRI...results showed that a small number of trans- planted cells differentiated and formed muscle fibers , which could potentially contribute to force genera...the wet weight of the muscle (in g); q is the angle of fiber pinnation (12.8 for TA); Lf is the mean fiber length (57% of TAmuscle length); and r is

Adaptive treatment-length optimization in spatiobiologically integrated radiotherapy

NASA Astrophysics Data System (ADS)

Ajdari, Ali; Ghate, Archis; Kim, Minsun

2018-04-01

Recent theoretical research on spatiobiologically integrated radiotherapy has focused on optimization models that adapt fluence-maps to the evolution of tumor state, for example, cell densities, as observed in quantitative functional images acquired over the treatment course. We propose an optimization model that adapts the length of the treatment course as well as the fluence-maps to such imaged tumor state. Specifically, after observing the tumor cell densities at the beginning of a session, the treatment planner solves a group of convex optimization problems to determine an optimal number of remaining treatment sessions, and a corresponding optimal fluence-map for each of these sessions. The objective is to minimize the total number of tumor cells remaining (TNTCR) at the end of this proposed treatment course, subject to upper limits on the biologically effective dose delivered to the organs-at-risk. This fluence-map is administered in future sessions until the next image is available, and then the number of sessions and the fluence-map are re-optimized based on the latest cell density information. We demonstrate via computer simulations on five head-and-neck test cases that such adaptive treatment-length and fluence-map planning reduces the TNTCR and increases the biological effect on the tumor while employing shorter treatment courses, as compared to only adapting fluence-maps and using a pre-determined treatment course length based on one-size-fits-all guidelines.

Analysis of the Growth Process of Neural Cells in Culture Environment Using Image Processing Techniques

NASA Astrophysics Data System (ADS)

Mirsafianf, Atefeh S.; Isfahani, Shirin N.; Kasaei, Shohreh; Mobasheri, Hamid

Here we present an approach for processing neural cells images to analyze their growth process in culture environment. We have applied several image processing techniques for: 1- Environmental noise reduction, 2- Neural cells segmentation, 3- Neural cells classification based on their dendrites' growth conditions, and 4- neurons' features Extraction and measurement (e.g., like cell body area, number of dendrites, axon's length, and so on). Due to the large amount of noise in the images, we have used feed forward artificial neural networks to detect edges more precisely.

Heterogeneity of the Axon Initial Segment in Interneurons and Pyramidal Cells of Rodent Visual Cortex

PubMed Central

Höfflin, Felix; Jack, Alexander; Riedel, Christian; Mack-Bucher, Julia; Roos, Johannes; Corcelli, Corinna; Schultz, Christian; Wahle, Petra; Engelhardt, Maren

2017-01-01

The microdomain that orchestrates action potential initiation in neurons is the axon initial segment (AIS). It has long been considered to be a rather homogeneous domain at the very proximal axon hillock with relatively stable length, particularly in cortical pyramidal cells. However, studies in other brain regions paint a different picture. In hippocampal CA1, up to 50% of axons emerge from basal dendrites. Further, in about 30% of thick-tufted layer V pyramidal neurons in rat somatosensory cortex, axons have a dendritic origin. Consequently, the AIS is separated from the soma. Recent in vitro and in vivo studies have shown that cellular excitability is a function of AIS length/position and somatodendritic morphology, undermining a potentially significant impact of AIS heterogeneity for neuronal function. We therefore investigated neocortical axon morphology and AIS composition, hypothesizing that the initial observation of seemingly homogeneous AIS is inadequate and needs to take into account neuronal cell types. Here, we biolistically transfected cortical neurons in organotypic cultures to visualize the entire neuron and classify cell types in combination with immunolabeling against AIS markers. Using confocal microscopy and morphometric analysis, we investigated axon origin, AIS position, length, diameter as well as distance to the soma. We find a substantial AIS heterogeneity in visual cortical neurons, classified into three groups: (I) axons with somatic origin with proximal AIS at the axon hillock; (II) axons with somatic origin with distal AIS, with a discernible gap between the AIS and the soma; and (III) axons with dendritic origin (axon-carrying dendrite cell, AcD cell) and an AIS either starting directly at the axon origin or more distal to that point. Pyramidal cells have significantly longer AIS than interneurons. Interneurons with vertical columnar axonal projections have significantly more distal AIS locations than all other cells with their prevailing phenotype as an AcD cell. In contrast, neurons with perisomatic terminations display most often an axon originating from the soma. Our data contribute to the emerging understanding that AIS morphology is highly variable, and potentially a function of the cell type. PMID:29170630

Simultaneous determination of apparent tortuosity and microstructure length scale and shape: Application to rigid open cell foams

NASA Astrophysics Data System (ADS)

Gómez Álvarez-Arenas, T. E.; de la Fuente, S.; González Gómez, I.

2006-05-01

A novel experimental technique based on phase spectroscopy and through transmission of high-frequency airborne ultrasonic pulses is used to study rigid open cell foams. Phase velocity shows an anomalous relaxation like behavior which is attributed to a frequency variation of the apparent tortuosity. An explanation is proposed in terms of the relationship between the different length scales involved: microstructure and macroscopic behavior. The experimental technique together with the proposed apparent tortuosity scheme provides a novel and unique procedure to determine simultaneously tortuosity and characteristic length dimension and shape of the solid constituent of foams and porous materials in general.

Donor Telomere Length SAA

Cancer.gov

A new NCI study has found that, among patients with severe aplastic anemia who received a hematopoietic cell transplant from an unrelated donor, those whose donor white blood cells had longer telomeres had higher survival rates five-years after transplant

Atg7-Mediated Autophagy Is Involved in the Neural Crest Cell Generation in Chick Embryo.

PubMed

Wang, Guang; Chen, En-Ni; Liang, Chang; Liang, Jianxin; Gao, Lin-Rui; Chuai, Manli; Münsterberg, Andrea; Bao, Yongping; Cao, Liu; Yang, Xuesong

2018-04-01

Autophagy plays a very important role in numerous physiological and pathological events. However, it still remains unclear whether Atg7-induced autophagy is involved in the regulation of neural crest cell production. In this study, we found the co-location of Atg7 and Pax7 + neural crest cells in early chick embryo development. Upregulation of Atg7 with unilateral transfection of full-length Atg7 increased Pax7 + and HNK-1 + cephalic and trunk neural crest cell numbers compared to either Control-GFP transfection or opposite neural tubes, suggesting that Atg7 over-expression in neural tubes could enhance the production of neural crest cells. BMP4 in situ hybridization and p-Smad1/5/8 immunofluorescent staining demonstrated that upregulation of Atg7 in neural tubes suppressed the BMP4/Smad signaling, which is considered to promote the delamination of neural crest cells. Interestingly, upregulation of Atg7 in neural tubes could significantly accelerate cell progression into the S phase, implying that Atg7 modulates cell cycle progression. However, β-catenin expression was not significantly altered. Finally, we demonstrated that upregulation of the Atg7 gene could activate autophagy as did Atg8. We have also observed that similar phenotypes, such as more HNK-1 + neural crest cells in the unilateral Atg8 transfection side of neural tubes, and the transfection with full-length Atg8-GFP certainly promote the numbers of BrdU + neural crest cells in comparison to the GFP control. Taken together, we reveal that Atg7-induced autophagy is involved in regulating the production of neural crest cells in early chick embryos through the modification of the cell cycle.

Where hearing starts: the development of the mammalian cochlea.

PubMed

Basch, Martin L; Brown, Rogers M; Jen, Hsin-I; Groves, Andrew K

2016-02-01

The mammalian cochlea is a remarkable sensory organ, capable of perceiving sound over a range of 10(12) in pressure, and discriminating both infrasonic and ultrasonic frequencies in different species. The sensory hair cells of the mammalian cochlea are exquisitely sensitive, responding to atomic-level deflections at speeds on the order of tens of microseconds. The number and placement of hair cells are precisely determined during inner ear development, and a large number of developmental processes sculpt the shape, size and morphology of these cells along the length of the cochlear duct to make them optimally responsive to different sound frequencies. In this review, we briefly discuss the evolutionary origins of the mammalian cochlea, and then describe the successive developmental processes that lead to its induction, cell cycle exit, cellular patterning and the establishment of topologically distinct frequency responses along its length. © 2015 Anatomical Society.

Effect of Cell Aspect Ratio on Swarming Bacteria

NASA Astrophysics Data System (ADS)

Ilkanaiv, Bella; Kearns, Daniel B.; Ariel, Gil; Be'er, Avraham

2017-04-01

Swarming bacteria collectively migrate on surfaces using flagella, forming dynamic whirls and jets that consist of millions of individuals. Because some swarming bacteria elongate prior to actual motion, cell aspect ratio may play a significant role in the collective dynamics. Extensive research on self-propelled rodlike particles confirms that elongation promotes alignment, strongly affecting the dynamics. Here, we study experimentally the collective dynamics of variants of swarming Bacillus subtilis that differ in length. We show that the swarming statistics depends on the aspect ratio in a critical, fundamental fashion not predicted by theory. The fastest motion was obtained for the wild-type and variants that are similar in length. However, shorter and longer cells exhibit anomalous, non-Gaussian statistics and nonexponential decay of the autocorrelation function, indicating lower collective motility. These results suggest that the robust mechanisms to maintain aspect ratios may be important for efficient swarming motility. Wild-type cells are optimal in this sense.

Ammonium Inhibits Primary Root Growth by Reducing the Length of Meristem and Elongation Zone and Decreasing Elemental Expansion Rate in the Root Apex in Arabidopsis thaliana

PubMed Central

Gao, Kun; Chen, Fanjun; Yuan, Lixing; Mi, Guohua

2013-01-01

The inhibitory effect of ammonium on primary root growth has been well documented; however the underlying physiological and molecular mechanisms are still controversial. To avoid ammonium toxicity to shoot growth, we used a vertical two-layer split plate system, in which the upper layer contained nitrate and the lower layer contained ammonium. In this way, nitrogen status was maintained and only the apical part of the root system was exposed to ammonium. Using a kinematic approach, we show here that 1 mM ammonium reduces primary root growth, decreasing both elemental expansion and cell production. Ammonium inhibits the length of elongation zone and the maximum elemental expansion rate. Ammonium also decreases the apparent length of the meristem as well as the number of dividing cells without affecting cell division rate. Moreover, ammonium reduces the number of root cap cells but appears to affect neither the status of root stem cell niche nor the distal auxin maximum at the quiescent center. Ammonium also inhibits root gravitropism and concomitantly down-regulates the expression of two pivotal auxin transporters, AUX1 and PIN2. Insofar as ammonium inhibits root growth rate in AUX1 and PIN2 loss-of-function mutants almost as strongly as in wild type, we conclude that ammonium inhibits root growth and gravitropism by largely distinct pathways. PMID:23577185

Fibroblast growth factor receptor inhibition induces loss of matrix MCL1 and necrosis in cholangiocarcinoma.

PubMed

Kabashima, Ayano; Hirsova, Petra; Bronk, Steven F; Hernandez, Matthew C; Truty, Mark J; Rizvi, Sumera; Kaufmann, Scott H; Gores, Gregory J

2018-03-08

Myeloid cell leukemia 1 (MCL1), a prosurvival member of the BCL2 protein family, has a pivotal role in human cholangiocarcinoma (CCA) cell survival. We previously reported that fibroblast growth factor receptor (FGFR) signalling mediates MCL1-dependent survival of CCA cells in vitro and in vivo. However, the mode and mechanisms of cell death in this model were not delineated. Human CCA cell lines were treated with the pan-FGFR inhibitor LY2874455 and the mode of cell death examined by several complementary assays. Mitochondrial oxidative metabolism was examined using a XF24 extracellular flux analyser. The efficiency of FGFR inhibition in patient-derived xenografts (PDX) was also assessed. CCA cells expressed two species of MCL1, a full-length form localised to the outer mitochondrial membrane, and an N terminus-truncated species compartmentalised within the mitochondrial matrix. The pan-FGFR inhibitor LY2874455 induced non-apoptotic cell death in the CCA cell lines associated with cellular depletion of both MCL1 species. The cell death was accompanied by failure of mitochondrial oxidative metabolism and was most consistent with necrosis. Enforced expression of N terminus-truncated MCL1 targeted to the mitochondrial matrix, but not full-length MCL1 targeted to the outer mitochondrial membrane, rescued cell death and mitochondrial function. LY2874455 treatment of PDX-bearing mice was associated with tumour cell loss of MCL1 and cell necrosis. FGFR inhibition induces loss of matrix MCL1, resulting in cell necrosis. These observations support a heretofore unidentified, alternative MCL1 survival function, namely prevention of cell necrosis, and have implications for treatment of human CCA. Herein, we report that therapeutic inhibition of a cell receptor expressed by bile duct cancer cells resulted in the loss of a critical survival protein termed MCL1. Cellular depletion of MCL1 resulted in the death of the cancer cells by a process characterised by cell rupture. Cell death by this process can stimulate the immune system and has implications for combination therapy using receptor inhibition with immunotherapy. Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Role of spatial averaging in multicellular gradient sensing.

PubMed

Smith, Tyler; Fancher, Sean; Levchenko, Andre; Nemenman, Ilya; Mugler, Andrew

2016-05-20

Gradient sensing underlies important biological processes including morphogenesis, polarization, and cell migration. The precision of gradient sensing increases with the length of a detector (a cell or group of cells) in the gradient direction, since a longer detector spans a larger range of concentration values. Intuition from studies of concentration sensing suggests that precision should also increase with detector length in the direction transverse to the gradient, since then spatial averaging should reduce the noise. However, here we show that, unlike for concentration sensing, the precision of gradient sensing decreases with transverse length for the simplest gradient sensing model, local excitation-global inhibition. The reason is that gradient sensing ultimately relies on a subtraction of measured concentration values. While spatial averaging indeed reduces the noise in these measurements, which increases precision, it also reduces the covariance between the measurements, which results in the net decrease in precision. We demonstrate how a recently introduced gradient sensing mechanism, regional excitation-global inhibition (REGI), overcomes this effect and recovers the benefit of transverse averaging. Using a REGI-based model, we compute the optimal two- and three-dimensional detector shapes, and argue that they are consistent with the shapes of naturally occurring gradient-sensing cell populations.

Role of spatial averaging in multicellular gradient sensing

NASA Astrophysics Data System (ADS)

Smith, Tyler; Fancher, Sean; Levchenko, Andre; Nemenman, Ilya; Mugler, Andrew

2016-06-01

Gradient sensing underlies important biological processes including morphogenesis, polarization, and cell migration. The precision of gradient sensing increases with the length of a detector (a cell or group of cells) in the gradient direction, since a longer detector spans a larger range of concentration values. Intuition from studies of concentration sensing suggests that precision should also increase with detector length in the direction transverse to the gradient, since then spatial averaging should reduce the noise. However, here we show that, unlike for concentration sensing, the precision of gradient sensing decreases with transverse length for the simplest gradient sensing model, local excitation-global inhibition. The reason is that gradient sensing ultimately relies on a subtraction of measured concentration values. While spatial averaging indeed reduces the noise in these measurements, which increases precision, it also reduces the covariance between the measurements, which results in the net decrease in precision. We demonstrate how a recently introduced gradient sensing mechanism, regional excitation-global inhibition (REGI), overcomes this effect and recovers the benefit of transverse averaging. Using a REGI-based model, we compute the optimal two- and three-dimensional detector shapes, and argue that they are consistent with the shapes of naturally occurring gradient-sensing cell populations.

The role of telomere dynamics in aging and cancer

NASA Astrophysics Data System (ADS)

Blagoev, Krastan; Goodwin, Edwin

2006-03-01

Telomere length changes are far more dynamic than previously thought. In addition to a gradual loss of ˜100 base pairs per telomere in each cell division, losses as well as gains may occur within a single cell cycle. We are investigating how telomere exchange, extension, and deletion affect the proliferative potential of telomerase-negative somatic cells. Experimental techniques are being devised to detect dynamic telomere processes and quantify both the frequency and length changes of each. In parallel, a ``dynamic telomere model'' is being used that incorporates telomere dynamics to study how the telomere size distribution evolves with time. This is an essential step towards understanding the role that telomere dynamics play in the normal aging of tissues and organisms. The model casts light on relationships not otherwise easily explained by a deterministic ``mitotic clock,'' or to what extent the shortest initial telomere determines the onset of senescence. We also expect to identify biomarkers that will correlate with aging better than average telomere length and to shed light on the transition to unlimited growth found in telomerase-negative tumor cells having the ALT (alternative lengthening of telomeres) phenotype, and to evaluate strategies to suppress the growth of these tumors.

Design and Optimization of Copper Indium Gallium Selenide Thin Film Solar Cells

DTIC Science & Technology

2015-09-01

determined by the intensity of the illumination that the solar cell is exposed to. The diffusion lengths L can be further defined by n n nL D τ...absorbers with graded Ga concentrations. (3) Back Contact Model Models for back contact silicon solar cells have been created with results that closely...Radiation. New York, NY: Academic Press, 2012. [12] B. Richards, “Enhancing the performance of silicon solar cells via the application of passive

Optimizing homeostatic cell renewal in hierarchical tissues

PubMed Central

Fider, Nicole A.

2018-01-01

In order to maintain homeostasis, mature cells removed from the top compartment of hierarchical tissues have to be replenished by means of differentiation and self-renewal events happening in the more primitive compartments. As each cell division is associated with a risk of mutation, cell division patterns have to be optimized, in order to minimize or delay the risk of malignancy generation. Here we study this optimization problem, focusing on the role of division tree length, that is, the number of layers of cells activated in response to the loss of terminally differentiated cells, which is related to the balance between differentiation and self-renewal events in the compartments. Using both analytical methods and stochastic simulations in a metapopulation-style model, we find that shorter division trees are advantageous if the objective is to minimize the total number of one-hit mutants in the cell population. Longer division trees on the other hand minimize the accumulation of two-hit mutants, which is a more likely evolutionary goal given the key role played by tumor suppressor genes in cancer initiation. While division tree length is the most important property determining mutant accumulation, we also find that increasing the size of primitive compartments helps to delay two-hit mutant generation. PMID:29447149

Simulation of action potential propagation in plants.

PubMed

Sukhov, Vladimir; Nerush, Vladimir; Orlova, Lyubov; Vodeneev, Vladimir

2011-12-21

Action potential is considered to be one of the primary responses of a plant to action of various environmental factors. Understanding plant action potential propagation mechanisms requires experimental investigation and simulation; however, a detailed mathematical model of plant electrical signal transmission is absent. Here, the mathematical model of action potential propagation in plants has been worked out. The model is a two-dimensional system of excitable cells; each of them is electrically coupled with four neighboring ones. Ion diffusion between excitable cell apoplast areas is also taken into account. The action potential generation in a single cell has been described on the basis of our previous model. The model simulates active and passive signal transmission well enough. It has been used to analyze theoretically the influence of cell to cell electrical conductivity and H(+)-ATPase activity on the signal transmission in plants. An increase in cell to cell electrical conductivity has been shown to stimulate an increase in the length constant, the action potential propagation velocity and the temperature threshold, while the membrane potential threshold being weakly changed. The growth of H(+)-ATPase activity has been found to induce the increase of temperature and membrane potential thresholds and the reduction of the length constant and the action potential propagation velocity. Copyright © 2011 Elsevier Ltd. All rights reserved.

Cell response to quasi-monochromatic light with different coherence

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

Budagovsky, A V; Solovykh, N V; Budagovskaya, O N

The problem of the light coherence effect on the magnitude of the photoinduced cell response is discussed. The origins of ambiguous interpretation of the known experimental results are considered. Using the biological models, essentially differing in anatomy, morphology and biological functions (acrospires of radish, blackberry microsprouts cultivated in vitro, plum pollen), the effect of statistical properties of quasi-monochromatic light (λ{sub max} = 633 nm) on the magnitude of the photoinduced cell response is shown. It is found that for relatively low spatial coherence, the cell functional activity changes insignificantly. The maximal enhancement of growing processes (stimulating effect) is observed whenmore » the coherence length L{sub coh} and the correlation radius r{sub cor} are greater than the cell size, i.e., the entire cell fits into the field coherence volume. In this case, the representative indicators (germination of seeds and pollen, the spears length) exceeds those of non-irradiated objects by 1.7 – 3.9 times. For more correct assessment of the effect of light statistical properties on photocontrol processes, it is proposed to replace the qualitative description (coherent – incoherent) with the quantitative one, using the determination of spatial and temporal correlation functions and comparing them with the characteristic dimensions of the biological structures, e.g., the cell size. (biophotonics)« less

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.

A free-boundary model of a motile cell explains turning behavior.

PubMed

Nickaeen, Masoud; Novak, Igor L; Pulford, Stephanie; Rumack, Aaron; Brandon, Jamie; Slepchenko, Boris M; Mogilner, Alex

2017-11-01

To understand shapes and movements of cells undergoing lamellipodial motility, we systematically explore minimal free-boundary models of actin-myosin contractility consisting of the force-balance and myosin transport equations. The models account for isotropic contraction proportional to myosin density, viscous stresses in the actin network, and constant-strength viscous-like adhesion. The contraction generates a spatially graded centripetal actin flow, which in turn reinforces the contraction via myosin redistribution and causes retraction of the lamellipodial boundary. Actin protrusion at the boundary counters the retraction, and the balance of the protrusion and retraction shapes the lamellipodium. The model analysis shows that initiation of motility critically depends on three dimensionless parameter combinations, which represent myosin-dependent contractility, a characteristic viscosity-adhesion length, and a rate of actin protrusion. When the contractility is sufficiently strong, cells break symmetry and move steadily along either straight or circular trajectories, and the motile behavior is sensitive to conditions at the cell boundary. Scanning of a model parameter space shows that the contractile mechanism of motility supports robust cell turning in conditions where short viscosity-adhesion lengths and fast protrusion cause an accumulation of myosin in a small region at the cell rear, destabilizing the axial symmetry of a moving cell.

Inhibition of mTOR by Rapamycin Results in Auditory Hair Cell Damage and Decreased Spiral Ganglion Neuron Outgrowth and Neurite Formation In Vitro

PubMed Central

Leitmeyer, Katharina; Glutz, Andrea; Radojevic, Vesna; Setz, Cristian; Huerzeler, Nathan; Bumann, Helen; Bodmer, Daniel; Brand, Yves

2015-01-01

Rapamycin is an antifungal agent with immunosuppressive properties. Rapamycin inhibits the mammalian target of rapamycin (mTOR) by blocking the mTOR complex 1 (mTORC1). mTOR is an atypical serine/threonine protein kinase, which controls cell growth, cell proliferation, and cell metabolism. However, less is known about the mTOR pathway in the inner ear. First, we evaluated whether or not the two mTOR complexes (mTORC1 and mTORC2, resp.) are present in the mammalian cochlea. Next, tissue explants of 5-day-old rats were treated with increasing concentrations of rapamycin to explore the effects of rapamycin on auditory hair cells and spiral ganglion neurons. Auditory hair cell survival, spiral ganglion neuron number, length of neurites, and neuronal survival were analyzed in vitro. Our data indicates that both mTOR complexes are expressed in the mammalian cochlea. We observed that inhibition of mTOR by rapamycin results in a dose dependent damage of auditory hair cells. Moreover, spiral ganglion neurite number and length of neurites were significantly decreased in all concentrations used compared to control in a dose dependent manner. Our data indicate that the mTOR may play a role in the survival of hair cells and modulates spiral ganglion neuronal outgrowth and neurite formation. PMID:25918725

Intermediate progenitors are increased by lengthening of the cell cycle through calcium signaling and p53 expression in human neural progenitors

PubMed Central

García-García, Elisa; Pino-Barrio, María José; López-Medina, Laura; Martínez-Serrano, Alberto

2012-01-01

During development, neurons can be generated directly from a multipotent progenitor or indirectly through an intermediate progenitor (IP). This last mode of division amplifies the progeny of neurons. The mechanisms governing the generation and behavior of IPs are not well understood. In this work, we demonstrate that the lengthening of the cell cycle enhances the generation of neurons in a human neural progenitor cell system in vitro and also the generation and expansion of IPs. These IPs are insulinoma-associated 1 (Insm1)+/BTG family member 2 (Btg2)−, which suggests an increase in a self-amplifying IP population. Later the cultures express neurogenin 2 (Ngn2) and become neurogenic. The signaling responsible for this cell cycle modulation is investigated. It is found that the release of calcium from the endoplasmic reticulum to the cytosol in response to B cell lymphoma-extra large overexpression or ATP addition lengths the cell cycle and increases the number of IPs and, in turn, the final neuron outcome. Moreover, data suggest that the p53–p21 pathway is responsible for the changes in cell cycle. In agreement with this, increased p53 levels are necessary for a calcium-induced increase in neurons. Our findings contribute to understand how calcium signaling can modulate cell cycle length during neurogenesis. PMID:22323293

Inhibition of mTOR by Rapamycin Results in Auditory Hair Cell Damage and Decreased Spiral Ganglion Neuron Outgrowth and Neurite Formation In Vitro.

PubMed

Leitmeyer, Katharina; Glutz, Andrea; Radojevic, Vesna; Setz, Cristian; Huerzeler, Nathan; Bumann, Helen; Bodmer, Daniel; Brand, Yves

2015-01-01

Rapamycin is an antifungal agent with immunosuppressive properties. Rapamycin inhibits the mammalian target of rapamycin (mTOR) by blocking the mTOR complex 1 (mTORC1). mTOR is an atypical serine/threonine protein kinase, which controls cell growth, cell proliferation, and cell metabolism. However, less is known about the mTOR pathway in the inner ear. First, we evaluated whether or not the two mTOR complexes (mTORC1 and mTORC2, resp.) are present in the mammalian cochlea. Next, tissue explants of 5-day-old rats were treated with increasing concentrations of rapamycin to explore the effects of rapamycin on auditory hair cells and spiral ganglion neurons. Auditory hair cell survival, spiral ganglion neuron number, length of neurites, and neuronal survival were analyzed in vitro. Our data indicates that both mTOR complexes are expressed in the mammalian cochlea. We observed that inhibition of mTOR by rapamycin results in a dose dependent damage of auditory hair cells. Moreover, spiral ganglion neurite number and length of neurites were significantly decreased in all concentrations used compared to control in a dose dependent manner. Our data indicate that the mTOR may play a role in the survival of hair cells and modulates spiral ganglion neuronal outgrowth and neurite formation.

Mathematical models of tumor heterogeneity and drug resistance

NASA Astrophysics Data System (ADS)

Greene, James

In this dissertation we develop mathematical models of tumor heterogeneity and drug resistance in cancer chemotherapy. Resistance to chemotherapy is one of the major causes of the failure of cancer treatment. Furthermore, recent experimental evidence suggests that drug resistance is a complex biological phenomena, with many influences that interact nonlinearly. Here we study the influence of such heterogeneity on treatment outcomes, both in general frameworks and under specific mechanisms. We begin by developing a mathematical framework for describing multi-drug resistance to cancer. Heterogeneity is reflected by a continuous parameter, which can either describe a single resistance mechanism (such as the expression of P-gp in the cellular membrane) or can account for the cumulative effect of several mechanisms and factors. The model is written as a system of integro-differential equations, structured by the continuous "trait," and includes density effects as well as mutations. We study the limiting behavior of the model, both analytically and numerically, and apply it to study treatment protocols. We next study a specific mechanism of tumor heterogeneity and its influence on cell growth: the cell-cycle. We derive two novel mathematical models, a stochastic agent-based model and an integro-differential equation model, each of which describes the growth of cancer cells as a dynamic transition between proliferative and quiescent states. By examining the role all parameters play in the evolution of intrinsic tumor heterogeneity, and the sensitivity of the population growth to parameter values, we show that the cell-cycle length has the most significant effect on the growth dynamics. In addition, we demonstrate that the agent-based model can be approximated well by the more computationally efficient integro-differential equations, when the number of cells is large. The model is closely tied to experimental data of cell growth, and includes a novel implementation of transition rates as a function of global density. Finally, we extend the model of cell-cycle heterogeneity to include spatial variables. Cells are modeled as soft spheres and exhibit attraction/repulsion/random forces. A fundamental hypothesis is that cell-cycle length increases with local density, thus producing a distribution of observed division lengths. Apoptosis occurs primarily through an extended period of unsuccessful proliferation, and the explicit mechanism of the drug (Paclitaxel) is modeled as an increase in cell-cycle duration. We show that the distribution of cell-cycle lengths is highly time-dependent, with close time-averaged agreement with the distribution used in the previous work. Furthermore, survival curves are calculated and shown to qualitatively agree with experimental data in different densities and geometries, thus relating the cellular microenvironment to drug resistance.

Adiabatic quantum-flux-parametron cell library adopting minimalist design

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

Takeuchi, Naoki, E-mail: takeuchi-naoki-kx@ynu.jp; Yamanashi, Yuki; Yoshikawa, Nobuyuki

We herein build an adiabatic quantum-flux-parametron (AQFP) cell library adopting minimalist design and a symmetric layout. In the proposed minimalist design, every logic cell is designed by arraying four types of building block cells: buffer, NOT, constant, and branch cells. Therefore, minimalist design enables us to effectively build and customize an AQFP cell library. The symmetric layout reduces unwanted parasitic magnetic coupling and ensures a large mutual inductance in an output transformer, which enables very long wiring between logic cells. We design and fabricate several logic circuits using the minimal AQFP cell library so as to test logic cells inmore » the library. Moreover, we experimentally investigate the maximum wiring length between logic cells. Finally, we present an experimental demonstration of an 8-bit carry look-ahead adder designed using the minimal AQFP cell library and demonstrate that the proposed cell library is sufficiently robust to realize large-scale digital circuits.« less

Adiabatic quantum-flux-parametron cell library adopting minimalist design

NASA Astrophysics Data System (ADS)

Takeuchi, Naoki; Yamanashi, Yuki; Yoshikawa, Nobuyuki

2015-05-01

We herein build an adiabatic quantum-flux-parametron (AQFP) cell library adopting minimalist design and a symmetric layout. In the proposed minimalist design, every logic cell is designed by arraying four types of building block cells: buffer, NOT, constant, and branch cells. Therefore, minimalist design enables us to effectively build and customize an AQFP cell library. The symmetric layout reduces unwanted parasitic magnetic coupling and ensures a large mutual inductance in an output transformer, which enables very long wiring between logic cells. We design and fabricate several logic circuits using the minimal AQFP cell library so as to test logic cells in the library. Moreover, we experimentally investigate the maximum wiring length between logic cells. Finally, we present an experimental demonstration of an 8-bit carry look-ahead adder designed using the minimal AQFP cell library and demonstrate that the proposed cell library is sufficiently robust to realize large-scale digital circuits.

Log (TG)/HDL-C ratio as a predictor of decreased islet beta cell function in patients with type 2 diabetes: 6-year cohort study.

PubMed

Zhou, Meicen; Li, Zengyi; Min, Rui; Dong, Yaxiu; Sun, Qi; Li, Yuxiu

2015-09-01

The aim of the present study was to explore whether the triglyceride to high density lipoprotein cholesterol ratio [log (TG)/HDL-C] and peripheral blood leukocytes DNA telomere length could predict future islet beta cell function decreased in Chinese type 2 diabetes mellitus (T2DM) during a 6-year cohort. Sixty T2DM patients (without insulin treatment at baseline) were included in the 6-year cohort study. Peripheral blood leukocytes DNA telomere length, HbA1c, blood lipid profile, fatty fat acid, glucose, insulin and C peptide (3 h after a mixed meal) were determined. Delta C peptide area under curve (Delta CP AUC) was used to reflect change in beta cell secretion function (Delta CP AUC = baseline CP AUC - CP AUC after 6 years). Subjects were divided into slow decrease of beta cell function group (Delta CP AUCslow group) and fast decrease group (Delta CP AUCfast group) according to median of Delta CP AUC. Baseline demographic characteristics, clinical variables between two groups were compared. Correlations between baseline data and Delta CP AUC were analyzed. Baseline log (TG)/HDL-C was positively correlated with Delta CP AUC (r = 0.306, P = 0.027); log (TG)/HDL-C in Delta CP AUCfast group was higher than that in Delta CP AUCslow group (0.103 ± 0.033 vs 0.083 ± 0.030, P = 0.027). There was no significant difference in DNA telomere length between the two groups. Change in DNA telomere length over 6 years was not significantly correlated with baseline blood lipid. In Chinese T2DM patients, high baseline log (TG)/HDL-C ratio predicts fast progression of islet beta cell dysfunction. It may be a simple index to predict progression speed of islet beta cell dysfunction. © 2014 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd.

The availability of filament ends modulates actin stochastic dynamics in live plant cells

PubMed Central

Li, Jiejie; Staiger, Benjamin H.; Henty-Ridilla, Jessica L.; Abu-Abied, Mohamad; Sadot, Einat; Blanchoin, Laurent; Staiger, Christopher J.

2014-01-01

A network of individual filaments that undergoes incessant remodeling through a process known as stochastic dynamics comprises the cortical actin cytoskeleton in plant epidermal cells. From images at high spatial and temporal resolution, it has been inferred that the regulation of filament barbed ends plays a central role in choreographing actin organization and turnover. How this occurs at a molecular level, whether different populations of ends exist in the array, and how individual filament behavior correlates with the overall architecture of the array are unknown. Here we develop an experimental system to modulate the levels of heterodimeric capping protein (CP) and examine the consequences for actin dynamics, architecture, and cell expansion. Significantly, we find that all phenotypes are the opposite for CP-overexpression (OX) cells compared with a previously characterized cp-knockdown line. Specifically, CP OX lines have fewer filament–filament annealing events, as well as reduced filament lengths and lifetimes. Further, cp-knockdown and OX lines demonstrate the existence of a subpopulation of filament ends sensitive to CP concentration. Finally, CP levels correlate with the biological process of axial cell expansion; for example, epidermal cells from hypocotyls with reduced CP are longer than wild-type cells, whereas CP OX lines have shorter cells. On the basis of these and other genetic studies in this model system, we hypothesize that filament length and lifetime positively correlate with the extent of axial cell expansion in dark-grown hypocotyls. PMID:24523291

Distinct Domains of CheA Confer Unique Functions in Chemotaxis and Cell Length in Azospirillum brasilense Sp7.

PubMed

Gullett, Jessica M; Bible, Amber; Alexandre, Gladys

2017-07-01

Chemotaxis is the movement of cells in response to gradients of diverse chemical cues. Motile bacteria utilize a conserved chemotaxis signal transduction system to bias their motility and navigate through a gradient. A central regulator of chemotaxis is the histidine kinase CheA. This cytoplasmic protein interacts with membrane-bound receptors, which assemble into large polar arrays, to propagate the signal. In the alphaproteobacterium Azospirillum brasilense , Che1 controls transient increases in swimming speed during chemotaxis, but it also biases the cell length at division. However, the exact underlying molecular mechanisms for Che1-dependent control of multiple cellular behaviors are not known. Here, we identify specific domains of the CheA1 histidine kinase implicated in modulating each of these functions. We show that CheA1 is produced in two isoforms: a membrane-anchored isoform produced as a fusion with a conserved seven-transmembrane domain of unknown function (TMX) at the N terminus and a soluble isoform similar to prototypical CheA. Site-directed and deletion mutagenesis combined with behavioral assays confirm the role of CheA1 in chemotaxis and implicate the TMX domain in mediating changes in cell length. Fluorescence microscopy further reveals that the membrane-anchored isoform is distributed around the cell surface while the soluble isoform localizes at the cell poles. Together, the data provide a mechanism for the role of Che1 in controlling multiple unrelated cellular behaviors via acquisition of a new domain in CheA1 and production of distinct functional isoforms. IMPORTANCE Chemotaxis provides a significant competitive advantage to bacteria in the environment, and this function has been transferred laterally multiple times, with evidence of functional divergence in different genomic contexts. The molecular principles that underlie functional diversification of chemotaxis in various genomic contexts are unknown. Here, we provide a molecular mechanism by which a single CheA protein controls two unrelated functions: chemotaxis and cell length. Acquisition of this multifunctionality is seemingly a recent evolutionary event. The findings illustrate a mechanism by which chemotaxis function may be co-opted to regulate additional cellular functions. Copyright © 2017 American Society for Microbiology.

Distinct Domains of CheA Confer Unique Functions in Chemotaxis and Cell Length in Azospirillum brasilense Sp7

PubMed Central

Gullett, Jessica M.

2017-01-01

ABSTRACT Chemotaxis is the movement of cells in response to gradients of diverse chemical cues. Motile bacteria utilize a conserved chemotaxis signal transduction system to bias their motility and navigate through a gradient. A central regulator of chemotaxis is the histidine kinase CheA. This cytoplasmic protein interacts with membrane-bound receptors, which assemble into large polar arrays, to propagate the signal. In the alphaproteobacterium Azospirillum brasilense, Che1 controls transient increases in swimming speed during chemotaxis, but it also biases the cell length at division. However, the exact underlying molecular mechanisms for Che1-dependent control of multiple cellular behaviors are not known. Here, we identify specific domains of the CheA1 histidine kinase implicated in modulating each of these functions. We show that CheA1 is produced in two isoforms: a membrane-anchored isoform produced as a fusion with a conserved seven-transmembrane domain of unknown function (TMX) at the N terminus and a soluble isoform similar to prototypical CheA. Site-directed and deletion mutagenesis combined with behavioral assays confirm the role of CheA1 in chemotaxis and implicate the TMX domain in mediating changes in cell length. Fluorescence microscopy further reveals that the membrane-anchored isoform is distributed around the cell surface while the soluble isoform localizes at the cell poles. Together, the data provide a mechanism for the role of Che1 in controlling multiple unrelated cellular behaviors via acquisition of a new domain in CheA1 and production of distinct functional isoforms. IMPORTANCE Chemotaxis provides a significant competitive advantage to bacteria in the environment, and this function has been transferred laterally multiple times, with evidence of functional divergence in different genomic contexts. The molecular principles that underlie functional diversification of chemotaxis in various genomic contexts are unknown. Here, we provide a molecular mechanism by which a single CheA protein controls two unrelated functions: chemotaxis and cell length. Acquisition of this multifunctionality is seemingly a recent evolutionary event. The findings illustrate a mechanism by which chemotaxis function may be co-opted to regulate additional cellular functions. PMID:28416707

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

Impact of alemtuzumab on HIV persistence in an HIV-infected individual on antiretroviral therapy with Sezary syndrome.

PubMed

Rasmussen, Thomas A; McMahon, James; Chang, J Judy; Symons, Jori; Roche, Michael; Dantanarayana, Ashanti; Okoye, Afam; Hiener, Bonnie; Palmer, Sarah; Lee, Wen Shi; Kent, Stephen J; Van Der Weyden, Carrie; Prince, H Miles; Cameron, Paul U; Lewin, Sharon R

2017-08-24

To study the effects of alemtuzumab on HIV persistence in an HIV-infected individual on antiretroviral therapy (ART) with Sezary syndrome, a rare malignancy of CD4 T cells. Case report. Blood was collected 30 and 18 months prior to presentation with Sezary syndrome, at the time of presentation and during alemtuzumab. T-cell subsets in malignant (CD7-CD26-TCR-VBeta2+) and nonmalignant cells were quantified by flow cytometry. HIV-DNA in total CD4 T cells, in sorted malignant and nonmalignant CD4 T cells, was quantified by PCR and clonal expansion of HIV-DNA assessed by full-length next-generation sequencing. HIV-hepatitis B virus coinfection was diagnosed and antiretroviral therapy initiated 4 years prior to presentation with Sezary syndrome and primary cutaneous anaplastic large cell lymphoma. The patient received alemtuzumab 10 mg three times per week for 4 weeks but died 6 weeks post alemtuzumab. HIV-DNA was detected in nonmalignant but not in malignant CD4 T cells, consistent with expansion of a noninfected CD4 T-cell clone. Full-length HIV-DNA sequencing demonstrated multiple defective viruses but no identical or expanded sequences. Alemtuzumab extensively depleted T cells, including more than 1 log reduction in total T cells and more than 3 log reduction in CD4 T cells. Finally, alemtuzumab decreased HIV-DNA in CD4 T cells by 57% but HIV-DNA remained detectable at low levels even after depletion of nearly all CD4 T cells. Alemtuzumab extensively depleted multiple T-cell subsets and decreased the frequency of but did not eliminate HIV-infected CD4 T cells. Studying the effects on HIV persistence following immune recovery in HIV-infected individuals who require alemtuzumab for malignancy or in animal studies may provide further insights into novel cure strategies.

Development of a Space-Rated Proton Exchange Membrane Fuel Cell

NASA Technical Reports Server (NTRS)

Hoffman, William C., III; Vasquez, Arturo; Lazaroff, Scott M.; Downey, Michael G.

1999-01-01

Power systems for human spacecraft have historically included fuel cells due to the superior energy density they offer over battery systems depending on mission length and power consumption. As space exploration focuses on the evolution of reusable spacecraft and also considers planetary exploration power system requirements, fuel cells continue to be a factor in the potential system solutions.

Mechanism of Telomerase Activation by v-Rel and Its Contribution to Transformation

PubMed Central

Hrdličková, Radmila; Nehyba, Jiří; Liss, Andrew S.; Bose, Henry R.

2006-01-01

Telomerase is activated during the transformation of lymphoid cells and fibroblasts by v-Rel, the oncogenic member of the Rel/NF-κB family of transcription factors. v-Rel-transformed cell lines have longer telomeres than untransformed chicken lymphoid cells and have high levels of telomerase activity. v-Rel-mediated activation of telomerase is achieved by multiple mechanisms. The expression of the gene encoding the catalytic subunit of telomerase (TERT) was directly upregulated by v-Rel. Moreover, the expression of v-Rel altered the ratio of alternatively spliced and full-length TERT transcripts in favor of the full-length forms. The activation of telomerase by v-Rel in lymphocytes was also accompanied by inactivation of nuclear inhibitors. The inhibition of telomerase activity in v-Rel-transformed cell lines led to apoptosis within 24 h. The expression of v-Rel in a macrophage cell line resulted in elevated levels of reactive oxygen species (ROS), increased telomerase activity, and increased sensitivity to telomerase inhibitors. In contrast, the ectopic expression of TERT decreased the extent of apoptosis induced by ROS. The activation of telomerase by v-Rel may, therefore, partially protect the transformed cells from apoptosis induced by ROS. PMID:16352553

Halogenated auxins affect microtubules and root elongation in Lactuca sativa

NASA Technical Reports Server (NTRS)

Zhang, N.; Hasenstein, K. H.

2000-01-01

We studied the effect of 4,4,4-trifluoro-3-(indole-3-)butyric acid (TFIBA), a recently described root growth stimulator, and 5,6-dichloro-indole-3-acetic acid (DCIAA) on growth and microtubule (MT) organization in roots of Lactuca sativa L. DCIAA and indole-3-butyric acid (IBA) inhibited root elongation and depolymerized MTs in the cortex of the elongation zone, inhibited the elongation of stele cells, and promoted xylem maturation. Both auxins caused the plane of cell division to shift from anticlinal to periclinal. In contrast, TFIBA (100 micromolar) promoted elongation of primary roots by 40% and stimulated the elongation of lateral roots, even in the presence of IBA, the microtubular inhibitors oryzalin and taxol, or the auxin transport inhibitor naphthylphthalamic acid. However, TFIBA inhibited the formation of lateral root primordia. Immunostaining showed that TFIBA stabilized MTs orientation perpendicular to the root axis, doubled the cortical cell length, but delayed xylem maturation. The data indicate that the auxin-induced inhibition of elongation and swelling of roots results from reoriented phragmoplasts, the destabilization of MTs in elongating cells, and promotion of vessel formation. In contrast, TFIBA induced promotion of root elongation by enhancing cell length, prolonging transverse MT orientation, delaying cell and xylem maturation.

Modulation of flagellum attachment zone protein FLAM3 and regulation of the cell shape in Trypanosoma brucei life cycle transitions

PubMed Central

Sunter, Jack D.; Benz, Corinna; Andre, Jane; Whipple, Sarah; McKean, Paul G.; Gull, Keith; Ginger, Michael L.; Lukeš, Julius

2015-01-01

ABSTRACT The cell shape of Trypanosoma brucei is influenced by flagellum-to-cell-body attachment through a specialised structure – the flagellum attachment zone (FAZ). T. brucei exhibits numerous morphological forms during its life cycle and, at each stage, the FAZ length varies. We have analysed FLAM3, a large protein that localises to the FAZ region within the old and new flagellum. Ablation of FLAM3 expression causes a reduction in FAZ length; however, this has remarkably different consequences in the tsetse procyclic form versus the mammalian bloodstream form. In procyclic form cells FLAM3 RNAi results in the transition to an epimastigote-like shape, whereas in bloodstream form cells a severe cytokinesis defect associated with flagellum detachment is observed. Moreover, we demonstrate that the amount of FLAM3 and its localisation is dependent on ClpGM6 expression and vice versa. This evidence demonstrates that FAZ is a key regulator of trypanosome shape, with experimental perturbations being life cycle form dependent. An evolutionary cell biology explanation suggests that these differences are a reflection of the division process, the cytoskeleton and intrinsic structural plasticity of particular life cycle forms. PMID:26148511

Interconnection of bundled solid oxide fuel cells

DOEpatents

Brown, Michael; Bessette, II, Norman F; Litka, Anthony F; Schmidt, Douglas S

2014-01-14

A system and method for electrically interconnecting a plurality of fuel cells to provide dense packing of the fuel cells. Each one of the plurality of fuel cells has a plurality of discrete electrical connection points along an outer surface. Electrical connections are made directly between the discrete electrical connection points of adjacent fuel cells so that the fuel cells can be packed more densely. Fuel cells have at least one outer electrode and at least one discrete interconnection to an inner electrode, wherein the outer electrode is one of a cathode and and anode and wherein the inner electrode is the other of the cathode and the anode. In tubular solid oxide fuel cells the discrete electrical connection points are spaced along the length of the fuel cell.

The role of telomeres and telomerase complex in haematological neoplasia: the length of telomeres as a marker of carcinogenesis and prognosis of disease.

PubMed

Gancarcíková, M; Zemanová, Z; Brezinová, J; Berková, A; Vcelíková, S; Smigová, J; Michalová, K

2010-01-01

Human telomeres (discovery of telomere structure and function has been recently awarded The Nobel Prize) consist of approximately 5-12 kb of tandem repeated sequences (TTAGGG)n and associated proteins capping chromosome ends which prevent degradation, loss of genetic information, end-to-end fusion, senescence and apoptosis. Due to the end-replication problem, telomere repeats are lost with each cell division, eventually leading to genetic instability and cellular senescence when telomeres become critically short. Stabilization of the telomeric DNA through telomerase activation, unique reverse transcriptase, or activation of the alternative mechanism of telomere maintenance is essential if the cells are to survive and proliferate indefinitely. Telomerase is expressed during early development and remains fully active in specific germline cells, but is undetectable in most normal somatic cells. High level of telomerase activity is detected in almost 90% of human tumours and immortalized cell lines. The hematopoietic compartment may develop genetic instability as a consequence of telomere erosion, resulting in aplastic anaemia (AA) and increased risk of myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). Genetic instability associated with telomere dysfunction (i.e. short telomeres) is an early event in carcinogenesis. The molecular cytogenetic method telomere/centromere fluorescence in situ hybridization (T/C-FISH) can be used to characterize the telomere length of hematopoietic cells. This review describes recent advances in the molecular characterization of telomere system, the regulation of telomerase activity in cancer pathogenesis and shows that the telomeric length could be a potential clinical marker of hematologic neoplasia and prognosis of disease.

Anisotropic Cell Expansion Is Affected through the Bidirectional Mobility of Cellulose Synthase Complexes and Phosphorylation at Two Critical Residues on CESA31[OPEN

PubMed Central

Liu, Yanmei; Bauer, Stefan

2016-01-01

Here we report that phosphorylation status of S211 and T212 of the CESA3 component of Arabidopsis (Arabidopsis thaliana) cellulose synthase impacts the regulation of anisotropic cell expansion as well as cellulose synthesis and deposition and microtubule-dependent bidirectional mobility of CESA complexes. Mutation of S211 to Ala caused a significant decrease in the length of etiolated hypocotyls and primary roots, while root hairs were not significantly affected. By contrast, the S211E mutation stunted the growth of root hairs, but primary roots were not significantly affected. Similarly, T212E caused a decrease in the length of root hairs but not root length. However, T212E stunted the growth of etiolated hypocotyls. Live-cell imaging of fluorescently labeled CESA showed that the rate of movement of CESA particles was directionally asymmetric in etiolated hypocotyls of S211A and T212E mutants, while similar bidirectional velocities were observed with the wild-type control and S211E and T212A mutant lines. Analysis of cell wall composition and the innermost layer of cell wall suggests a role for phosphorylation of CESA3 S211 and T212 in cellulose aggregation into fibrillar bundles. These results suggest that microtubule-guided bidirectional mobility of CESA complexes is fine-tuned by phosphorylation of CESA3 S211 and T212, which may, in turn, modulate cellulose synthesis and organization, resulting in or contributing to the observed defects of anisotropic cell expansion. PMID:26969722

A Short Isoform of Human Cytomegalovirus US3 Functions as a Dominant Negative Inhibitor of the Full-Length Form

PubMed Central

Shin, Jinwook; Park, Boyoun; Lee, Sungwook; Kim, Youngkyun; Biegalke, Bonita J.; Kang, Seongman; Ahn, Kwangseog

2006-01-01

Human cytomegalovirus encodes four unique short (US) region proteins, each of which is independently sufficient for causing the down-regulation of major histocompatibility complex (MHC) class I molecules on the cell surface. This down-regulation enables infected cells to evade recognition by cytotoxic T lymphocytes (CTLs) but makes them vulnerable to lysis by natural killer (NK) cells, which lyse those cells that lack MHC class I molecules. The 22-kDa US3 glycoprotein is able to down-regulate the surface expression of MHC class I molecules by dual mechanisms: direct endoplasmic reticulum retention by physical association and/or tapasin inhibition. The alternative splicing of the US3 gene generates two additional products, including 17-kDa and 3.5-kDa truncated isoforms; however, the functional significance of these isoforms during viral infection is unknown. Here, we describe a novel mode of self-regulation of US3 function that uses the endogenously produced truncated isoform. The truncated isoform itself neither binds to MHC class I molecules nor prevents the full-length US3 from interacting with MHC class I molecules. Instead, the truncated isoform associates with tapasin and competes with full-length US3 for binding to tapasin; thus, it suppresses the action of US3 that causes the disruption of the function of tapasin. Our results indicate that the truncated isoform of the US3 locus acts as a dominant negative regulator of full-length US3 activity. These data reflect the manner in which the virus has developed temporal survival strategies during viral infection against immune surveillance involving both CTLs and NK cells. PMID:16699020

Functional PAK-2 knockout and replacement with a caspase cleavage-deficient mutant in mice reveals differential requirements of full-length PAK-2 and caspase-activated PAK-2p34.

PubMed

Marlin, Jerry W; Chang, Yu-Wen E; Ober, Margaret; Handy, Amy; Xu, Wenhao; Jakobi, Rolf

2011-06-01

p21-Activated protein kinase 2 (PAK-2) has both anti- and pro-apoptotic functions depending on its mechanism of activation. Activation of full-length PAK-2 by the monomeric GTPases Cdc42 or Rac stimulates cell survival, whereas caspase activation of PAK-2 to the PAK-2p34 fragment is involved in the apoptotic response. In this study we use functional knockout of PAK-2 and gene replacement with the caspase cleavage-deficient PAK-2D212N mutant to differentiate the biological functions of full-length PAK-2 and caspase-activated PAK-2p34. Knockout of PAK-2 results in embryonic lethality at early stages before organ development, whereas replacement with the caspase cleavage-deficient PAK-2D212N results in viable and healthy mice, indicating that early embryonic lethality is caused by deficiency of full-length PAK-2 rather than lack of caspase activation to the PAK-2p34 fragment. However, deficiency of caspase activation of PAK-2 decreased spontaneous cell death of primary mouse embryonic fibroblasts and increased cell growth at high cell density. In contrast, stress-induced cell death by treatment with the anti-cancer drug cisplatin was not reduced by deficiency of caspase activation of PAK-2, but switched from an apoptotic to a nonapoptotic, caspase-independent mechanism. Homozygous PAK-2D212N primary mouse embryonic fibroblasts that lack the ability to generate the proapoptotic PAK-2p34 show less activation of the effector caspase 3, 6, and 7, indicating that caspase activation of PAK-2 amplifies the apoptotic response through a positive feedback loop resulting in more activation of effector caspases.

[Telomere lengthening by trichostatin A treatment in cloned pigs].

PubMed

Xie, Bing-Teng; Ji, Guang-Zhen; Kong, Qing-Ran; Mao, Jian; Shi, Yong-Qian; Liu, Shi-Chao; Wu, Mei-Ling; Wang, Juan; Liu, Lin; Liu, Zhong-Hua

2012-12-01

Telomeres are repeated GC rich sequences at the end of chromosomes, and shorten with each cell division due to DNA end replication problem. Previously, reprogrammed somatic cells of cloned animals display variable telomere elongation. However, it was reported that the cloned animals including Dolly do not reset telomeres and show premature aging. In this study, we investigated telomere function in cloned or transgenic cloned pigs, including the cloned Northeast Min pigs, eGFP, Mx, and PGC1α transgenic cloned pigs, and found that the telomere lengths of cloned pigs were significantly shorter than the nuclear donor adult fibroblasts and age-matched noncloned pigs (P<0.05), indicating that nuclear reprogramming did not restore cellular age of donor cells after somatic cell nuclear transfer (SCNT). Trichostatin A (TSA), an inhibitor of histone deacetylase, has proven to enhance the efficiency of nuclear reprogramming in several species. In order to test whether TSA also can effectively enhance reprogramming of telomeres, TSA (40 nmol/L) was used to treat porcine cloned embryos at 1-cell stage for 24 h. Consistent with previous reports, the developmental rate of SCNT embryos to the blastocyst stage was significantly increased compared with those of the control group (16.35% vs. 27.09%, 21.60% vs. 34.90%, P<0.05). Notably, the telomere length of cloned porcine blastocysts was also significantly elongated (P<0.05). Although TSA did not improve the cloning efficiency (1.3% vs. 1.7%, TSA vs. control), the telomere lengths of cloned pig-lets were significantly longer compared with those of the control group and the donor fibroblasts (P<0.05). In conclusion, telomeres have not been effectively restored by SCNT in pigs but TSA can effectively lengthen the telomere lengths of cloned pigs.

A short isoform of human cytomegalovirus US3 functions as a dominant negative inhibitor of the full-length form.

PubMed

Shin, Jinwook; Park, Boyoun; Lee, Sungwook; Kim, Youngkyun; Biegalke, Bonita J; Kang, Seongman; Ahn, Kwangseog

2006-06-01

Human cytomegalovirus encodes four unique short (US) region proteins, each of which is independently sufficient for causing the down-regulation of major histocompatibility complex (MHC) class I molecules on the cell surface. This down-regulation enables infected cells to evade recognition by cytotoxic T lymphocytes (CTLs) but makes them vulnerable to lysis by natural killer (NK) cells, which lyse those cells that lack MHC class I molecules. The 22-kDa US3 glycoprotein is able to down-regulate the surface expression of MHC class I molecules by dual mechanisms: direct endoplasmic reticulum retention by physical association and/or tapasin inhibition. The alternative splicing of the US3 gene generates two additional products, including 17-kDa and 3.5-kDa truncated isoforms; however, the functional significance of these isoforms during viral infection is unknown. Here, we describe a novel mode of self-regulation of US3 function that uses the endogenously produced truncated isoform. The truncated isoform itself neither binds to MHC class I molecules nor prevents the full-length US3 from interacting with MHC class I molecules. Instead, the truncated isoform associates with tapasin and competes with full-length US3 for binding to tapasin; thus, it suppresses the action of US3 that causes the disruption of the function of tapasin. Our results indicate that the truncated isoform of the US3 locus acts as a dominant negative regulator of full-length US3 activity. These data reflect the manner in which the virus has developed temporal survival strategies during viral infection against immune surveillance involving both CTLs and NK cells.

Population mixture model for nonlinear telomere dynamics

NASA Astrophysics Data System (ADS)

Itzkovitz, Shalev; Shlush, Liran I.; Gluck, Dan; Skorecki, Karl

2008-12-01

Telomeres are DNA repeats protecting chromosomal ends which shorten with each cell division, eventually leading to cessation of cell growth. We present a population mixture model that predicts an exponential decrease in telomere length with time. We analytically solve the dynamics of the telomere length distribution. The model provides an excellent fit to available telomere data and accounts for the previously unexplained observation of telomere elongation following stress and bone marrow transplantation, thereby providing insight into the nature of the telomere clock.

In vitro generation of helper T cells and suppressor T cells that regulate the cytolytic T lymphocyte response to trinitrophenyl-modified syngeneic cells.

PubMed

Gualde, N; Weinberger, O; Ratnofsky, S; Benacerraf, B; Burakoff, S J

1982-04-01

Helper T cells and suppressor T cells have been generated in vitro that regulate the cytolytic T lymphocyte (CTL) response to trinitrophenyl (TNP)-modified syngeneic cells. B6D2F1 helper cells generated to TNP-modified parental (P1) cells augment the CTL response to those P1-TNP-modified antigens but not to P2-TNP-modified antigens. The generation of these helper T cells requires the presence of splenic adherent cells and these helper T cells are radioresistant. A soluble factor can be obtained from the helper T cell cultures that can also augment the CTL response. The suppressor T cells generated in culture do not demonstrate the specificity observed with the helper T cells; however, they are antigen-dependent in their induction. Whether helper or suppressor activity is obtained depends upon the length of time cells are cultured in vitro.

In vitro generation of helper T cells and suppressor T cells that regulate the cytolytic T lymphocyte response to trinitrophenyl-modified syngeneic cells

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

Gualde, N.; Weinberger, O.; Ratnofsky, S.

1982-04-01

Helper T cells and suppressor T cells have been generated in vitro that regulate the cytolytic T lymphocyte (CTL) response to trinitrophenyl (TNP)-modified syngeneic cells. B6D2F1 helper cells generated to TNP-modified parental (P1) cells augment the CTL response to those P1-TNP-modified antigens but not to P2-TNP-modified antigens. The generation of these helper T cells requires the presence of splenic adherent cells and these helper T cells are radioresistant. A soluble factor can be obtained from the helper T cell cultures that can also augment the CTL response. The suppressor T cells generated in culture do not demonstrate the specificity observedmore » with the helper T cells; however, they are antigen-dependent in their induction. Whether helper or suppressor activity is obtained depends upon the length of time cells are cultured in vitro.« less

Control of scavenger receptor-mediated endocytosis by novel ligands of different length.

PubMed

Guaderrama-Díaz, Margarita; Solís, Carlos F; Velasco-Loyden, Gabriela; Laclette, Juan P; Mas-Oliva, Jaime

2005-03-01

The scavenger receptor recognized as a multiligand family of receptors falls in the group that is internalised through endocytosis. In this report we used several recombinant fragments of the tapeworm protein paramyosin, known to form filamentous dimers that bind collagenous structures as ligands of different length for the class A type I scavenger receptor (SR-AI). While native CHO cells are unresponsive to any of the recombinant fragments, it is shown that CHO cells transfected with this receptor efficiently internalise recombinant fragments that correspond to two thirds of the full-length paramyosin. In contrast, recombinant products corresponding to one-third of the full-length paramyiosin are not internalised. It is also shown that important molecules in the organization of the coated pit, are enriched when the two-thirds long paramyosin fragments were bound and internalised through the SR-AI. Moreover, internalisation of these fragments trigger a classical apoptotic pathway shown by the presence of TUNEL positive cells and the appearance of apoptotic bodies. We report paramyosin as a new ligand for the scavenger receptor and provide evidence supporting the notion that these receptors upon the formation of arrays with length-specific molecules, not only trigger endocytosis but also seem to regulate the synthesis of molecules involved in the organization of coated pits.

Cell-specific targeting by heterobivalent ligands.

PubMed

Josan, Jatinder S; Handl, Heather L; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M; Vagner, Josef; Mash, Eugene A; Hruby, Victor J; Gillies, Robert J

2011-07-20

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach--to specifically target combinations of cell-surface receptors using heteromultivalent ligands ("receptor combination approach"). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle(4), dPhe(7)]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20-50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH(2). Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging.

Cell-Specific Targeting by Heterobivalent Ligands

PubMed Central

Josan, Jatinder S.; Handl, Heather L.; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M.; Vagner, Josef; Mash, Eugene A.; Hruby, Victor J.; Gillies, Robert J.

2012-01-01

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach—to specifically target combinations of cell-surface receptors using heteromultivalent ligands (“receptor combination approach”). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle4, DPhe7]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20–50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH2. Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging. PMID:21639139

Cell Protrusion and Retraction Driven by Fluctuations in Actin Polymerization: A Two-Dimensional Model

PubMed Central

Ryan, Gillian L.; Holz, Danielle; Yamashiro, Sawako; Taniguchi, Daisuke; Watanabe, Naoki; Vavylonis, Dimitrios

2017-01-01

Animal cells that spread onto a surface often rely on actin-rich lamellipodial extensions to execute protrusion. Many cell types recently adhered on a two-dimensional substrate exhibit protrusion and retraction of their lamellipodia, even though the cell is not translating. Traveling waves of protrusion have also been observed, similar to those observed in crawling cells. These regular patterns of protrusion and retraction allow quantitative analysis for comparison to mathematical models. The periodic fluctuations in leading edge position of XTC cells have been linked to excitable actin dynamics using a one-dimensional model of actin dynamics, as a function of arc-length along the cell. In this work we extend this earlier model of actin dynamics into two dimensions (along the arc-length and radial directions of the cell) and include a model membrane that protrudes and retracts in response to the changing number of free barbed ends of actin filaments near the membrane. We show that if the polymerization rate at the barbed ends changes in response to changes in their local concentration at the leading edge and/or the opposing force from the cell membrane, the model can reproduce the patterns of membrane protrusion and retraction seen in experiment. We investigate both Brownian ratchet and switch-like force-velocity relationships between the membrane load forces and actin polymerization rate. The switch-like polymerization dynamics recover the observed patterns of protrusion and retraction as well as the fluctuations in F-actin concentration profiles. The model generates predictions for the behavior of cells after local membrane tension perturbations. PMID:28752950

Promise and problems in relating cellular senescence in vitro to aging in vivo.

PubMed

Rubin, Harry

2002-01-01

According to the 'Hayflick limit', human fetal fibroblasts have a uniform, limited replicative lifespan of about 50 population doublings in cell culture. This concept was extrapolated to diverse cells in the body. It seemed to decrease with the age of the cell donor and, as a form of cell senescence, was thought to underlie the aging process. More discriminating analysis, however, showed that the fibroblasts decayed in a stochastic manner from the time of their explantation, at a rate that increased with the number of population doublings in culture. There was no consistent relation to the age of the donor. Despite the contradictory evidence, the original version of the Hayflick limit retained its general acceptance. Cell senescence was attributed to the absence of telomerase in the fibroblasts, which resulted in shortening of telomeres at each division until they fell below a critical length needed for further division. However, it is well established that stem cells in renewing tissues undergo many more than 50 divisions in a lifetime, without apparent senescence. Contrary to early findings of no telomerase in most tissues, their stem cells retain telomerase and presumably telomere length despite many divisions in vivo. Massive accumulation of lipofuscin granules occurs under stress in long term crowded cultures, but the granules dissipate on subculture or neoplastic transformation. The overall results indicate a critical disjunction between cell senescence in vitro and aging in vivo. By contrast, cell culture has been useful in showing a need for telomere capping in maintaining cell stability and viability. It may also provide information about the biochemical mechanism of lipofuscin production.

Phosphorylation of the Peptidoglycan Synthase PonA1 Governs the Rate of Polar Elongation in Mycobacteria

PubMed Central

Kieser, Karen J.; Baer, Christina E.; Barczak, Amy K.; Meniche, Xavier; Chao, Michael C.; Rego, E. Hesper; Sassetti, Christopher M.; Fortune, Sarah M.; Rubin, Eric J.

2015-01-01

Cell growth and division are required for the progression of bacterial infections. Most rod-shaped bacteria grow by inserting new cell wall along their mid-section. However, mycobacteria, including the human pathogen Mycobacterium tuberculosis, produce new cell wall material at their poles. How mycobacteria control this different mode of growth is incompletely understood. Here we find that PonA1, a penicillin binding protein (PBP) capable of transglycosylation and transpeptidation of cell wall peptidoglycan (PG), is a major governor of polar growth in mycobacteria. PonA1 is required for growth of Mycobacterium smegmatis and is critical for M. tuberculosis during infection. In both cases, PonA1’s catalytic activities are both required for normal cell length, though loss of transglycosylase activity has a more pronounced effect than transpeptidation. Mutations that alter the amount or the activity of PonA1 result in abnormal formation of cell poles and changes in cell length. Moreover, altered PonA1 activity results in dramatic differences in antibiotic susceptibility, suggesting that a balance between the two enzymatic activities of PonA1 is critical for survival. We also find that phosphorylation of a cytoplasmic region of PonA1 is required for normal activity. Mutations in a critical phosphorylated residue affect transglycosylase activity and result in abnormal rates of cell elongation. Together, our data indicate that PonA1 is a central determinant of polar growth in mycobacteria, and its governance of cell elongation is required for robust cell fitness during both host-induced and antibiotic stress. PMID:26114871

An increase in telomere sister chromatid exchange in murine embryonic stem cells possessing critically shortened telomeres

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

Wang, Yisong; Giannone, Richard J; Wu, Jun

Telomerase deficiency leads to a progressive loss of telomeric DNA that eventually triggers cell apoptosis in human primary cells during prolonged growth in culture. Rare survivors can maintain telomere length through either activation of telomerase or recombination-based telomere lengthening, and thus proliferate indefinitely. We have explored the possibility that telomeres may be maintained through telomere sister chromatid exchange (T-SCE) in murine telomere reverse transcriptase-deficient (mTert -/-) splenocytes and ES cells. Because telomerase deficiency leads to gradual loss of telomeric DNA in mTert -/- splenocytes and ES cells and eventually to chromosomes with telomere signal-free ends (SFEs), we examined these cellmore » types for evidence of sister chromatid exchange at telomeres, and observed an increase in T-SCEs only in a subset of mTert -/- splenocytes or ES cells that possessed multiple SFEs. Furthermore, T-SCEs were more often detected in ES cells than in splenocytes that harbored a similar frequency of SFEs. In mTert heterozygous (mTert +/-) ES cells or splenocytes, which are known to exhibit a decrease in average telomere length but no SFEs, no increase in T-SCE was observed. In addition to T-SCE, other genomic rearrangements (i.e., SCE) were also significantly increased in mTert -/- ES cells possessing critically short telomeres, but not in splenocytes. Our results suggest that animals and cell culture differ in their ability to carry out genomic rearrangements as a means of maintaining telomere integrity when telomeres become critically shortened.« less

Reduced Ang2 expression in aging endothelial cells

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

Hohensinner, P.J., E-mail: philipp.hohensinner@meduniwien.ac.at; Ebenbauer, B.; Ludwig Boltzmann Cluster for Cardiovascular Research, Vienna

Aging endothelial cells are characterized by increased cell size, reduced telomere length and increased expression of proinflammatory cytokines. In addition, we describe here that aging reduces the migratory distance of endothelial cells. Furthermore, we observe an increase of the quiescence protein Ang1 and a decrease of the endothelial activation protein Ang2 upon aging. Supplementing Ang2 to aged endothelial cells restored their migratory capacity. We conclude that aging shifts the balance of the Ang1/Ang2 network favouring a quiescent state. Activation of endothelial cells in aging might be necessary to enhance wound healing capacities. -- Highlights: •Endothelial cells display signs of agingmore » before reaching proliferative senescence. •Aging endothelial cells express more angiopoietin 1 and less angiopoietin 2 than young endothelial cells. •Migratory capacity is reduced in aging endothelial cells.« less

Exploring the human mesenchymal stem cell tubule communication network through electron microscopy.

PubMed

Valente, Sabrina; Rossi, Roberta; Resta, Leonardo; Pasquinelli, Gianandrea

2015-04-01

Cells use several mechanisms to transfer information to other cells. In this study, we describe micro/nanotubular connections and exosome-like tubule fragments in multipotent mesenchymal stem cells (MSCs) from human arteries. Scanning and transmission electron microscopy allowed characterization of sinusoidal microtubular projections (700 nm average size, 200 µm average length, with bulging mitochondria and actin microfilaments); short, uniform, variously shaped nanotubular projections (100 nm, bidirectional communication); and tubule fragments (50 nm). This is the first study demonstrating that MSCs from human arteries constitutively interact through an articulate and dynamic tubule network allowing long-range cell to cell communication.

Reduced Ang2 expression in aging endothelial cells.

PubMed

Hohensinner, P J; Ebenbauer, B; Kaun, C; Maurer, G; Huber, K; Wojta, J

2016-06-03

Aging endothelial cells are characterized by increased cell size, reduced telomere length and increased expression of proinflammatory cytokines. In addition, we describe here that aging reduces the migratory distance of endothelial cells. Furthermore, we observe an increase of the quiescence protein Ang1 and a decrease of the endothelial activation protein Ang2 upon aging. Supplementing Ang2 to aged endothelial cells restored their migratory capacity. We conclude that aging shifts the balance of the Ang1/Ang2 network favouring a quiescent state. Activation of endothelial cells in aging might be necessary to enhance wound healing capacities. Copyright © 2016 Elsevier Inc. All rights reserved.

Extracellular guanosine and GTP promote expression of differentiation markers and induce S-phase cell-cycle arrest in human SH-SY5Y neuroblastoma cells.

PubMed

Guarnieri, S; Pilla, R; Morabito, C; Sacchetti, S; Mancinelli, R; Fanò, G; Mariggiò, M A

2009-04-01

SH-SY5Y neuroblastoma cells, a model for studying neuronal differentiation, are able to differentiate into either cholinergic or dopaminergic/adrenergic phenotypes depending on media conditions. Using this system, we asked whether guanosine (Guo) or guanosine-5'-triphosphate (GTP) are able to drive differentiation towards one particular phenotype. Differentiation was determined by evaluating the frequency of cells bearing neurites and assessing neurite length after exposure to different concentrations of Guo or GTP for different durations. After 6 days, 0.3 mM Guo or GTP induced a significant increase in the number of cells bearing neurites and increased neurite length. Western blot analyses confirmed that purines induced differentiation; cells exposed to purines showed increases in the levels of GAP43, MAP2, and tyrosine hydroxylase. Proliferation assays and cytofluorimetric analyses indicated a significant anti-proliferative effect of purines, and a concentration-dependent accumulation of cells in S-phase, starting after 24 h of purine exposure and extending for up to 6 days. A transcriptional profile analysis using gene arrays showed that an up-regulation of cyclin E2/cdk2 evident after 24 h was responsible for S-phase entry, and a concurrent down-regulation of cell-cycle progression-promoting cyclin B1/B2 prevented S-phase exit. In addition, patch-clamp recordings revealed that 0.3 mM Guo or GTP, after 6 day incubation, significantly decreased Na(+) currents. In conclusion, we showed Guo- and GTP-induced cell-cycle arrest in neuroblastoma cells and suggest that this makes these cells more responsive to differentiation processes that favor the dopaminergic/adrenergic phenotype.

The dimensions and characteristics of the subepidermal nerve plexus in human skin--terminal Schwann cells constitute a substantial cell population within the superficial dermis.

PubMed

Reinisch, Christina M; Tschachler, Erwin

2012-03-01

The skin constitutes the largest sensorial organ. Its nervous system consists of different types of afferent nerve fibers which spread out immediately beneath the skin surface to sense temperature, touch and pain. Our aim was to investigate the dimension and topographic relationship of the different nerve fibers of the subepidermal nerve plexus in human hairy skin and to analyze numbers and marker expression of terminal Schwann cells. Nerve fibers and Schwann cells were investigated on dermal sheet preparations and thick sections of skin from various body regions of 10 individuals. The dimension of subepidermal nerve fibers varied between different body sites with highest values in chest skin (100 ± 18 mm/mm(2)) and lowest in posterior forearm skin (53 ± 10 mm/mm(2)). The majority of fibers (85.79%) were unmyelinated, thus representing C-fibers, of which 7.84% were peptidergic. Neurofilament-positive fibers (A-fibers) accounted for 14.21% and fibers positive for both neurofilament and myelin (Aβ-fibers) for only 0.18%. The number of Schwann cells varied in accordance with nerve fiber length from 453 ± 108 on chest skin to 184 ± 58/mm(2) in skin of the posterior forearm. Terminal Schwann cells showed a marker profile comparable to Schwann cells in peripheral nerves with the notable exception of expression of NGFr, NCAM, L1CAM and CD146 on myelinating Schwann cells in the dermis but not in peripheral nerves. Our data show that terminal Schwann cells constitute a substantial cell population within the papillary dermis and that both nerve fiber length and Schwann cell numbers vary considerably between different body sites. Copyright © 2011 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Microtubules and epithem-cell morphogenesis in hydathodes of Pilea cadierei.

PubMed

Galatis, B

1988-12-01

When cell divisions have ceased, the epithem of the hydathodes of Pilea cadierei Gagnep. et Guill. consists of small polyhedral cells exhibiting a meristematic appearance, and completely lacks intercellular spaces. The cortical microtubules in epithem cells exhibit a unique organization: they are not scattered along the whole wall surface but form groups lying at some distance from each other. In sections, from two to eight groups of microtubules can be observed, each lining a wall region averaging between 0.5 and 1.5 μm in length. These groups represent sections of microtubule bundles girdling a major part or the whole of the cell periphery. They are connected to one another by anastomoses, forming a microtubular reticulum. The assembly of microtubule bundles is followed by the appearance of distinct local thickenings in the adjacent wall areas. The cellulose microfibrils in the thickenings are deposited in parallel to the underlying microtubules. Gradually, the vacuolating epithem cells undergo swelling, except for the areas bounded by the wall thickenings. Since the latter, and actually their constituent bundles of cellulose microfibrils, cannot extend in length the differential cell growth results in schizogenous formation of intercellular spaces between contiguous cell walls at their thickened regions. The spaces then broaden and merge to become an extensive intercellular space system. As a result of the above processes, the epithem cells become constricted and finally deeply lobed. The observations show that (i) the cortical microtubules are intimately involved in the morphogenesis of the epithem cells and (ii) the initiation and development of the epithem intercellular spaces is a phenomenon directly related to cell morphogenesis and therefore to the cortical microtubule cytoskeleton. The sites of initiation of these spaces are highly predictable.

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.

Histological study of some Echium vulgare, Pulmonaria officinalis and Symphytum officinale populations.

PubMed

Papp, Nóra; Bencsik, Tímea; Németh, Kitti; Gyergyák, Kinga; Sulc, Alexandra; Farkas, Agnes

2011-10-01

Plants living in different ecological habitats can show significant variability in their histological and phytochemical characters. The main histological features of various populations of three medicinal plants from the Boraginaceae family were studied. Stems, petioles and leaves were investigated by light microscopy in vertical and transverse sections. The outline of the epidermal cells, as well as the shape and cell number of trichomes was studied in leaf surface casts. Differences were measured among the populations of Echium vulgare in the width and height of epidermis cells in the stem, petiole and leaf, as well as in the size of palisade cells in the leaves. Among the populations of Pulmonaria officinalis significant differences were found in the length of trichomes and in the slightly or strongly wavy outline of epidermal radial cell walls. Populations of Symphytum officinale showed variance in the height of epidermal cells in leaves and stems, length of palisade cells and number of intercellular spaces in leaves, and the size of the central cavity in the stem. Boraginaceae bristles were found to be longer in plants in windy/shady habitats as opposed to sunny habitats, both in the leaves and stems ofP. officinalis and S. officinale, which might be connected to varying levels of exposure to wind. Longer epidermal cells were detected in the leaves and stems of both E. vulgare and S. officinale plants living in shady habitats, compared with shorter cells in sunny habitats. Leaf mesophyll cells were shorter in shady habitats as opposed to longer cells in sunny habitats, both in E. vulgare and S. officinale. This combination of histological characters may contribute to the plant's adaptation to various amounts of sunshine. The reported data prove the polymorphism of the studied taxa, as well as their ability to adapt to various ecological circumstances.

Electrochemical cell apparatus having axially distributed entry of a fuel-spent fuel mixture transverse to the cell lengths

DOEpatents

Reichner, P.; Dollard, W.J.

1991-01-08

An electrochemical apparatus is made having a generator section containing axially elongated electrochemical cells, a fresh gaseous feed fuel inlet, a gaseous feed oxidant inlet, and at least one gaseous spent fuel exit channel, where the spent fuel exit channel passes from the generator chamber to combine with the fresh feed fuel inlet at a mixing apparatus, reformable fuel mixture channel passes through the length of the generator chamber and connects with the mixing apparatus, that channel containing entry ports within the generator chamber, where the axis of the ports is transverse to the fuel electrode surfaces, where a catalytic reforming material is distributed near the reformable fuel mixture entry ports. 2 figures.

Sensitive detection of methane at 3.3 μm using an integrating sphere and interband cascade laser

NASA Astrophysics Data System (ADS)

Davis, N. M.; Hodgkinson, J.; Francis, D.; Tatam, R. P.

2016-04-01

Detection of methane at 3.3μm using a DFB Interband Cascade Laser and gold coated integrating sphere is performed. A 10cm diameter sphere with effective path length of 54.5cm was adapted for use as a gas cell. A comparison between this system and one using a 25cm path length single-pass gas cell is made using direct TDLS and methane concentrations between 0 and 1000 ppm. Initial investigations suggest a limit of detection of 1.0ppm for the integrating sphere and 2.2ppm for the single pass gas cell. The system has potential applications in challenging or industrial environments subject to high levels of vibration.

The effect of diffusion induced lattice stress on the open-circuit voltage in silicon solar cells

NASA Technical Reports Server (NTRS)

Weizer, V. G.; Godlewski, M. P.

1984-01-01

It is demonstrated that diffusion induced stresses in low resistivity silicon solar cells can significantly reduce both the open-circuit voltage and collection efficiency. The degradation mechanism involves stress induced changes in both the minority carrier mobility and the diffusion length. Thermal recovery characteristics indicate that the stresses are relieved at higher temperatures by divacancy flow (silicon self diffusion). The level of residual stress in as-fabricated cells was found to be negligible in the cells tested.

Intracavity brillouin scattering from passive Q-spoiling cells.

PubMed

Wick, R V; Guenther, A H

1968-01-01

Stimulated Brillouin scattering from the methanol solvent used in conjunction with cryptocyanine bleachable dye in a ruby laser cavity has been observed at low megawatt output powers. The frequency shifts of the Brillouin scattered radiation produced within the laser cavity are slightly less than frequency shifts produced in an external methanol cell. The Brillouin radiation was eliminated even at output power levels in excess of 250 MW when a 3-mm length cell was used in place of the 25.4-mm commercial cell.

Canine Mesenchymal Stem Cell Potential and the Importance of Dog Breed: Implication for Cell-Based Therapies.

PubMed

Bertolo, Alessandro; Steffen, Frank; Malonzo-Marty, Cherry; Stoyanov, Jivko

2015-01-01

The study of canine bone marrow-derived mesenchymal stem cells (MSCs) has a prominent position in veterinary cell-based applications. Yet the plethora of breeds, their different life spans, and interbreed variations provide unclearness on what can be achieved specifically by such therapies. In this study, we compared a set of morphological, physiological, and genetic markers of MSCs derived from large dog breeds, namely, Border collie, German shepherd, Labrador, Malinois, Golden retriever, and Hovawart. We compared colony-forming units (CFUs) assay, population doubling time (PDT), senescence-associated β-galactosidase (SA-β-gal) activity, telomere length, and gene expression of MSCs, as well as the ability of cells to differentiate to osteogenic, adipogenic, and chondrogenic phenotypes. The influence of the culture media α-MEM, low-glucose DMEM, and high-glucose DMEM, used in cell isolation and expansion, was investigated in the presence and absence of basic fibroblast growth factor (bFGF). Initial cell yield was not affected by culturing medium, but MSCs expanded best in α-MEM supplemented with bFGF. After isolation, the number of MSCs was similar among breeds--as shown by equivalent CFUs--except in the Hovawart samples, which had fivefold less CFU. Telomere lengths were similar among breeds. MSCs divided actively only for 4 weeks in culture (PDT = ∼50 h/division), except Border collie cells divided for a longer time than cells from other groups. The percentage of senescent cells increased linearly in all breeds with time, with a faster rate in German shepherd, Labrador, and Golden retriever. Border collie cells underwent efficient osteogenic differentiation, Hovawart cells performed the best in chondrogenic differentiation, and Labrador cells in both, while German shepherd cells had the lower differentiation potential. MSCs from all breeds preserved the same adipogenic differentiation potential. In conclusion, despite variations, isolated MSCs can be expanded and differentiated in vitro, and all breeds are eligible for MSC-based therapies.

Multifunctional superparamagnetic iron oxide nanoparticles for combined chemotherapy and hyperthermia cancer treatment

NASA Astrophysics Data System (ADS)

Quinto, Christopher A.; Mohindra, Priya; Tong, Sheng; Bao, Gang

2015-07-01

Superparamagnetic iron oxide (SPIO) nanoparticles have the potential for use as a multimodal cancer therapy agent due to their ability to carry anticancer drugs and generate localized heat when exposed to an alternating magnetic field, resulting in combined chemotherapy and hyperthermia. To explore this potential, we synthesized SPIOs with a phospholipid-polyethylene glycol (PEG) coating, and loaded Doxorubicin (DOX) with a 30.8% w/w loading capacity when the PEG length is optimized. We found that DOX-loaded SPIOs exhibited a sustained DOX release over 72 hours where the release kinetics could be altered by the PEG length. In contrast, the heating efficiency of the SPIOs showed minimal change with the PEG length. With a core size of 14 nm, the SPIOs could generate sufficient heat to raise the local temperature to 43 °C, sufficient to trigger apoptosis in cancer cells. Further, we found that DOX-loaded SPIOs resulted in cell death comparable to free DOX, and that the combined effect of DOX and SPIO-induced hyperthermia enhanced cancer cell death in vitro. This study demonstrates the potential of using phospholipid-PEG coated SPIOs for chemotherapy-hyperthermia combinatorial cancer treatment with increased efficacy.Superparamagnetic iron oxide (SPIO) nanoparticles have the potential for use as a multimodal cancer therapy agent due to their ability to carry anticancer drugs and generate localized heat when exposed to an alternating magnetic field, resulting in combined chemotherapy and hyperthermia. To explore this potential, we synthesized SPIOs with a phospholipid-polyethylene glycol (PEG) coating, and loaded Doxorubicin (DOX) with a 30.8% w/w loading capacity when the PEG length is optimized. We found that DOX-loaded SPIOs exhibited a sustained DOX release over 72 hours where the release kinetics could be altered by the PEG length. In contrast, the heating efficiency of the SPIOs showed minimal change with the PEG length. With a core size of 14 nm, the SPIOs could generate sufficient heat to raise the local temperature to 43 °C, sufficient to trigger apoptosis in cancer cells. Further, we found that DOX-loaded SPIOs resulted in cell death comparable to free DOX, and that the combined effect of DOX and SPIO-induced hyperthermia enhanced cancer cell death in vitro. This study demonstrates the potential of using phospholipid-PEG coated SPIOs for chemotherapy-hyperthermia combinatorial cancer treatment with increased efficacy. Electronic supplementary information (ESI) available: Core size distribution; temperature increase for specific absorption rate calculations; effect of DOX loading on zeta potential; combined effect of hyperthermia and free DOX; cell morphology following DOX/hyperthermia treatment. See DOI: 10.1039/c5nr02718g

General working principles of CH3NH3PbX3 perovskite solar cells.

PubMed

Gonzalez-Pedro, Victoria; Juarez-Perez, Emilio J; Arsyad, Waode-Sukmawati; Barea, Eva M; Fabregat-Santiago, Francisco; Mora-Sero, Ivan; Bisquert, Juan

2014-02-12

Organometal halide perovskite-based solar cells have recently realized large conversion efficiency over 15% showing great promise for a new large scale cost-competitive photovoltaic technology. Using impedance spectroscopy measurements we are able to separate the physical parameters of carrier transport and recombination in working devices of the two principal morphologies and compositions of perovskite solar cells, viz. compact thin films of CH3NH3PbI(3-x)Clx and CH3NH3PbI3 infiltrated on nanostructured TiO2. The results show nearly identical spectral characteristics indicating a unique photovoltaic operating mechanism that provides long diffusion lengths (1 μm). Carrier conductivity in both devices is closely matched, so that the most significant differences in performance are attributed to recombination rates. These results highlight the central role of the CH3NH3PbX3 semiconductor absorber in carrier collection and provide a new tool for improved optimization of perovskite solar cells. We report for the first time a measurement of the diffusion length in a nanostructured perovskite solar cell.

Effect of low-energy hydrogen ion implantation on dendritic web silicon solar cells

NASA Technical Reports Server (NTRS)

Rohatgi, A.; Meier, D. L.; Rai-Choudhury, P.; Fonash, S. J.; Singh, R.

1986-01-01

The effect of a low-energy (0.4 keV), short-time (2-min), heavy-dose (10 to the 18th/sq cm) hydrogen ion implant on dendritic web silicon solar cells and material was investigated. Such an implant was observed to improve the cell open-circuit voltage and short-circuit current appreciably for a number of cells. In spite of the low implant energy, measurements of internal quantum efficiency indicate that it is the base of the cell, rather than the emitter, which benefits from the hydrogen implant. This is supported by the observation that the measured minority-carrier diffusion length in the base did not change when the emitter was removed. In some cases, a threefold increase of the base diffusion length was observed after implantation. The effects of the hydrogen implantation were not changed by a thermal stress test at 250 C for 111 h in nitrogen. It is speculated that hydrogen enters the bulk by traveling along dislocations, as proposed recently for edge-defined film-fed growth silicon ribbon.

Data on correlations between T cell subset frequencies and length of partial remission in type 1 diabetes.

PubMed

Narsale, Aditi; Moya, Rosita; Robertson, Hannah Kathryn; Davies, Joanna Davida

2016-09-01

Partial remission in patients newly diagnosed with type 1 diabetes is a period of good glucose control that can last from several weeks to over a year. The clinical significance of the remission period is that patients might be more responsive to immunotherapy if treated within this period. This article provides clinical data that indicates the level of glucose control and insulin-secreting β-cell function of each patient in the study at baseline (within 3 months of diagnosis), and at 3, 6, 9, 12, 18 and 24 months post-baseline. The relative frequency of immune cell subsets in the PBMC of each patient and the association between the frequency of immune cell subsets measured and length of remission is also shown. These data support the findings reported in the accompanying publication, "A pilot study showing associations between frequency of CD4+ memory cell subsets at diagnosis and duration of partial remission in type 1 diabetes" (Moya et al., 2016) [1], where a full interpretation, including biological relevance of the study can be found.

The improved Allium/Vicia root tip micronucleus assay for clastogenicity of environmental pollutants.

PubMed

Ma, T H; Xu, Z; Xu, C; McConnell, H; Rabago, E V; Arreola, G A; Zhang, H

1995-04-01

The meristematic mitotic cells of plant roots are appropriate and efficient cytogenetic materials for the detection of clastogenicity of environmental pollutants, especially for in situ monitoring of water contaminants. Among several cytological endpoints in these fast dividing cells, such as chromosome/chromatid aberrations, sister-chromatid exchanges and micronuclei, the most effective and simplest indicator of cytological damage is micronucleus formation. Although the Allium cepa and Vicia faba root meristem micronucleus assays (Allium/Vicia root MCN) have been used in clastogenicity studies about 12 times by various authors in the last 25 years, there is no report on the comparison of the efficiency of these two plant systems and in different cell populations (meristem and F1) of the root tip as well as under adequate recovery duration. In order to maximize the efficiency of these bioassays, the current study was designed to compare the Allium and the Vicia root MCN assays on the basis of chromosome length, peak sensitivity of the mitotic cells, and the regions of the root tip where the MCN are formed. The total length of the 2n complement of Allium chromosomes is 14.4 microns and the total length of the 2n complement of Vicia is 9.32 microns. The peak sensitivity determined by serial fixation at 12-h intervals after 100 R of X-irradiation is 44 h. The slope of the X-ray dose-response curve of Allium roots derived from the meristematic regions was lower than that derived from cells in the F1 region. Higher efficiency was also demonstrated when the MCN frequencies were scored from the F1 cells in both Allium and Vicia treated with formaldehyde (FA), mitomycin C (MMC), and maleic hydrazide (MH). The results indicated that scoring of MCN frequencies from the F1 cell region of the root tip was more efficient than scoring from the meristematic region. The X-ray linear regression dose-response curves were established in both Allium and Vicia cell systems and the coefficients of correlations, slope values were used to verify the reliability and efficiency of these two plant cell systems. Based on the dose-response slope value of 0.894 for Allium and 0.643 for Vicia, the Allium root MCN was a more efficient test system. The greater sensitivity of the Allium roots is probably due to the greater total length of the diploid complement and the higher number of metacentric chromosomes.(ABSTRACT TRUNCATED AT 400 WORDS)

Stokes injected Raman capillary waveguide amplifier

DOEpatents

Kurnit, Norman A.

1980-01-01

A device for producing stimulated Raman scattering of CO.sub.2 laser radiation by rotational states in a diatomic molecular gas utilizing a Stokes injection signal. The system utilizes a cryogenically cooled waveguide for extending focal interaction length. The waveguide, in conjunction with the Stokes injection signal, reduces required power density of the CO.sub.2 radiation below the breakdown threshold for the diatomic molecular gas. A Fresnel rhomb is employed to circularly polarize the Stokes injection signal and CO.sub.2 laser radiation in opposite circular directions. The device can be employed either as a regenerative oscillator utilizing optical cavity mirrors or as a single pass amplifier. Additionally, a plurality of Raman gain cells can be staged to increase output power magnitude. Also, in the regenerative oscillator embodiment, the Raman gain cell cavity length and CO.sub.2 cavity length can be matched to provide synchronism between mode locked CO.sub.2 pulses and pulses produced within the Raman gain cell.

Spermatozoon structure and motility in the anuran Lepidobatrachus laevis.

PubMed

Waggener, W L; Carroll, E J

1998-02-01

Synthetic human gonadotropin releasing hormone (GnRH) injections were used for induction of spermatozoon release followed by cloacal lavage or mechanical stimulation of sperm release in Lepidobatrachus laevis. Light microscopic observations of Lepidobatrachus laevis spermatozoa indicated an acrosomal segment with a length of 4.1 microm delineated by an indentation, a nuclear region of 12.6 microm in length and a midpiece of 0.87 microm in length. The tail was 54.9 microm long by 1.35 microm wide with two lateral axial fibers and a central undulating membrane. At the electron microscopic level, the unusual tail had two complete axonemes that emanated from the distal centriole. The tail also contained two axial fibers 77 nm in diameter medial to the axonemes and was connected by an undulating membrane. An unusual accessory cell adherent to the head of the spermatozoon was noted in freshly obtained suspensions of spermatozoa. Spermatozoa with the accessory cell were motile and a subsequent loss of motility was correlated with the shedding of the accessory cell.

Evaluation of Shape and Textural Features from CT as Prognostic Biomarkers in Non-small Cell Lung Cancer.

PubMed

Bianconi, Francesco; Fravolini, Mario Luca; Bello-Cerezo, Raquel; Minestrini, Matteo; Scialpi, Michele; Palumbo, Barbara

2018-04-01

We retrospectively investigated the prognostic potential (correlation with overall survival) of 9 shape and 21 textural features from non-contrast-enhanced computed tomography (CT) in patients with non-small-cell lung cancer. We considered a public dataset of 203 individuals with inoperable, histologically- or cytologically-confirmed NSCLC. Three-dimensional shape and textural features from CT were computed using proprietary code and their prognostic potential evaluated through four different statistical protocols. Volume and grey-level run length matrix (GLRLM) run length non-uniformity were the only two features to pass all four protocols. Both features correlated negatively with overall survival. The results also showed a strong dependence on the evaluation protocol used. Tumour volume and GLRLM run-length non-uniformity from CT were the best predictor of survival in patients with non-small-cell lung cancer. We did not find enough evidence to claim a relationship with survival for the other features. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Biochemical basis for the biological clock

NASA Technical Reports Server (NTRS)

Morre, D. James; Chueh, Pin-Ju; Pletcher, Jake; Tang, Xiaoyu; Wu, Lian-Ying; Morre, Dorothy M.

2002-01-01

NADH oxidases at the external surface of plant and animal cells (ECTO-NOX proteins) exhibit stable and recurring patterns of oscillations with potentially clock-related, entrainable, and temperature-compensated period lengths of 24 min. To determine if ECTO-NOX proteins might represent the ultradian time keepers (pacemakers) of the biological clock, COS cells were transfected with cDNAs encoding tNOX proteins having a period length of 22 min or with C575A or C558A cysteine to alanine replacements having period lengths of 36 or 42 min. Here we demonstrate that such transfectants exhibited 22, 36, or 40 to 42 h circadian patterns in the activity of glyceraldehyde-3-phosphate dehydrogenase, a common clock-regulated protein, in addition to the endogenous 24 h circadian period length. The fact that the expression of a single oscillatory ECTO-NOX protein determines the period length of a circadian biochemical marker (60 X the ECTO-NOX period length) provides compelling evidence that ECTO-NOX proteins are the biochemical ultradian drivers of the cellular biological clock.

Development of Pulsating Tubules with Chiral Inversion

DTIC Science & Technology

2013-09-21

assembly of rod–coil block molecules. These supramolecular ligands agglutinated effectively specific bacterial cells through carbohydrate-mediated...transduction and cause Jurkat cells to release 100 to 300 times as much IL-2 as lectin-stimulated normal human peripheral blood lymphocytes. In our...were found to regulate T cell activation. The lengths as well as stability of the protein-coated supramolecular nanofibers could be manipulated by a

PbCl2-tuned inorganic cubic CsPbBr3(Cl) perovskite solar cells with enhanced electron lifetime, diffusion length and photovoltaic performance

NASA Astrophysics Data System (ADS)

Li, Bo; Zhang, Yanan; Zhang, Luyuan; Yin, Longwei

2017-08-01

Inorganic CsPbBr3 perovskite is arousing great interest following after organic-inorganic hybrid halide perovskites, and is found as a good candidate for photovoltaic devices for its prominent photoelectric property and stability. Herein, we for the first time report on PbCl2-tuned inorganic Cl-doped CsPbBr3(Cl) perovskite solar cells with adjustable crystal structure and Cl doping for enhanced carrier lifetime, extraction rate and photovoltaic performance. The effect of PbCl2 on the morphologies, structures, optical, and photovoltaic performance of CsPbBr3 perovskite solar cells is investigated systemically. Compared with orthorhombic CsPbBr3, cubic CsPbBr3 demonstrates a significant improvement for electron lifetime (from 6.7 ns to 12.3 ns) and diffusion length (from 69 nm to 197 nm), as well as the enhanced electron extraction rate from CsPbBr3 to TiO2. More importantly, Cl doping benefits the further enhancement of carrier lifetime (14.3 ns) and diffusion length (208 nm). The Cl doped cubic CsPbBr3(Cl) perovskite solar cell exhibits a Jsc of 8.47 mA cm-2 and a PCE of 6.21%, superior to that of pure orthorhombic CsPbBr3 (6.22 mA cm-2 and 3.78%). The improvement of photovoltaic performance can be attributed to enhanced carrier lifetime, diffusion length and extraction rates, as well as suppressed nonradiative recombination.

Top down and bottom up engineering of bone.

PubMed

Knothe Tate, Melissa L

2011-01-11

The goal of this retrospective article is to place the body of my lab's multiscale mechanobiology work in context of top-down and bottom-up engineering of bone. We have used biosystems engineering, computational modeling and novel experimental approaches to understand bone physiology, in health and disease, and across time (in utero, postnatal growth, maturity, aging and death, as well as evolution) and length scales (a single bone like a femur, m; a sample of bone tissue, mm-cm; a cell and its local environment, μm; down to the length scale of the cell's own skeleton, the cytoskeleton, nm). First we introduce the concept of flow in bone and the three calibers of porosity through which fluid flows. Then we describe, in the context of organ-tissue, tissue-cell and cell-molecule length scales, both multiscale computational models and experimental methods to predict flow in bone and to understand the flow of fluid as a means to deliver chemical and mechanical cues in bone. Addressing a number of studies in the context of multiple length and time scales, the importance of appropriate boundary conditions, site specific material parameters, permeability measures and even micro-nanoanatomically correct geometries are discussed in context of model predictions and their value for understanding multiscale mechanobiology of bone. Insights from these multiscale computational modeling and experimental methods are providing us with a means to predict, engineer and manufacture bone tissue in the laboratory and in the human body. Copyright © 2010 Elsevier Ltd. All rights reserved.

Longitudinal growth of skeletal myotubes in vitro in a new horizontal mechanical cell stimulator

NASA Technical Reports Server (NTRS)

Vandenburgh, Herman H.; Karlisch, Patricia

1989-01-01

A tissue-culture model system for growing skeletal-muscle cells under more dynamic conditions than found in normal tissue-culture environments is described. A computerized device presented allows mechanical stimulation of the cell's substratum by 300 to 400 pct in length in the horizontal plane. Cell growth rates and skeletal-muscle organogenesis are stimulated in this in vitro system. It is noted that longitudinal myotube growth observed is accompanied by increased rates of cell proliferation and myoblast fusion. Prestretching the collagen-coated substratum before cell plating is shown to lead to increased cell proliferation, myotube orientation, and longitudinal myotube growth. The effects of substratum stretching on myogenesis in the model system are also assessed and attributed to alterations in the cell's extracellular matrix.

Oncogenicity of L-type amino-acid transporter 1 (LAT1) revealed by targeted gene disruption in chicken DT40 cells: LAT1 is a promising molecular target for human cancer therapy

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

Ohkawa, Mayumi; Ohno, Yoshiya; Masuko, Kazue

Highlights: {yields} We established LAT1 amino-acid transporter-disrupted DT40 cells. {yields} LAT1-disrupted cells showed slow growth and lost the oncogenicity. {yields} siRNA and mAb inhibited human tumor growth in vitro and in vivo. {yields} LAT1 is a promising target molecule for cancer therapy. -- Abstract: L-type amino-acid transporter 1 (LAT1) is the first identified light chain of CD98 molecule, disulfide-linked to a heavy chain of CD98. Following cDNA cloning of chicken full-length LAT1, we have constructed targeting vectors for the disruption of chicken LAT1 gene from genomic DNA of chicken LAT1 consisting of 5.4 kb. We established five homozygous LAT1-disrupted (LAT1{supmore » -/-}) cell clones, derived from a heterozygous LAT1{sup +/-} clone of DT40 chicken B cell line. Reactivity of anti-chicken CD98hc monoclonal antibody (mAb) with LAT1{sup -/-} DT40 cells was markedly decreased compared with that of wild-type DT40 cells. All LAT1{sup -/-} cells were deficient in L-type amino-acid transporting activity, although alternative-splice variant but not full-length mRNA of LAT1 was detected in these cells. LAT1{sup -/-} DT40 clones showed outstandingly slow growth in liquid culture and decreased colony-formation capacity in soft agar compared with wild-type DT40 cells. Cell-cycle analyses indicated that LAT1{sup -/-} DT40 clones have prolonged cell-cycle phases compared with wild-type or LAT1{sup +/-} DT40 cells. Knockdown of human LAT1 by small interfering RNAs resulted in marked in vitro cell-growth inhibition of human cancer cells, and in vivo tumor growth of HeLa cells in athymic mice was significantly inhibited by anti-human LAT1 mAb. All these results indicate essential roles of LAT1 in the cell proliferation and occurrence of malignant phenotypes and that LAT1 is a promising candidate as a molecular target of human cancer therapy.« less

The C-terminal domain of TRPV4 is essential for plasma membrane localization.

PubMed

Becker, Daniel; Müller, Margarethe; Leuner, Kristina; Jendrach, Marina

2008-02-01

Many members of the TRP superfamily oligomerize in the ER before trafficking to the plasma membrane. For membrane localization of the non-selective cation channel TRPV4 specific domains in the N-terminus are required, but the role of the C-terminus in the oligomerization and trafficking process has been not determined until now. Therefore, the localization of recombinant TRPV4 in two cell models was analyzed: HaCaT keratinocytes that express TRPV4 endogenously were compared to CHO cells that are devoid of endogenous TRPV4. When deletions were introduced in the C-terminal domain three states of TRPV4 localization were defined: a truncated TRPV4 protein of 855 amino acids was exported to the plasma membrane like the full-length channel (871 aa) and was also functional. Mutants with a length of 828 to 844 amino acids remained in the ER of CHO cells, but in HaCaT cells plasma membrane localization was partially rescued by oligomerization with endogenous TRPV4. This was confirmed by coexpression of recombinant full-length TRPV4 together with these deletion mutants, which resulted in an almost complete plasma membrane localization of both proteins and significant FRET in the plasma membrane and the ER. All deletions upstream of amino acid 828 resulted in total ER retention that could not rescued by coexpression with the full-length protein. However, these deletion mutants did not impair export of full-length TRPV4, implying that no oligomerization took place. These data indicate that the C-terminus of TRPV4 is required for oligomerization, which takes place in the ER and precedes plasma membrane trafficking.

Actin filaments, stereocilia, and hair cells of the bird cochlea. V. How the staircase pattern of stereociliary lengths is generated.

PubMed

Tilney, L G; Tilney, M S; Cotanche, D A

1988-02-01

The stereocilia on each hair cell are arranged into rows of ascending height, resulting in what we refer to as a "staircase-like" profile. At the proximal end of the cochlea the length of the tallest row of stereocilia in the staircase is 1.5 micron, with the shortest row only 0.3 micron. As one proceeds towards the distal end of the cochlea the length of the stereocilia progressively increases so that at the extreme distal end the length of the tallest row of the staircase is 5.5 micron and the shortest row is 2 micron. During development hair cells form their staircases in four phases of growth separated from each other by developmental time. First, stereocilia sprout from the apical surfaces of the hair cells (8-10-d embryos). Second (10-12-d embryos), what will be the longest row of the staircase begins to elongate. As the embryo gets older successive rows of stereocilia initiate elongation. Thus the staircase is set up by the sequential initiation of elongation of stereociliary rows located at increased distances from the row that began elongation. Third (12-17-d embryos), all the stereocilia in the newly formed staircase elongate until those located on the first step of the staircase have reached the prescribed length. In the final phase (17-d embryos to hatchlings) there is a progressive cessation of elongation beginning with the shortest step and followed by taller and taller rows with the tallest step stopping last. Thus, to obtain a pattern of stereocilia in rows of increasing height what transpires are progressive go signals followed by a period when all the stereocilia grow and ending with progressive stop signals. We discuss how such a sequence could be controlled.

Programming Cell Adhesion for On-Chip Sequential Boolean Logic Functions.

PubMed

Qu, Xiangmeng; Wang, Shaopeng; Ge, Zhilei; Wang, Jianbang; Yao, Guangbao; Li, Jiang; Zuo, Xiaolei; Shi, Jiye; Song, Shiping; Wang, Lihua; Li, Li; Pei, Hao; Fan, Chunhai

2017-08-02

Programmable remodelling of cell surfaces enables high-precision regulation of cell behavior. In this work, we developed in vitro constructed DNA-based chemical reaction networks (CRNs) to program on-chip cell adhesion. We found that the RGD-functionalized DNA CRNs are entirely noninvasive when interfaced with the fluidic mosaic membrane of living cells. DNA toehold with different lengths could tunably alter the release kinetics of cells, which shows rapid release in minutes with the use of a 6-base toehold. We further demonstrated the realization of Boolean logic functions by using DNA strand displacement reactions, which include multi-input and sequential cell logic gates (AND, OR, XOR, and AND-OR). This study provides a highly generic tool for self-organization of biological systems.

Study of living single cells in culture: automated recognition of cell behavior.

PubMed

Bodin, P; Papin, S; Meyer, C; Travo, P

1988-07-01

An automated system capable of analyzing the behavior, in real time, of single living cells in culture, in a noninvasive and nondestructive way, has been developed. A large number of cell positions in single culture dishes were recorded using a computer controlled, robotized microscope. During subsequent observations, binary images obtained from video image analysis of the microscope visual field allowed the identification of the recorded cells. These cells could be revisited automatically every few minutes. Long-term studies of the behavior of cells make possible the analysis of cellular locomotary and mitotic activities as well as determination of cell shape (chosen from a defined library) for several hours or days in a fully automated way with observations spaced up to 30 minutes. Short-term studies of the behavior of cells permit the study, in a semiautomatic way, of acute effects of drugs (5 to 15 minutes) on changes of surface area and length of cells.

Nanostructured Polyaniline Coating on ITO Glass Promotes the Neurite Outgrowth of PC 12 Cells by Electrical Stimulation.

PubMed

Wang, Liping; Huang, Qianwei; Wang, Jin-Ye

2015-11-10

A conducting polymer polyaniline (PANI) with nanostructure was synthesized on indium tin oxide (ITO) glass. The effect of electrical stimulation on the proliferation and the length of neurites of PC 12 cells was investigated. The dynamic protein adsorption on PANI and ITO surfaces in a cell culture medium was also compared with and without electrical stimulation. The adsorbed proteins were characterized using SDS-PAGE. A PANI coating on ITO surface was shown with 30-50 nm spherical nanostructure. The number of PC 12 cells was significantly greater on the PANI/ITO surface than on ITO and plate surfaces after cell seeding for 24 and 36 h. This result confirmed that the PANI coating is nontoxic to PC 12 cells. The electrical stimulation for 1, 2, and 4 h significantly enhanced the cell numbers for both PANI and ITO conducting surfaces. Moreover, the application of electrical stimulation also improved the neurite outgrowth of PC 12 cells, and the number of PC 12 cells with longer neurite lengths increased obviously under electrical stimulation for the PANI surface. From the mechanism, the adsorption of DMEM proteins was found to be enhanced by electrical stimulation for both PANI/ITO and ITO surfaces. A new band 2 (around 37 kDa) was observed from the collected adsorbed proteins when PC 12 cells were cultured on these surfaces, and culturing PC 12 cells also seemed to increase the amount of band 1 (around 90 kDa). When immersing PANI/ITO and ITO surfaces in a DMEM medium without a cell culture, the number of band 3 (around 70 kDa) and band 4 (around 45 kDa) proteins decreased compared to that of PC 12 cell cultured surfaces. These results are valuable for the design and improvement of the material performance for neural regeneration.

Predicting cell viability within tissue scaffolds under equiaxial strain: multi-scale finite element model of collagen-cardiomyocytes constructs.

PubMed

Elsaadany, Mostafa; Yan, Karen Chang; Yildirim-Ayan, Eda

2017-06-01

Successful tissue engineering and regenerative therapy necessitate having extensive knowledge about mechanical milieu in engineered tissues and the resident cells. In this study, we have merged two powerful analysis tools, namely finite element analysis and stochastic analysis, to understand the mechanical strain within the tissue scaffold and residing cells and to predict the cell viability upon applying mechanical strains. A continuum-based multi-length scale finite element model (FEM) was created to simulate the physiologically relevant equiaxial strain exposure on cell-embedded tissue scaffold and to calculate strain transferred to the tissue scaffold (macro-scale) and residing cells (micro-scale) upon various equiaxial strains. The data from FEM were used to predict cell viability under various equiaxial strain magnitudes using stochastic damage criterion analysis. The model validation was conducted through mechanically straining the cardiomyocyte-encapsulated collagen constructs using a custom-built mechanical loading platform (EQUicycler). FEM quantified the strain gradients over the radial and longitudinal direction of the scaffolds and the cells residing in different areas of interest. With the use of the experimental viability data, stochastic damage criterion, and the average cellular strains obtained from multi-length scale models, cellular viability was predicted and successfully validated. This methodology can provide a great tool to characterize the mechanical stimulation of bioreactors used in tissue engineering applications in providing quantification of mechanical strain and predicting cellular viability variations due to applied mechanical strain.

Epitope topography controls bioactivity in supramolecular nanofibers

PubMed Central

Sur, Shantanu; Tantakitti, Faifan; Matson, John B.; Stupp, Samuel I.

2015-01-01

Incorporating bioactivity into artificial scaffolds using peptide epitopes present in the extracellular matrix (ECM) is a well-known approach. A common strategy has involved epitopes that provide cells with attachment points and external cues through interaction with integrin receptors. Although a variety of bioactive sequences have been identified so far, less is known about their optimal display in a scaffold. We report here on the use of self-assembled peptide amphiphile (PA) nanofiber matrices to investigate the impact of spatial presentation of the fibronectin derived epitope RGDS on cell response. Using one, three, or five glycine residues, RGDS epitopes were systematically spaced out from the surface of the rigid nanofibers. We found that cell morphology was strongly affected by the separation of the epitope from the nanofiber surface, with the longest distance yielding the most cell-spreading, bundling of actin filaments, and a round-to-polygonal transformation of cell shape. Cell response to this type of epitope display was also accompanied with activated integrin-mediated signaling and formation of stronger adhesions between cells and substrate. Interestingly, unlike length, changing the molecular flexibility of the linker had minimal influence on cell behavior on the substrate for reasons that remain poorly understood. The use in this study of high persistence length nanofibers rather than common flexible polymers allows us to conclude that epitope topography at the nanoscale structure of a scaffold influences its bioactive properties independent of epitope density and mechanical properties. PMID:25745558

Merkel Cell Polyomavirus Large T Antigen Has Growth-Promoting and Inhibitory Activities

PubMed Central

Cheng, Jingwei; Rozenblatt-Rosen, Orit; Paulson, Kelly G.; Nghiem, Paul

2013-01-01

Merkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer. In at least 80% of all MCC, Merkel cell polyomavirus (MCPyV) DNA has undergone clonal integration into the host cell genome, and most tumors express the MCPyV large and small T antigens. In all cases of MCC reported to date, the integrated MCPyV genome has undergone mutations in the large T antigen. These mutations result in expression of a truncated large T antigen that retains the Rb binding or LXCXE motif but deletes the DNA binding and helicase domains. However, the transforming functions of full-length and truncated MCPyV large T antigen are unknown. We compared the transforming activities of full-length, truncated, and alternatively spliced 57kT forms of MCPyV large T antigen. MCPyV large T antigen could bind to Rb but was unable to bind to p53. Furthermore, MCPyV-truncated large T antigen was more effective than full-length and 57kT large T antigen in promoting the growth of human and mouse fibroblasts. In contrast, expression of the MCPyV large T antigen C-terminal 100 residues could inhibit the growth of several different cell types. These data imply that the deletion of the C terminus of MCPyV large T antigen found in MCC serves not only to disrupt viral replication but also results in the loss of a distinct growth-inhibitory function intrinsic to this region. PMID:23514892

Fluorescence image-guided photodynamic therapy of cancer cells using a scanning fiber endoscope

NASA Astrophysics Data System (ADS)

Woldetensae, Mikias H.; Kirshenbaum, Mark R.; Kramer, Greg M.; Zhang, Liang; Seibel, Eric J.

2013-03-01

A scanning fiber endoscope (SFE) and the cancer biomarker 5-aminolevulinic acid (5-ALA) were used to fluorescently detect and destroy superficial cancerous lesions, while experimenting with different dosimetry levels for concurrent or sequential imaging and laser therapy. The 1.6-mm diameter SFE was used to fluorescently image a confluent monolayer of A549 human lung cancer cells from culture, previously administered with 5 mM solution of 5-ALA for 4 hours. Twenty hours after therapy, cell cultures were stained to distinguish between living and dead cells using a laser scanning confocal microscope. To determine relative dosimetry for photodynamic therapy (PDT), 405-nm laser illumination was varied from 1 to 5 minutes with power varying from 5 to 18 mW, chosen to compare equal amounts of energy delivered to the cell culture. The SFE produced 500-line images of fluorescence at 15 Hz using the red detection channel centered at 635 nm. The results show that PDT of A549 cancer cell monolayers using 405nm light for imaging and 5-ALAinduced PpIX therapy was possible using the same SFE system. Increased duration and power of laser illumination produced an increased area of cell death upon live/dead staining. The ultrathin and flexible SFE was able to direct PDT using wide-field fluorescence imaging of a monolayer of cultured cancer cells after uptaking 5-ALA. The correlation between light intensity and duration of PDT was measured. Increased length of exposure and decreased light intensity yields larger areas of cell death than decreased length of exposure with increased light intensity.

Atomistic Tight-Binding Theory Applied to Structural and Optical Properties of Silicon Nanodisks

NASA Astrophysics Data System (ADS)

Sukkabot, Worasak

2018-05-01

The use of ultrathin crystalline silicon (c-Si) wafers in solar cells necessitates a highly effective light absorber to compensate for poor light absorption. One route to overcoming this problem is to use a periodic array of Si nanodisks on ultrathin c-Si. In the present manuscript, we numerically investigate the effects of the geometrical parameters of the Si nanodisks, including disk diameter (D) and length (L), on the structural and optical properties, using atomistic tight-binding theory. These computations confirm that the electronic structure and optical properties are sensitive to the structural parameters. As the disk diameter and length increase, the single-electron energies decrease, and the single-hole energies increase. These calculations also reveal that, because of the quantum confinement effect, the optical band gaps gradually decrease independently of the increasing disk diameter and length. The optical spectra can be tuned across the visible region by varying the disk diameter and length, which is a useful feature for optimizing light absorption in solar cell applications. As the disk diameter and length increased, the optical intensities also increased; however, the atomistic electron-hole interactions and ground electron-hole wave function overlap progressively decreased. The ground electron-hole wave function overlap, Stokes shift, and fine structure splitting decreased as the disk diameter and length were increased. Thus, Si nanodisks with a large diameter and length might be a suitable candidate source of entangled photons. The Si nanodisks in this study also show promise for applications to solar cells based on ultrathin c-Si wafers.

Application of a roughness-length representation to parameterize energy loss in 3-D numerical simulations of large rivers

NASA Astrophysics Data System (ADS)

Sandbach, S. D.; Lane, S. N.; Hardy, R. J.; Amsler, M. L.; Ashworth, P. J.; Best, J. L.; Nicholas, A. P.; Orfeo, O.; Parsons, D. R.; Reesink, A. J. H.; Szupiany, R. N.

2012-12-01

Recent technological advances in remote sensing have enabled investigation of the morphodynamics and hydrodynamics of large rivers. However, measuring topography and flow in these very large rivers is time consuming and thus often constrains the spatial resolution and reach-length scales that can be monitored. Similar constraints exist for computational fluid dynamics (CFD) studies of large rivers, requiring maximization of mesh- or grid-cell dimensions and implying a reduction in the representation of bedform-roughness elements that are of the order of a model grid cell or less, even if they are represented in available topographic data. These "subgrid" elements must be parameterized, and this paper applies and considers the impact of roughness-length treatments that include the effect of bed roughness due to "unmeasured" topography. CFD predictions were found to be sensitive to the roughness-length specification. Model optimization was based on acoustic Doppler current profiler measurements and estimates of the water surface slope for a variety of roughness lengths. This proved difficult as the metrics used to assess optimal model performance diverged due to the effects of large bedforms that are not well parameterized in roughness-length treatments. However, the general spatial flow patterns are effectively predicted by the model. Changes in roughness length were shown to have a major impact upon flow routing at the channel scale. The results also indicate an absence of secondary flow circulation cells in the reached studied, and suggest simpler two-dimensional models may have great utility in the investigation of flow within large rivers.

Spatial distribution of the capacity to initiate a secondary embryo in the 32-cell embryo of Xenopus laevis.

PubMed

Kageura, H

1990-12-01

To examine the spatial distribution of dorsal determinants in the early embryos of Xenopus laevis, individual cells from the 32-cell embryo were transplanted into the same tier of the ventral side of a synchronous recipient. Their abilities to initiate a secondary embryo were measured by the incidence of secondary embryos and by the length of the secondary axis relative to the primary embryo. The ability was found to be localized in all cells (A1, B1, C1, and D1) of the dorsal most column and in the vegetal cells (C2 and D2) of the dorsolateral column. Transplanted C1 (subequatorial) cells caused the highest incidence of a secondary embryo and the average relative length of the secondary embryo was also greatest. Effectiveness decreased in the order: D1, B1, D2, C2, and A1. When these results were compared with Dale and Slack's fate map of the 32-cell embryo, it was concluded that the distribution of dorsal determinants is unique and does not coincide with the prospective regions for any tissues, though it is somewhat similar to the prospective region of dorsal endoderm or notochord. From these results it seems that dorsal determinants do not determine a particular tissue in an embryo but rather the "dorsal" region of an embryo.

Linking macroscopic with microscopic neuroanatomy using synthetic neuronal populations.

PubMed

Schneider, Calvin J; Cuntz, Hermann; Soltesz, Ivan

2014-10-01

Dendritic morphology has been shown to have a dramatic impact on neuronal function. However, population features such as the inherent variability in dendritic morphology between cells belonging to the same neuronal type are often overlooked when studying computation in neural networks. While detailed models for morphology and electrophysiology exist for many types of single neurons, the role of detailed single cell morphology in the population has not been studied quantitatively or computationally. Here we use the structural context of the neural tissue in which dendritic trees exist to drive their generation in silico. We synthesize the entire population of dentate gyrus granule cells, the most numerous cell type in the hippocampus, by growing their dendritic trees within their characteristic dendritic fields bounded by the realistic structural context of (1) the granule cell layer that contains all somata and (2) the molecular layer that contains the dendritic forest. This process enables branching statistics to be linked to larger scale neuroanatomical features. We find large differences in dendritic total length and individual path length measures as a function of location in the dentate gyrus and of somatic depth in the granule cell layer. We also predict the number of unique granule cell dendrites invading a given volume in the molecular layer. This work enables the complete population-level study of morphological properties and provides a framework to develop complex and realistic neural network models.

Linking Macroscopic with Microscopic Neuroanatomy Using Synthetic Neuronal Populations

PubMed Central

Schneider, Calvin J.; Cuntz, Hermann; Soltesz, Ivan

2014-01-01

Dendritic morphology has been shown to have a dramatic impact on neuronal function. However, population features such as the inherent variability in dendritic morphology between cells belonging to the same neuronal type are often overlooked when studying computation in neural networks. While detailed models for morphology and electrophysiology exist for many types of single neurons, the role of detailed single cell morphology in the population has not been studied quantitatively or computationally. Here we use the structural context of the neural tissue in which dendritic trees exist to drive their generation in silico. We synthesize the entire population of dentate gyrus granule cells, the most numerous cell type in the hippocampus, by growing their dendritic trees within their characteristic dendritic fields bounded by the realistic structural context of (1) the granule cell layer that contains all somata and (2) the molecular layer that contains the dendritic forest. This process enables branching statistics to be linked to larger scale neuroanatomical features. We find large differences in dendritic total length and individual path length measures as a function of location in the dentate gyrus and of somatic depth in the granule cell layer. We also predict the number of unique granule cell dendrites invading a given volume in the molecular layer. This work enables the complete population-level study of morphological properties and provides a framework to develop complex and realistic neural network models. PMID:25340814

Phosphatidylinositol 4,5-Bisphosphate Clusters the Cell Adhesion Molecule CD44 and Assembles a Specific CD44-Ezrin Heterocomplex, as Revealed by Small Angle Neutron Scattering*

PubMed Central

Chen, Xiaodong; Khajeh, Jahan Ali; Ju, Jeong Ho; Gupta, Yogesh K.; Stanley, Christopher B.; Do, Changwoo; Heller, William T.; Aggarwal, Aneel K.; Callaway, David J. E.; Bu, Zimei

2015-01-01

The cell adhesion molecule CD44 regulates diverse cellular functions, including cell-cell and cell-matrix interaction, cell motility, migration, differentiation, and growth. In cells, CD44 co-localizes with the membrane-cytoskeleton adapter protein Ezrin that links the CD44 assembled receptor signaling complexes to the cytoskeletal actin network, which organizes the spatial and temporal localization of signaling events. Here we report that the cytoplasmic tail of CD44 (CD44ct) is largely disordered. Upon binding to the signaling lipid phosphatidylinositol 4,5-bisphosphate (PIP2), CD44ct clusters into aggregates. Further, contrary to the generally accepted model, CD44ct does not bind directly to the FERM domain of Ezrin or to the full-length Ezrin but only forms a complex with FERM or with the full-length Ezrin in the presence of PIP2. Using contrast variation small angle neutron scattering, we show that PIP2 mediates the assembly of a specific heterotetramer complex of CD44ct with Ezrin. This study reveals the role of PIP2 in clustering CD44 and in assembling multimeric CD44-Ezrin complexes. We hypothesize that polyvalent electrostatic interactions are responsible for the assembly of CD44 clusters and the multimeric PIP2-CD44-Ezrin complexes. PMID:25572402

Phosphatidylinositol 4,5-bisphosphate clusters the cell adhesion molecule CD44 and assembles a specific CD44-Ezrin heterocomplex, as revealed by small angle neutron scattering.

PubMed

Chen, Xiaodong; Khajeh, Jahan Ali; Ju, Jeong Ho; Gupta, Yogesh K; Stanley, Christopher B; Do, Changwoo; Heller, William T; Aggarwal, Aneel K; Callaway, David J E; Bu, Zimei

2015-03-06

The cell adhesion molecule CD44 regulates diverse cellular functions, including cell-cell and cell-matrix interaction, cell motility, migration, differentiation, and growth. In cells, CD44 co-localizes with the membrane-cytoskeleton adapter protein Ezrin that links the CD44 assembled receptor signaling complexes to the cytoskeletal actin network, which organizes the spatial and temporal localization of signaling events. Here we report that the cytoplasmic tail of CD44 (CD44ct) is largely disordered. Upon binding to the signaling lipid phosphatidylinositol 4,5-bisphosphate (PIP2), CD44ct clusters into aggregates. Further, contrary to the generally accepted model, CD44ct does not bind directly to the FERM domain of Ezrin or to the full-length Ezrin but only forms a complex with FERM or with the full-length Ezrin in the presence of PIP2. Using contrast variation small angle neutron scattering, we show that PIP2 mediates the assembly of a specific heterotetramer complex of CD44ct with Ezrin. This study reveals the role of PIP2 in clustering CD44 and in assembling multimeric CD44-Ezrin complexes. We hypothesize that polyvalent electrostatic interactions are responsible for the assembly of CD44 clusters and the multimeric PIP2-CD44-Ezrin complexes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Regional heterogeneity of endothelial cells in the porcine vortex vein system.

PubMed

Tan, Priscilla Ern Zhi; Yu, Paula K; Cringle, Stephen J; Morgan, William H; Yu, Dao-Yi

2013-09-01

The aim of this study was to investigate whether region-dependent endothelial heterogeneity is present within the porcine vortex vein system. The superior temporal vortex vein in young adult pig eyes were dissected out and cannulated. The intact vortex vein system down to the choroidal veins was then perfused with labels for f-actin and nucleic acid. The endothelial cells within the choroidal veins, pre-ampulla, anterior portion of the ampulla, mid-ampulla, posterior portion of the ampulla, post-ampulla, intra-scleral canal and the extra-ocular vortex vein regions were studied in detail using a confocal microscopy technique. The endothelial cell and nuclei length, width, area and perimeter were measured and compared between the different regions. Significant regional differences in the endothelial cell and nuclei length, width, area and perimeter were observed throughout the porcine vortex vein system. Most notably, very narrow and elongated endothelia were found in the post-ampulla region. A lack of smooth muscle cells was noted in the ampulla region compared to other regions. Heterogeneity in endothelial cell morphology is present throughout the porcine vortex vein system and there is a lack of smooth muscle cells in the ampulla region. This likely reflects the highly varied haemodynamic conditions and potential blood flow control mechanisms in different regions of the vortex vein system. Copyright © 2013 Elsevier Inc. All rights reserved.

Use of a corrugated surface to enhance radiation tolerance in a GaAs solar cell

NASA Technical Reports Server (NTRS)

Leon, Rosa P.; Piszczor, Michael F., Jr.

1985-01-01

The use of a corrugated surface on a GaAs solar cell and its effects on radiation resistance were studied. A compute code was developed to determine the performance of the cell for various geometric parameters. The large optical absorption coefficient of GaAs allows grooves to be only 4-5 micrometers deep. Using accepted material parameters for GaAs solar cells the theoretical performances were compared for various corrugated cells before and after minority carrier diffusion length degradation. The total power output was maximized for both n(+)/p and p(+)/n cells. Optimum values of 1.0-1.5 and 5.0 micrometers for groove and ridge widths respectively were determined.

Precise mass determination of single cell with cantilever-based microbiosensor system.

PubMed

Łabędź, Bogdan; Wańczyk, Aleksandra; Rajfur, Zenon

2017-01-01

Having determined the mass of a single cell of brewer yeast Saccharomyces cerevisiae by means of a microcantilever-based biosensor Cantisens CSR-801 (Concentris, Basel, Switzerland), it was found that its dry mass is 47,65 ± 1,05 pg. Found to be crucial in this mass determination was the cell position along the length of the cantilever. Moreover, calculations including cells positions on the cantilever provide a threefold better degree of accuracy than those which assume uniform mass distribution. We have also examined the influence of storage time on the single cell mass. Our results show that after 6 months there is an increase in the average mass of a single yeast cell.

Fine mapping and identification of a novel locus qGL12.2 control grain length in wild rice (Oryza rufipogon Griff.).

PubMed

Qi, Lan; Ding, Yingbin; Zheng, Xiaoming; Xu, Rui; Zhang, Lizhen; Wang, Yanyan; Wang, Xiaoning; Zhang, Lifang; Cheng, Yunlian; Qiao, Weihua; Yang, Qingwen

2018-04-19

A wild rice QTL qGL12.2 for grain length was fine mapped to an 82-kb interval in chromosome 12 containing six candidate genes and none was reported previously. Grain length is an important trait for yield and commercial value in rice. Wild rice seeds have a very slender shape and have many desirable genes that have been lost in cultivated rice during domestication. In this study, we identified a quantitative trait locus, qGL12.2, which controls grain length in wild rice. First, a wild rice chromosome segment substitution line, CSSL41, was selected that has longer glume and grains than does the Oryza sativa indica cultivar, 9311. Next, an F 2 population was constructed from a cross between CSSL41 and 9311. Using the next-generation sequencing combined with bulked-segregant analysis and F 3 recombinants analysis, qGL12.2 was finally fine mapped to an 82-kb interval in chromosome 12. Six candidate genes were found, and no reported grain length genes were found in this interval. Using scanning electron microscopy, we found that CSSL41 cells are significantly longer than those of 9311, but there is no difference in cell widths. These data suggest that qGL12.2 is a novel gene that controls grain cell length in wild rice. Our study provides a new genetic resource for rice breeding and a starting point for functional characterization of the wild rice GL gene.

Carbon flux to growth or polyhydroxyalkanoate synthesis under microaerophilic conditions is affected by fatty acid chain-length in Pseudomonas putida LS46.

PubMed

Blunt, Warren; Dartiailh, Christopher; Sparling, Richard; Gapes, Daniel; Levin, David B; Cicek, Nazim

2018-05-24

Economical production of medium-chain length polyhydroxyalkanoates (mcl-PHA) is dependent on efficient cultivation processes. This work describes growth and mcl-PHA synthesis characteristics of Pseudomonas putida LS46 when grown on medium-chain length fatty acids (octanoic acid) and lower-cost long-chain fatty acids (LCFAs, derived from hydrolyzed canola oil) in microaerophilic environments. Growth on octanoic acid ceased when the oxygen uptake rate was limited by the oxygen transfer rate, and mcl-PHA accumulated to 61.9% of the cell dry mass. From LCFAs, production of non-PHA cell mass continued at a rate of 0.36 g L -1  h -1 under oxygen-limited conditions, while mcl-PHA accumulated simultaneously to 31% of the cell dry mass. The titer of non-PHA cell mass from LCFAs at 14 h post-inoculation was double that obtained from octanoic acid in bioreactors operated with identical feeding and aeration conditions. While the productivity for octanoic acid was higher by 14 h, prolonged cultivation on LCFAs achieved similar productivity but with twice the PHA titer. Simultaneous co-feeding of each substrate demonstrated the continued cell growth under microaerophilic conditions characteristic of LCFAs, and the resulting polymer was dominant in C8 monomers. Furthermore, co-feeding resulted in improved PHA titer and volumetric productivity compared to either substrate individually. These results suggest that LCFAs improve growth of P. putida in oxygen-limited environments and could reduce production costs since more non-PHA cell mass, the cellular factories required to produce mcl-PHA and the most oxygen-intensive cellular process, can be produced for a given oxygen transfer rate.

Protective dendritic cell responses against listeriosis induced by the short form of the deubiquitinating enzyme CYLD are inhibited by full-length CYLD.

PubMed

Wurm, Rebecca; Just, Sissy; Wang, Xu; Wex, Katharina; Schmid, Ursula; Blanchard, Nicolas; Waisman, Ari; Schild, Hans-Jörg; Deckert, Martina; Naumann, Michael; Schlüter, Dirk; Nishanth, Gopala

2015-05-01

The deubiquitinating enzyme CYLD is an important tumor suppressor and inhibitor of immune responses. In contrast to full-length CYLD, the immunological function of the naturally occurring short splice variant of CYLD (sCYLD) is insufficiently described. Previously, we showed that DCs, which lack full-length CYLD but express sCYLD, exhibit augmented NF-κB and DC activation. To explore the function of sCYLD in infection, we investigated whether DC-specific sCYLD regulates the pathogenesis of listeriosis. Upon Listeria monocytogenes infection of CD11c-Cre Cyld(ex7/8 fl/fl) mice, infection of CD8α(+) DCs, which are crucial for the establishment of listeriosis in the spleen, was not affected. However, NF-κB activity of CD11c-Cre Cyld(ex7/8 fl/fl) DCs was increased, while activation of ERK and p38 was normal. In addition, CD11c-Cre Cyld(ex7/8 fl/fl) DCs produced more TNF, IL-10, and IL-12 upon infection, which led to enhanced stimulation of IFN-γ-producing NK cells. In addition CD11c-Cre Cyld(ex7/8 fl/fl) DCs presented Listeria Ag more efficiently to CD8(+) T cells resulting in a stronger pathogen-specific CD8(+) T-cell proliferation and more IFN-γ production. Collectively, the improved innate and adaptive immunity and survival during listeriosis identify the DC-specific FL-CYLD/sCYLD balance as a potential target to modulate NK-cell and Ag-specific CD8(+) T-cell responses. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Phenotypic integration in an extended phenotype: among-individual variation in nest-building traits of the alfalfa leafcutting bee (Megachile rotundata).

PubMed

Royauté, Raphaël; Wilson, Elisabeth S; Helm, Bryan R; Mallinger, Rachel E; Prasifka, Jarrad; Greenlee, Kendra J; Bowsher, Julia H

2018-03-02

Structures such as nests and burrows are an essential component of many organisms' life-cycle and require a complex sequence of behaviours. Because behaviours can vary consistently among individuals and be correlated with one another, we hypothesized that these structures would (1) show evidence of among-individual variation, (2) be organized into distinct functional modules and (3) show evidence of trade-offs among functional modules due to limits on energy budgets. We tested these hypotheses using the alfalfa leafcutting bee, Megachile rotundata, a solitary bee and important crop pollinator. Megachile rotundata constructs complex nests by gathering leaf materials to form a linear series of cells in pre-existing cavities. In this study, we examined variation in the following nest construction traits: reproduction (number of cells per nest and nest length), nest protection (cap length and number of leaves per cap), cell construction (cell size and number of leaves per cell) and cell provisioning (cell mass) from 60 nests. We found a general decline in investment in cell construction and provisioning with each new cell built. In addition, we found evidence for both repeatability and plasticity in cell provisioning with little evidence for trade-offs among traits. Instead, most traits were positively, albeit weakly, correlated (r ~ 0.15), and traits were loosely organized into covarying modules. Our results show that individual differences in nest construction are detectable at a level similar to that of other behavioural traits and that these traits are only weakly integrated. This suggests that nest components are capable of independent evolutionary trajectories. © 2018 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2018 European Society For Evolutionary Biology.