Methylene Blue Protects Astrocytes against Glucose Oxygen Deprivation by Improving Cellular Respiration

PubMed Central

Roy Choudhury, Gourav; Winters, Ali; Rich, Ryan M.; Ryou, Myoung-Gwi; Gryczynski, Zygmunt; Yuan, Fang; Yang, Shao-Hua; Liu, Ran

2015-01-01

Astrocytes outnumber neurons and serve many metabolic and trophic functions in the mammalian brain. Preserving astrocytes is critical for normal brain function as well as for protecting the brain against various insults. Our previous studies have indicated that methylene blue (MB) functions as an alternative electron carrier and enhances brain metabolism. In addition, MB has been shown to be protective against neurodegeneration and brain injury. In the current study, we investigated the protective role of MB in astrocytes. Cell viability assays showed that MB treatment significantly protected primary astrocytes from oxygen-glucose deprivation (OGD) & reoxygenation induced cell death. We also studied the effect of MB on cellular oxygen and glucose metabolism in primary astrocytes following OGD-reoxygenation injury. MB treatment significantly increased cellular oxygen consumption, glucose uptake and ATP production in primary astrocytes. In conclusion our study demonstrated that MB protects astrocytes against OGD-reoxygenation injury by improving astrocyte cellular respiration. PMID:25848957

Activation of cellular death programs associated with immunosenescence-like phenotype in TPPII knockout mice

PubMed Central

Huai, Jisen; Firat, Elke; Nil, Ahmed; Million, Daniele; Gaedicke, Simone; Kanzler, Benoit; Freudenberg, Marina; van Endert, Peter; Kohler, Gabriele; Pahl, Heike L.; Aichele, Peter; Eichmann, Klaus; Niedermann, Gabriele

2008-01-01

The giant cytosolic protease tripeptidyl peptidase II (TPPII) has been implicated in the regulation of proliferation and survival of malignant cells, particularly lymphoma cells. To address its functions in normal cellular and systemic physiology we have generated TPPII-deficient mice. TPPII deficiency activates cell type-specific death programs, including proliferative apoptosis in several T lineage subsets and premature cellular senescence in fibroblasts and CD8+ T cells. This coincides with up-regulation of p53 and dysregulation of NF-κB. Prominent degenerative alterations at the organismic level were a decreased lifespan and symptoms characteristic of immunohematopoietic senescence. These symptoms include accelerated thymic involution, lymphopenia, impaired proliferative T cell responses, extramedullary hematopoiesis, and inflammation. Thus, TPPII is important for maintaining normal cellular and systemic physiology, which may be relevant for potential therapeutic applications of TPPII inhibitors. PMID:18362329

Characteristics of functional enrichment and gene expression level of human putative transcriptional target genes.

PubMed

Osato, Naoki

2018-01-19

Transcriptional target genes show functional enrichment of genes. However, how many and how significantly transcriptional target genes include functional enrichments are still unclear. To address these issues, I predicted human transcriptional target genes using open chromatin regions, ChIP-seq data and DNA binding sequences of transcription factors in databases, and examined functional enrichment and gene expression level of putative transcriptional target genes. Gene Ontology annotations showed four times larger numbers of functional enrichments in putative transcriptional target genes than gene expression information alone, independent of transcriptional target genes. To compare the number of functional enrichments of putative transcriptional target genes between cells or search conditions, I normalized the number of functional enrichment by calculating its ratios in the total number of transcriptional target genes. With this analysis, native putative transcriptional target genes showed the largest normalized number of functional enrichments, compared with target genes including 5-60% of randomly selected genes. The normalized number of functional enrichments was changed according to the criteria of enhancer-promoter interactions such as distance from transcriptional start sites and orientation of CTCF-binding sites. Forward-reverse orientation of CTCF-binding sites showed significantly higher normalized number of functional enrichments than the other orientations. Journal papers showed that the top five frequent functional enrichments were related to the cellular functions in the three cell types. The median expression level of transcriptional target genes changed according to the criteria of enhancer-promoter assignments (i.e. interactions) and was correlated with the changes of the normalized number of functional enrichments of transcriptional target genes. Human putative transcriptional target genes showed significant functional enrichments. Functional enrichments were related to the cellular functions. The normalized number of functional enrichments of human putative transcriptional target genes changed according to the criteria of enhancer-promoter assignments and correlated with the median expression level of the target genes. These analyses and characters of human putative transcriptional target genes would be useful to examine the criteria of enhancer-promoter assignments and to predict the novel mechanisms and factors such as DNA binding proteins and DNA sequences of enhancer-promoter interactions.

NAD(H) and NADP(H) Redox Couples and Cellular Energy Metabolism.

PubMed

Xiao, Wusheng; Wang, Rui-Sheng; Handy, Diane E; Loscalzo, Joseph

2018-01-20

The nicotinamide adenine dinucleotide (NAD + )/reduced NAD + (NADH) and NADP + /reduced NADP + (NADPH) redox couples are essential for maintaining cellular redox homeostasis and for modulating numerous biological events, including cellular metabolism. Deficiency or imbalance of these two redox couples has been associated with many pathological disorders. Recent Advances: Newly identified biosynthetic enzymes and newly developed genetically encoded biosensors enable us to understand better how cells maintain compartmentalized NAD(H) and NADP(H) pools. The concept of redox stress (oxidative and reductive stress) reflected by changes in NAD(H)/NADP(H) has increasingly gained attention. The emerging roles of NAD + -consuming proteins in regulating cellular redox and metabolic homeostasis are active research topics. The biosynthesis and distribution of cellular NAD(H) and NADP(H) are highly compartmentalized. It is critical to understand how cells maintain the steady levels of these redox couple pools to ensure their normal functions and simultaneously avoid inducing redox stress. In addition, it is essential to understand how NAD(H)- and NADP(H)-utilizing enzymes interact with other signaling pathways, such as those regulated by hypoxia-inducible factor, to maintain cellular redox homeostasis and energy metabolism. Additional studies are needed to investigate the inter-relationships among compartmentalized NAD(H)/NADP(H) pools and how these two dinucleotide redox couples collaboratively regulate cellular redox states and cellular metabolism under normal and pathological conditions. Furthermore, recent studies suggest the utility of using pharmacological interventions or nutrient-based bioactive NAD + precursors as therapeutic interventions for metabolic diseases. Thus, a better understanding of the cellular functions of NAD(H) and NADP(H) may facilitate efforts to address a host of pathological disorders effectively. Antioxid. Redox Signal. 28, 251-272.

Cellular Sites of Immunologic Unresponsiveness*

PubMed Central

Chiller, Jacques M.; Habicht, Gail S.; Weigle, William O.

1970-01-01

The reconstitution of the immune response of lethally irradiated mice to human γ-globulin is dependent on the synergistic action of bone marrow with thymus cells. Immunologic unresponsiveness appears to involve a functional defect at each of these cellular levels, inasmuch as neither bone marrow nor thymus cells from unresponsive donors are capable of demonstrating synergism in combination with their normal counterpart. PMID:4192271

Aggregation of polyglutamine-expanded ataxin-3 sequesters its specific interacting partners into inclusions: Implication in a loss-of-function pathology

PubMed Central

Yang, Hui; Li, Jing-Jing; Liu, Shuai; Zhao, Jian; Jiang, Ya-Jun; Song, Ai-Xin; Hu, Hong-Yu

2014-01-01

Expansion of polyglutamine (polyQ) tract may cause protein misfolding and aggregation that lead to cytotoxicity and neurodegeneration, but the underlying mechanism remains to be elucidated. We applied ataxin-3 (Atx3), a polyQ tract-containing protein, as a model to study sequestration of normal cellular proteins. We found that the aggregates formed by polyQ-expanded Atx3 sequester its interacting partners, such as P97/VCP and ubiquitin conjugates, into the protein inclusions through specific interactions both in vitro and in cells. Moreover, this specific sequestration impairs the normal cellular function of P97 in down-regulating neddylation. However, expansion of polyQ tract in Atx3 does not alter the conformation of its surrounding regions and the interaction affinities with the interacting partners, although it indeed facilitates misfolding and aggregation of the Atx3 protein. Thus, we propose a loss-of-function pathology for polyQ diseases that sequestration of the cellular essential proteins via specific interactions into inclusions by the polyQ aggregates causes dysfunction of the corresponding proteins, and consequently leads to neurodegeneration. PMID:25231079

Cellular Mechanisms of Somatic Stem Cell Aging

PubMed Central

Jung, Yunjoon

2014-01-01

Tissue homeostasis and regenerative capacity rely on rare populations of somatic stem cells endowed with the potential to self-renew and differentiate. During aging, many tissues show a decline in regenerative potential coupled with a loss of stem cell function. Cells including somatic stem cells have evolved a series of checks and balances to sense and repair cellular damage to maximize tissue function. However, during aging the mechanisms that protect normal cell function begin to fail. In this review, we will discuss how common cellular mechanisms that maintain tissue fidelity and organismal lifespan impact somatic stem cell function. We will highlight context-dependent changes and commonalities that define aging, by focusing on three age-sensitive stem cell compartments: blood, neural, and muscle. Understanding the interaction between extrinsic regulators and intrinsic effectors that operate within different stem cell compartments is likely to have important implications for identifying strategies to improve health span and treat age-related degenerative diseases. PMID:24439814

PNUTS functions as a proto-oncogene by sequestering PTEN

PubMed Central

Kavela, Sridhar; Shinde, Swapnil R; Ratheesh, Raman; Viswakalyan, Kotapalli; Bashyam, Murali D; Gowrishankar, Swarnalata; Vamsy, Mohana; Pattnaik, Sujit; Rao, Subramanyeshwar; Sastry, Regulagadda A; Srinivasulu, Mukta; Chen, Junjie; Maddika, Subbareddy

2012-01-01

PTEN is a well-defined tumor suppressor gene that antagonizes the PI3K/Akt pathway to regulate a multitude of cellular processes such as survival, growth, motility, invasiveness and angiogenesis. While the functions of PTEN have been studied extensively, the regulation of its activity during normal and disease conditions still remains incompletely understood. In this study, we identified the protein phosphatase-1 nuclear targeting subunit PNUTS (PPP1R10) as a PTEN associated protein. PNUTS directly interacted with the lipid-binding domain (C2 domain) of PTEN and sequestered it in the nucleus. Depletion of PNUTS leads to increased apoptosis and reduced cellular proliferation in a PTEN-dependent manner. PNUTS expression was elevated in certain cancers compared to matched normal tissues. Collectively, our studies reveal PNUTS as a novel PTEN regulator and a likely oncogene. PMID:23117887

Mechanics of the Nucleus

PubMed Central

Lammerding, Jan

2015-01-01

The nucleus is the distinguishing feature of eukaryotic cells. Until recently, it was often considered simply as a unique compartment containing the genetic information of the cell and associated machinery, without much attention to its structure and mechanical properties. This article provides compelling examples that illustrate how specific nuclear structures are associated with important cellular functions, and how defects in nuclear mechanics can cause a multitude of human diseases. During differentiation, embryonic stem cells modify their nuclear envelope composition and chromatin structure, resulting in stiffer nuclei that reflect decreased transcriptional plasticity. In contrast, neutrophils have evolved characteristic lobulated nuclei that increase their physical plasticity, enabling passage through narrow tissue spaces in their response to inflammation. Research on diverse cell types further demonstrates how induced nuclear deformations during cellular compression or stretch can modulate cellular function. Pathological examples of disturbed nuclear mechanics include the many diseases caused by mutations in the nuclear envelope proteins lamin A/C and associated proteins, as well as cancer cells that are often characterized by abnormal nuclear morphology. In this article, we will focus on determining the functional relationship between nuclear mechanics and cellular (dys-)function, describing the molecular changes associated with physiological and pathological examples, the resulting defects in nuclear mechanics, and the effects on cellular function. New insights into the close relationship between nuclear mechanics and cellular organization and function will yield a better understanding of normal biology and will offer new clues into therapeutic approaches to the various diseases associated with defective nuclear mechanics. PMID:23737203

Discoidin domain receptor 2 (DDR2) regulates proliferation of endochondral cells in mice

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

Kawai, Ikuma; Hisaki, Tomoka; Sugiura, Koji

2012-10-26

Highlights: Black-Right-Pointing-Pointer Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase. Black-Right-Pointing-Pointer DDR2 regulates cell proliferation, cell adhesion, migration, and extracellular matrix remodeling. Black-Right-Pointing-Pointer We produced in vitro and in vivo model to better understand the role of DDR2. Black-Right-Pointing-Pointer DDR2 might play an inhibitory role in the proliferation of chondrocyte. -- Abstract: Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase that is activated by fibrillar collagens. DDR2 regulates cell proliferation, cell adhesion, migration, and extracellular matrix remodeling. The decrement of endogenous DDR2 represses osteoblastic marker gene expression and osteogenic differentiation in murine preosteoblastic cells, but themore » functions of DDR2 in chondrogenic cellular proliferation remain unclear. To better understand the role of DDR2 signaling in cellular proliferation in endochondral ossification, we inhibited Ddr2 expression via the inhibitory effect of miRNA on Ddr2 mRNA (miDdr2) and analyzed the cellular proliferation and differentiation in the prechondrocyte ATDC5 cell lines. To investigate DDR2's molecular role in endochondral cellular proliferation in vivo, we also produced transgenic mice in which the expression of truncated, kinase dead (KD) DDR2 protein is induced, and evaluated the DDR2 function in cellular proliferation in chondrocytes. Although the miDdr2-transfected ATDC5 cell lines retained normal differentiation ability, DDR2 reduction finally promoted cellular proliferation in proportion to the decreasing ratio of Ddr2 expression, and it also promoted earlier differentiation to cartilage cells by insulin induction. The layer of hypertrophic chondrocytes in KD Ddr2 transgenic mice was not significantly thicker than that of normal littermates, but the layer of proliferative chondrocytes in KD-Ddr2 transgenic mice was significantly thicker than that of normal littermates. Taken together, our data demonstrated that DDR2 might play a local and essential role in the proliferation of chondrocytes.« less

Picornaviruses and nuclear functions: targeting a cellular compartment distinct from the replication site of a positive-strand RNA virus

PubMed Central

Flather, Dylan; Semler, Bert L.

2015-01-01

The compartmentalization of DNA replication and gene transcription in the nucleus and protein production in the cytoplasm is a defining feature of eukaryotic cells. The nucleus functions to maintain the integrity of the nuclear genome of the cell and to control gene expression based on intracellular and environmental signals received through the cytoplasm. The spatial separation of the major processes that lead to the expression of protein-coding genes establishes the necessity of a transport network to allow biomolecules to translocate between these two regions of the cell. The nucleocytoplasmic transport network is therefore essential for regulating normal cellular functioning. The Picornaviridae virus family is one of many viral families that disrupt the nucleocytoplasmic trafficking of cells to promote viral replication. Picornaviruses contain positive-sense, single-stranded RNA genomes and replicate in the cytoplasm of infected cells. As a result of the limited coding capacity of these viruses, cellular proteins are required by these intracellular parasites for both translation and genomic RNA replication. Being of messenger RNA polarity, a picornavirus genome can immediately be translated upon entering the cell cytoplasm. However, the replication of viral RNA requires the activity of RNA-binding proteins, many of which function in host gene expression, and are consequently localized to the nucleus. As a result, picornaviruses disrupt nucleocytoplasmic trafficking to exploit protein functions normally localized to a different cellular compartment from which they translate their genome to facilitate efficient replication. Furthermore, picornavirus proteins are also known to enter the nucleus of infected cells to limit host-cell transcription and down-regulate innate antiviral responses. The interactions of picornavirus proteins and host-cell nuclei are extensive, required for a productive infection, and are the focus of this review. PMID:26150805

Premature aging/senescence in cancer cells facing therapy: good or bad?

PubMed

Gonzalez, Llilians Calvo; Ghadaouia, Sabrina; Martinez, Aurélie; Rodier, Francis

2016-02-01

Normal and cancer cells facing their demise following exposure to radio-chemotherapy can actively participate in choosing their subsequent fate. These programmed cell fate decisions include true cell death (apoptosis-necroptosis) and therapy-induced cellular senescence (TIS), a permanent "proliferative arrest" commonly portrayed as premature cellular aging. Despite a permanent loss of proliferative potential, senescent cells remain viable and are highly bioactive at the microenvironment level, resulting in a prolonged impact on tissue architecture and functions. Cellular senescence is primarily documented as a tumor suppression mechanism that prevents cellular transformation. In the context of normal tissues, cellular senescence also plays important roles in tissue repair, but contributes to age-associated tissue dysfunction when senescent cells accumulate. Theoretically, in multi-step cancer progression models, cancer cells have already bypassed cellular senescence during their immortalization step (see hallmarks of cancer). It is then perhaps surprising to find that cancer cells often retain the ability to undergo TIS, or premature aging. This occurs because cellular senescence results from multiple signalling pathways, some retained in cancer cells, aiming to prevent cell cycle progression in damaged cells. Since senescent cancer cells persist after therapy and secrete an array of cytokines and growth factors that can modulate the tumor microenvironment, these cells may have beneficial and detrimental effects regarding immune modulation and survival of remaining proliferation-competent cancer cells. Similarly, while normal cells undergoing senescence are believed to remain indefinitely growth arrested, whether this is true for senescent cancer cells remains unclear, raising the possibility that these cells may represent a reservoir for cancer recurrence after treatment. This review discusses our current knowledge on cancer cell senescence and highlight questions that must be addressed to fully understand the beneficial and detrimental impacts of cellular senescence during cancer therapy.

Quantification of endocytosis using a folate functionalized silica hollow nanoshell platform

PubMed Central

Sandoval, Sergio; Mendez, Natalie; Alfaro, Jesus G.; Yang, Jian; Aschemeyer, Sharraya; Liberman, Alex; Trogler, William C.; Kummel, Andrew C.

2015-01-01

Abstract. A quantification method to measure endocytosis was designed to assess cellular uptake and specificity of a targeting nanoparticle platform. A simple N-hydroxysuccinimide ester conjugation technique to functionalize 100-nm hollow silica nanoshell particles with fluorescent reporter fluorescein isothiocyanate and folate or polyethylene glycol (PEG) was developed. Functionalized nanoshells were characterized using scanning electron microscopy and transmission electron microscopy and the maximum amount of folate functionalized on nanoshell surfaces was quantified with UV-Vis spectroscopy. The extent of endocytosis by HeLa cervical cancer cells and human foreskin fibroblast (HFF-1) cells was investigated in vitro using fluorescence and confocal microscopy. A simple fluorescence ratio analysis was developed to quantify endocytosis versus surface adhesion. Nanoshells functionalized with folate showed enhanced endocytosis by cancer cells when compared to PEG functionalized nanoshells. Fluorescence ratio analyses showed that 95% of folate functionalized silica nanoshells which adhered to cancer cells were endocytosed, while only 27% of PEG functionalized nanoshells adhered to the cell surface and underwent endocytosis when functionalized with 200 and 900  μg, respectively. Additionally, the endocytosis of folate functionalized nanoshells proved to be cancer cell selective while sparing normal cells. The developed fluorescence ratio analysis is a simple and rapid verification/validation method to quantify cellular uptake between datasets by using an internal control for normalization. PMID:26315280

Connexin 43 and ATP-sensitive potassium channels crosstalk: a missing link in hypoxia/ischemia stress.

PubMed

Ahmad Waza, Ajaz; Ahmad Bhat, Shabir; Ul Hussain, Mahboob; Ganai, Bashir A

2018-02-01

Connexin 43 (Cx43) is a gap junction protein expressed in various tissues and organs of vertebrates. Besides functioning as a gap junction, Cx43 also regulates diverse cellular processes like cell growth and differentiation, cell migration, cell survival, etc. Cx43 is critical for normal cardiac functioning and is therefore abundantly expressed in cardiomyocytes. On the other hand, ATP-sensitive potassium (K ATP ) channels are metabolic sensors converting metabolic changes into electrical activity. These channels are important in maintaining the neurotransmitter release, smooth muscle relaxation, cardiac action potential repolarization, normal physiology of cellular repolarization, insulin secretion and immune function. Cx43 and K ATP channels are part of the same signaling pathway, regulating cell survival during stress conditions and ischemia/hypoxia preconditioning. However, the underlying molecular mechanism for their combined role in ischemia/hypoxia preconditioning is largely unknown. The current review focuses on understanding the molecular mechanism responsible for the coordinated role of Cx43 and K ATP channel protein in protecting cardiomyocytes against ischemia/hypoxia stress.

PKCλ/ι signaling-a common node for normal cellular development and breast oncogenesis.

PubMed

Paul, Arindam; Paul, Soumen

2015-01-01

We recently demonstrated that PKCλ/ι signaling is an important contributor to breast cancer development. Strikingly, PKCλ/ι signaling is also important to balance self-renewal versus differentiation in pluripotent stem cells and is essential for embryonic development. This commentary highlights some key functions of PKCλ/ι signaling that are integral to both normal development and cancer progression.

Regulation of cell function by methionine oxidation and reduction

PubMed Central

Hoshi, Toshinori; Heinemann, Stefan H

2001-01-01

Reactive oxygen species (ROS) are generated during normal cellular activity and may exist in excess in some pathophysiological conditions, such as inflammation or reperfusion injury. These molecules oxidize a variety of cellular constituents, but sulfur-containing amino acid residues are especially susceptible. While reversible cysteine oxidation and reduction is part of well-established signalling systems, the oxidation and the enzymatically catalysed reduction of methionine is just emerging as a novel molecular mechanism for cellular regulation. Here we discuss how the oxidation of methionine to methionine sulfoxide in signalling proteins such as ion channels affects the function of these target proteins. Methionine sulfoxide reductase, which reduces methionine sulfoxide to methionine in a thioredoxin-dependent manner, is therefore not only an enzyme important for the repair of age- or degenerative disease-related protein modifications. It is also a potential missing link in the post-translational modification cycle involved in the specific oxidation and reduction of methionine residues in cellular signalling proteins, which may give rise to activity-dependent plastic changes in cellular excitability. PMID:11179387

A Modified Protocol for the Isolation of Primary Human Hepatocytes with Improved Viability and Function from Normal and Diseased Human Liver.

PubMed

Bartlett, David C; Newsome, Philip N

2017-01-01

Successful hepatocyte isolation is critical for continued development of cellular transplantation. However, most tissue available for research is from diseased liver and the results of hepatocyte isolation from such tissue are inferior compared to normal tissue. Here we describe a modified method, combining the use of Liberase and N-acetylcysteine (NAC), for the isolation of primary human hepatocytes with high viability from normal and diseased liver.

Vertebrate Presynaptic Active Zone Assembly: a Role Accomplished by Diverse Molecular and Cellular Mechanisms.

PubMed

Torres, Viviana I; Inestrosa, Nibaldo C

2018-06-01

Among all the biological systems in vertebrates, the central nervous system (CNS) is the most complex, and its function depends on specialized contacts among neurons called synapses. The assembly and organization of synapses must be exquisitely regulated for a normal brain function and network activity. There has been a tremendous effort in recent decades to understand the molecular and cellular mechanisms participating in the formation of new synapses and their organization, maintenance, and regulation. At the vertebrate presynapses, proteins such as Piccolo, Bassoon, RIM, RIM-BPs, CAST/ELKS, liprin-α, and Munc13 are constant residents and participate in multiple and dynamic interactions with other regulatory proteins, which define network activity and normal brain function. Here, we review the function of these active zone (AZ) proteins and diverse factors involved in AZ assembly and maintenance, with an emphasis on axonal trafficking of precursor vesicles, protein homo- and hetero-oligomeric interactions as a mechanism of AZ trapping and stabilization, and the role of F-actin in presynaptic assembly and its modulation by Wnt signaling.

Requirement for the Murine Zinc Finger Protein ZFR in Perigastrulation Growth and Survival

PubMed Central

Meagher, Madeleine J.; Braun, Robert E.

2001-01-01

The transition from preimplantation to postimplantation development leads to the initiation of complex cellular differentiation and morphogenetic movements, a dramatic decrease in cell cycle length, and a commensurate increase in the size of the embryo. Accompanying these changes is the need for the transfer of nutrients from the mother to the embryo and the elaboration of sophisticated genetic networks that monitor genomic integrity and the homeostatic control of cellular growth, differentiation, and programmed cell death. To determine the function of the murine zinc finger protein ZFR in these events, we generated mice carrying a null mutation in the gene encoding it. Homozygous mutant embryos form normal-appearing blastocysts that implant and initiate the process of gastrulation. Mutant embryos form mesoderm but they are delayed in their development and fail to form normal anterior embryonic structures. Loss of ZFR function leads to both an increase in programmed cell death and a decrease in mitotic index, especially in the region of the distal tip of the embryonic ectoderm. Mutant embryos also have an apparent reduction in apical vacuoles in the columnar visceral endoderm cells in the extraembryonic region. Together, these cellular phenotypes lead to a dramatic development delay and embryonic death by 8 to 9 days of gestation, which are independent of p53 function. PMID:11283266

Normal and leukaemic human haemopoietic cells in diffusion chamber. A morphological and functional CFU-C study.

PubMed

Laurent, M; Clémancey-Marcille, G; Hollard, D

1980-03-01

Leukaemic human bone marrow and peripheral blood cells were cultured for 25 d in diffusion chambers implanted into cyclophosphamide treated mice. Normal bone marrow cells were cultured simultaneously. These cells were studied both morphologically and functionally (CFU-C). The leukaemic cells behaved heterogeneously, 2 groups being distinguishable in accordance with their initial in vitro growth pattern (1: no growth or microcluster growth. 2: macrocluster growth). Group I showed progressive cellular death with a diminution of granulocytic progenitors and the appearance of a predominantly macrophagic population. This behaviour resembled that of the control group. The initial microcluster growth pattern remained identical throughout the entire culture period. Group 2, after considerable cellular death up to d 5, showed an explosive proliferation of the granulocytic progenitors and incomplete differentiation (up to myelocyte). The initial macrocluster growth pattern remained identical.

Extracellular matrix structure.

PubMed

Theocharis, Achilleas D; Skandalis, Spyros S; Gialeli, Chrysostomi; Karamanos, Nikos K

2016-02-01

Extracellular matrix (ECM) is a non-cellular three-dimensional macromolecular network composed of collagens, proteoglycans/glycosaminoglycans, elastin, fibronectin, laminins, and several other glycoproteins. Matrix components bind each other as well as cell adhesion receptors forming a complex network into which cells reside in all tissues and organs. Cell surface receptors transduce signals into cells from ECM, which regulate diverse cellular functions, such as survival, growth, migration, and differentiation, and are vital for maintaining normal homeostasis. ECM is a highly dynamic structural network that continuously undergoes remodeling mediated by several matrix-degrading enzymes during normal and pathological conditions. Deregulation of ECM composition and structure is associated with the development and progression of several pathologic conditions. This article emphasizes in the complex ECM structure as to provide a better understanding of its dynamic structural and functional multipotency. Where relevant, the implication of the various families of ECM macromolecules in health and disease is also presented. Copyright © 2015 Elsevier B.V. All rights reserved.

A Select Subset of Electron Transport Chain Genes Associated with Optic Atrophy Link Mitochondria to Axon Regeneration in Caenorhabditis elegans.

PubMed

Knowlton, Wendy M; Hubert, Thomas; Wu, Zilu; Chisholm, Andrew D; Jin, Yishi

2017-01-01

The role of mitochondria within injured neurons is an area of active interest since these organelles are vital for the production of cellular energy in the form of ATP. Using mechanosensory neurons of the nematode Caenorhabditis elegans to test regeneration after neuronal injury in vivo , we surveyed genes related to mitochondrial function for effects on axon regrowth after laser axotomy. Genes involved in mitochondrial transport, calcium uptake, mitophagy, or fission and fusion were largely dispensable for axon regrowth, with the exception of eat-3/Opa1 . Surprisingly, many genes encoding components of the electron transport chain were dispensable for regrowth, except for the iron-sulfur proteins gas-1, nduf-2.2, nduf-7 , and isp-1 , and the putative oxidoreductase rad-8 . In these mutants, axonal development was essentially normal and axons responded normally to injury by forming regenerative growth cones, but were impaired in subsequent axon extension. Overexpression of nduf-2.2 or isp-1 was sufficient to enhance regrowth, suggesting that mitochondrial function is rate-limiting in axon regeneration. Moreover, loss of function in isp-1 reduced the enhanced regeneration caused by either a gain-of-function mutation in the calcium channel EGL-19 or overexpression of the MAP kinase DLK-1. While the cellular function of RAD-8 remains unclear, our genetic analyses place rad-8 in the same pathway as other electron transport genes in axon regeneration. Unexpectedly, rad-8 regrowth defects were suppressed by altered function in the ubiquinone biosynthesis gene clk-1 . Furthermore, we found that inhibition of the mitochondrial unfolded protein response via deletion of atfs-1 suppressed the defective regrowth in nduf-2.2 mutants. Together, our data indicate that while axon regeneration is not significantly affected by general dysfunction of cellular respiration, it is sensitive to the proper functioning of a select subset of electron transport chain genes, or to the cellular adaptations used by neurons under conditions of injury.

A Select Subset of Electron Transport Chain Genes Associated with Optic Atrophy Link Mitochondria to Axon Regeneration in Caenorhabditis elegans

PubMed Central

Knowlton, Wendy M.; Hubert, Thomas; Wu, Zilu; Chisholm, Andrew D.; Jin, Yishi

2017-01-01

The role of mitochondria within injured neurons is an area of active interest since these organelles are vital for the production of cellular energy in the form of ATP. Using mechanosensory neurons of the nematode Caenorhabditis elegans to test regeneration after neuronal injury in vivo, we surveyed genes related to mitochondrial function for effects on axon regrowth after laser axotomy. Genes involved in mitochondrial transport, calcium uptake, mitophagy, or fission and fusion were largely dispensable for axon regrowth, with the exception of eat-3/Opa1. Surprisingly, many genes encoding components of the electron transport chain were dispensable for regrowth, except for the iron-sulfur proteins gas-1, nduf-2.2, nduf-7, and isp-1, and the putative oxidoreductase rad-8. In these mutants, axonal development was essentially normal and axons responded normally to injury by forming regenerative growth cones, but were impaired in subsequent axon extension. Overexpression of nduf-2.2 or isp-1 was sufficient to enhance regrowth, suggesting that mitochondrial function is rate-limiting in axon regeneration. Moreover, loss of function in isp-1 reduced the enhanced regeneration caused by either a gain-of-function mutation in the calcium channel EGL-19 or overexpression of the MAP kinase DLK-1. While the cellular function of RAD-8 remains unclear, our genetic analyses place rad-8 in the same pathway as other electron transport genes in axon regeneration. Unexpectedly, rad-8 regrowth defects were suppressed by altered function in the ubiquinone biosynthesis gene clk-1. Furthermore, we found that inhibition of the mitochondrial unfolded protein response via deletion of atfs-1 suppressed the defective regrowth in nduf-2.2 mutants. Together, our data indicate that while axon regeneration is not significantly affected by general dysfunction of cellular respiration, it is sensitive to the proper functioning of a select subset of electron transport chain genes, or to the cellular adaptations used by neurons under conditions of injury. PMID:28539870

Multiple functions of BCL-2 family proteins.

PubMed

Hardwick, J Marie; Soane, Lucian

2013-02-01

BCL-2 family proteins are the regulators of apoptosis, but also have other functions. This family of interacting partners includes inhibitors and inducers of cell death. Together they regulate and mediate the process by which mitochondria contribute to cell death known as the intrinsic apoptosis pathway. This pathway is required for normal embryonic development and for preventing cancer. However, before apoptosis is induced, BCL-2 proteins have critical roles in normal cell physiology related to neuronal activity, autophagy, calcium handling, mitochondrial dynamics and energetics, and other processes of normal healthy cells. The relative importance of these physiological functions compared to their apoptosis functions in overall organismal physiology is difficult to decipher. Apoptotic and noncanonical functions of these proteins may be intertwined to link cell growth to cell death. Disentanglement of these functions may require delineation of biochemical activities inherent to the characteristic three-dimensional shape shared by distantly related viral and cellular BCL-2 family members.

Derivation of the expressions for γ50 and D50 for different individual TCP and NTCP models

NASA Astrophysics Data System (ADS)

Stavreva, N.; Stavrev, P.; Warkentin, B.; Fallone, B. G.

2002-10-01

This paper presents a complete set of formulae for the position (D50) and the normalized slope (γ50) of the dose-response relationship based on the most commonly used radiobiological models for tumours as well as for normal tissues. The functional subunit response models (critical element and critical volume) are used in the derivation of the formulae for the normal tissue. Binomial statistics are used to describe the tumour control probability, the functional subunit response as well as the normal tissue complication probability. The formulae are derived for the single hit and linear quadratic models of cell kill in terms of the number of fractions and dose per fraction. It is shown that the functional subunit models predict very steep, almost step-like, normal tissue individual dose-response relationships. Furthermore, the formulae for the normalized gradient depend on the cellular parameters α and β when written in terms of number of fractions, but not when written in terms of dose per fraction.

Arctigenin preferentially induces tumor cell death under glucose deprivation by inhibiting cellular energy metabolism.

PubMed

Gu, Yuan; Qi, Chunting; Sun, Xiaoxiao; Ma, Xiuquan; Zhang, Haohao; Hu, Lihong; Yuan, Junying; Yu, Qiang

2012-08-15

Selectively eradicating cancer cells with minimum adverse effects on normal cells is a major challenge in the development of anticancer therapy. We hypothesize that nutrient-limiting conditions frequently encountered by cancer cells in poorly vascularized solid tumors might provide an opportunity for developing selective therapy. In this study, we investigated the function and molecular mechanisms of a natural compound, arctigenin, in regulating tumor cell growth. We demonstrated that arctigenin selectively promoted glucose-starved A549 tumor cells to undergo necrosis by inhibiting mitochondrial respiration. In doing so, arctigenin elevated cellular level of reactive oxygen species (ROS) and blocked cellular energy metabolism in the glucose-starved tumor cells. We also demonstrated that cellular ROS generation was caused by intracellular ATP depletion and played an essential role in the arctigenin-induced tumor cell death under the glucose-limiting condition. Furthermore, we combined arctigenin with the glucose analogue 2-deoxyglucose (2DG) and examined their effects on tumor cell growth. Interestingly, this combination displayed preferential cell-death inducing activity against tumor cells compared to normal cells. Hence, we propose that the combination of arctigenin and 2DG may represent a promising new cancer therapy with minimal normal tissue toxicity. Crown Copyright © 2012. Published by Elsevier Inc. All rights reserved.

Cortactin Branches Out: Roles in Regulating Protrusive Actin Dynamics

PubMed Central

Ammer, Amanda Gatesman; Weed, Scott A.

2008-01-01

Since its discovery in the early 1990’s, cortactin has emerged as a key signaling protein in many cellular processes, including cell adhesion, migration, endocytosis, and tumor invasion. While the list of cellular functions influenced by cortactin grows, the ability of cortactin to interact with and alter the cortical actin network is central to its role in regulating these processes. Recently, several advances have been made in our understanding of the interaction between actin and cortactin, providing insight into how these two proteins work together to provide a framework for normal and altered cellular function. This review examines how regulation of cortactin through post-translational modifications and interactions with multiple binding partners elicits changes in cortical actin cytoskeletal organization, impacting the regulation and formation of actin-rich motility structures. PMID:18615630

Evidence for Functional Differentiation among Drosophila Septins in Cytokinesis and Cellularization

PubMed Central

Adam, Jennifer C.; Pringle, John R.; Peifer, Mark

2000-01-01

The septins are a conserved family of proteins that are involved in cytokinesis and other aspects of cell-surface organization. In Drosophila melanogaster, null mutations in the pnut septin gene are recessive lethal, but homozygous pnut mutants complete embryogenesis and survive until the pupal stage. Because the completion of cellularization and other aspects of early development seemed likely to be due to maternally contributed Pnut product, we attempted to generate embryos lacking the maternal contribution in order to explore the roles of Pnut in these processes. We used two methods, the production of germline clones homozygous for a pnut mutation and the rescue of pnut homozygous mutant flies by a pnut+ transgene under control of the hsp70 promoter. Remarkably, the pnut germline-clone females produced eggs, indicating that stem-cell and cystoblast divisions in the female germline do not require Pnut. Moreover, the Pnut-deficient embryos obtained by either method completed early syncytial development and began cellularization of the embryo normally. However, during the later stages of cellularization, the organization of the actin cytoskeleton at the leading edge of the invaginating furrows became progressively more abnormal, and the embryos displayed widespread defects in cell and embryo morphology beginning at gastrulation. Examination of two other septins showed that Sep1 was not detectable at the cellularization front in the Pnut-deficient embryos, whereas Sep2 was still present in normal levels. Thus, it is possible that Sep2 (perhaps in conjunction with other septins such as Sep4 and Sep5) fulfills an essential septin role during the organization and initial ingression of the cellularization furrow even in the absence of Pnut and Sep1. Together, the results suggest that some cell-division events in Drosophila do not require septin function, that there is functional differentiation among the Drosophila septins, or both. PMID:10982405

Insulin-producing cells could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells

PubMed Central

2013-01-01

Objective The aim of this study was to compare the difference between insulin-producing cells (IPCs) and normal human pancreatic beta cells both in physiological function and morphological features in cellular level. Methods The levels of insulin secretion were measured by enzyme-linked immunosorbent assay. The insulin gene expression was determined by real-time quantitative polymerase chain reaction. The morphological features were detected by atomic force microscopy (AFM) and laser confocal scanning microscopy. Results IPCs and normal human pancreatic beta cells were similar to each other under the observation in AFM with the porous structure features in the cytoplasm. Both number of membrane particle size and average roughness of normal human beta cells were higher than those of IPCs. Conclusions Our results firstly revealed that the cellular ultrastructure of IPCs was closer to that of normal human pancreatic beta cells, but they still could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells. PMID:23421382

Adenovirus type 5 exerts genome-wide control over cellular programs governing proliferation, quiescence, and survival

PubMed Central

Miller, Daniel L; Myers, Chad L; Rickards, Brenden; Coller, Hilary A; Flint, S Jane

2007-01-01

Background Human adenoviruses, such as serotype 5 (Ad5), encode several proteins that can perturb cellular mechanisms that regulate cell cycle progression and apoptosis, as well as those that mediate mRNA production and translation. However, a global view of the effects of Ad5 infection on such programs in normal human cells is not available, despite widespread efforts to develop adenoviruses for therapeutic applications. Results We used two-color hybridization and oligonucleotide microarrays to monitor changes in cellular RNA concentrations as a function of time after Ad5 infection of quiescent, normal human fibroblasts. We observed that the expression of some 2,000 genes, about 10% of those examined, increased or decreased by a factor of two or greater following Ad5 infection, but were not altered in mock-infected cells. Consensus k-means clustering established that the temporal patterns of these changes were unexpectedly complex. Gene Ontology terms associated with cell proliferation were significantly over-represented in several clusters. The results of comparative analyses demonstrate that Ad5 infection induces reversal of the quiescence program and recapitulation of the core serum response, and that only a small subset of the observed changes in cellular gene expression can be ascribed to well characterized functions of the viral E1A and E1B proteins. Conclusion These findings establish that the impact of adenovirus infection on host cell programs is far greater than appreciated hitherto. Furthermore, they provide a new framework for investigating the molecular functions of viral early proteins and information relevant to the design of conditionally replicating adenoviral vectors. PMID:17430596

FAD-dependent lysine-specific demethylase-1 regulates cellular energy expenditure

PubMed Central

Hino, Shinjiro; Sakamoto, Akihisa; Nagaoka, Katsuya; Anan, Kotaro; Wang, Yuqing; Mimasu, Shinya; Umehara, Takashi; Yokoyama, Shigeyuki; Kosai, Ken-ichiro; Nakao, Mitsuyoshi

2012-01-01

Environmental factors such as nutritional state may act on the epigenome that consequently contributes to the metabolic adaptation of cells and the organisms. The lysine-specific demethylase-1 (LSD1) is a unique nuclear protein that utilizes flavin adenosine dinucleotide (FAD) as a cofactor. Here we show that LSD1 epigenetically regulates energy-expenditure genes in adipocytes depending on the cellular FAD availability. We find that the loss of LSD1 function, either by short interfering RNA or by selective inhibitors in adipocytes, induces a number of regulators of energy expenditure and mitochondrial metabolism such as PPARγ coactivator-1α resulting in the activation of mitochondrial respiration. In the adipose tissues from mice on a high-fat diet, expression of LSD1-target genes is reduced, compared with that in tissues from mice on a normal diet, which can be reverted by suppressing LSD1 function. Our data suggest a novel mechanism where LSD1 regulates cellular energy balance through coupling with cellular FAD biosynthesis. PMID:22453831

FAD-dependent lysine-specific demethylase-1 regulates cellular energy expenditure.

PubMed

Hino, Shinjiro; Sakamoto, Akihisa; Nagaoka, Katsuya; Anan, Kotaro; Wang, Yuqing; Mimasu, Shinya; Umehara, Takashi; Yokoyama, Shigeyuki; Kosai, Ken-Ichiro; Nakao, Mitsuyoshi

2012-03-27

Environmental factors such as nutritional state may act on the epigenome that consequently contributes to the metabolic adaptation of cells and the organisms. The lysine-specific demethylase-1 (LSD1) is a unique nuclear protein that utilizes flavin adenosine dinucleotide (FAD) as a cofactor. Here we show that LSD1 epigenetically regulates energy-expenditure genes in adipocytes depending on the cellular FAD availability. We find that the loss of LSD1 function, either by short interfering RNA or by selective inhibitors in adipocytes, induces a number of regulators of energy expenditure and mitochondrial metabolism such as PPARγ coactivator-1α resulting in the activation of mitochondrial respiration. In the adipose tissues from mice on a high-fat diet, expression of LSD1-target genes is reduced, compared with that in tissues from mice on a normal diet, which can be reverted by suppressing LSD1 function. Our data suggest a novel mechanism where LSD1 regulates cellular energy balance through coupling with cellular FAD biosynthesis.

Is the Cortical Deficit in Amblyopia Due to Reduced Cortical Magnification, Loss of Neural Resolution, or Neural Disorganization?

PubMed

Clavagnier, Simon; Dumoulin, Serge O; Hess, Robert F

2015-11-04

The neural basis of amblyopia is a matter of debate. The following possibilities have been suggested: loss of foveal cells, reduced cortical magnification, loss of spatial resolution of foveal cells, and topographical disarray in the cellular map. To resolve this we undertook a population receptive field (pRF) functional magnetic resonance imaging analysis in the central field in humans with moderate-to-severe amblyopia. We measured the relationship between averaged pRF size and retinal eccentricity in retinotopic visual areas. Results showed that cortical magnification is normal in the foveal field of strabismic amblyopes. However, the pRF sizes are enlarged for the amblyopic eye. We speculate that the pRF enlargement reflects loss of cellular resolution or an increased cellular positional disarray within the representation of the amblyopic eye. The neural basis of amblyopia, a visual deficit affecting 3% of the human population, remains a matter of debate. We undertook the first population receptive field functional magnetic resonance imaging analysis in participants with amblyopia and compared the projections from the amblyopic and fellow normal eye in the visual cortex. The projection from the amblyopic eye was found to have a normal cortical magnification factor, enlarged population receptive field sizes, and topographic disorganization in all early visual areas. This is consistent with an explanation of amblyopia as an immature system with a normal complement of cells whose spatial resolution is reduced and whose topographical map is disordered. This bears upon a number of competing theories for the psychophysical defect and affects future treatment therapies. Copyright © 2015 the authors 0270-6474/15/3514740-16$15.00/0.

Cigarette smoke–induced induction of antioxidant enzyme activities in airway leukocytes is absent in active smokers with COPD

PubMed Central

Dove, Rosamund E.; Leong-Smith, Pheneatia; Roos-Engstrand, Ester; Pourazar, Jamshid; Shah, Mittal; Behndig, Annelie F.; Mudway, Ian S.; Blomberg, Anders

2015-01-01

Background Oxidative injury to the airway has been proposed as an important underlying mechanism in the pathogenesis of chronic obstructive pulmonary disease (COPD). As the extent of oxidant-mediated damage is dependent on the endogenous antioxidant defences within the airways, we examined whether COPD was associated with deficiencies in the antioxidant network within the respiratory tract lining fluids (RTLFs) and resident airway leukocytes. We hypothesised that COPD would be associated with both basal depression of antioxidant defences and impaired adaptive antioxidant responses to cigarette smoke. Methods Low molecular weight and enzymatic antioxidants together with metal-handling proteins were quantified in bronchoalveolar lavage fluid and airway leukocytes, derived from current (n=9) and ex-smoking COPD patients (n=15), as well as from smokers with normal lung function (n=16) and healthy never smokers (n=13). Results Current cigarette smoking was associated with an increase in ascorbate and glutathione within peripheral RTLFs in both smokers with normal lung function compared with healthy never smokers and in COPD smokers compared with COPD ex-smokers. In contrast, intra-cellular antioxidant enzyme activities (glutathione peroxidase, glutathione reductase, and catalase) were only up-regulated in smokers with normal lung function compared with healthy never smokers and not in actively smoking COPD patients relative to COPD ex-smokers. Conclusions We found no evidence of impaired basal antioxidant defences, within either the RTLFs or airway leukocytes in stable ex-smoking COPD patients compared with healthy never smoking controls. Current cigarette smoking induced an up-regulation of low molecular weight antioxidants in the RTLFs of both control subjects with normal lung function and patients with COPD. Importantly, the present data demonstrated a cigarette smoke–induced increase in intra-cellular antioxidant enzyme activities only within the smokers with normal lung function, implying that patients with COPD who continue to smoke will experience enhanced oxidative stress, prompting disease progression. PMID:26557249

Histomorphometric analysis of nuclear and cellular volumetric alterations in oral lichen planus, lichenoid lesions and normal oral mucosa using image analysis software.

PubMed

Venkatesiah, Sowmya S; Kale, Alka D; Hallikeremath, Seema R; Kotrashetti, Vijayalakshmi S

2013-01-01

Lichen planus is a chronic inflammatory mucocutaneous disease that clinically and histologically resembles lichenoid lesions, although the latter has a different etiology. Though criteria have been suggested for differentiating oral lichen planus from lichenoid lesions, confusion still prevails. To study the cellular and nuclear volumetric features in the epithelium of normal mucosa, lichen planus, and lichenoid lesions to determine variations if any. A retrospective study was done on 25 histologically diagnosed cases each of oral lichen planus, oral lichenoid lesions, and normal oral mucosa. Cellular and nuclear morphometric measurements were assessed on hematoxylin and eosin sections using image analysis software. Analysis of variance test (ANOVA) and Tukey's post-hoc test. The basal cells of oral lichen planus showed a significant increase in the mean nuclear and cellular areas, and in nuclear volume; there was a significant decrease in the nuclear-cytoplasmic ratio as compared to normal mucosa. The suprabasal cells showed a significant increase in nuclear and cellular areas, nuclear diameter, and nuclear and cellular volumes as compared to normal mucosa. The basal cells of oral lichenoid lesions showed significant difference in the mean cellular area and the mean nuclear-cytoplasmic ratio as compared to normal mucosa, whereas the suprabasal cells differed significantly from normal mucosa in the mean nuclear area and the nuclear and cellular volumes. Morphometry can differentiate lesions of oral lichen planus and oral lichenoid lesions from normal oral mucosa. Thus, morphometry may serve to discriminate between normal and premalignant lichen planus and lichenoid lesions. These lesions might have a high risk for malignant transformation and may behave in a similar manner with respect to malignant transformation.

Specific targeting of proteins to outer envelope membranes of endosymbiotic organelles, chloroplasts, and mitochondria

PubMed Central

Lee, Junho; Kim, Dae Heon; Hwang, Inhwan

2014-01-01

Chloroplasts and mitochondria are endosymbiotic organelles thought to be derived from endosymbiotic bacteria. In present-day eukaryotic cells, these two organelles play pivotal roles in photosynthesis and ATP production. In addition to these major activities, numerous reactions, and cellular processes that are crucial for normal cellular functions occur in chloroplasts and mitochondria. To function properly, these organelles constantly communicate with the surrounding cellular compartments. This communication includes the import of proteins, the exchange of metabolites and ions, and interactions with other organelles, all of which heavily depend on membrane proteins localized to the outer envelope membranes. Therefore, correct and efficient targeting of these membrane proteins, which are encoded by the nuclear genome and translated in the cytosol, is critically important for organellar function. In this review, we summarize the current knowledge of the mechanisms of protein targeting to the outer membranes of mitochondria and chloroplasts in two different directions, as well as targeting signals and cytosolic factors. PMID:24808904

Fluorescence-based detection and quantification of features of cellular senescence.

PubMed

Cho, Sohee; Hwang, Eun Seong

2011-01-01

Cellular senescence is a spontaneous organismal defense mechanism against tumor progression which is raised upon the activation of oncoproteins or other cellular environmental stresses that must be circumvented for tumorigenesis to occur. It involves growth-arrest state of normal cells after a number of active divisions. There are multiple experimental routes that can drive cells into a state of senescence. Normal somatic cells and cancer cells enter a state of senescence upon overexpression of oncogenic Ras or Raf protein or by imposing certain kinds of stress such as cellular tumor suppressor function. Both flow cytometry and confocal imaging analysis techniques are very useful in quantitative analysis of cellular senescence phenomenon. They allow quantitative estimates of multiple different phenotypes expressed in multiple cell populations simultaneously. Here we review the various types of fluorescence methodologies including confocal imaging and flow cytometry that are frequently utilized to study a variety of senescence. First, we discuss key cell biological changes occurring during senescence and review the current understanding on the mechanisms of these changes with the goal of improving existing protocols and further developing new ones. Next, we list specific senescence phenotypes associated with each cellular trait along with the principles of their assay methods and the significance of the assay outcomes. We conclude by selecting appropriate references that demonstrate a typical example of each method. Copyright © 2011 Elsevier Inc. All rights reserved.

Mitochondrial damage and cytoskeleton reorganization in human dermal fibroblasts exposed to artificial visible light similar to screen-emitted light.

PubMed

Rascalou, Adeline; Lamartine, Jérôme; Poydenot, Pauline; Demarne, Frédéric; Bechetoille, Nicolas

2018-05-05

Artificial visible light is everywhere in modern life. Social communication confronts us with screens of all kinds, and their use is on the rise. We are therefore increasingly exposed to artificial visible light, the effects of which on skin are poorly known. The purpose of this study was to model the artificial visible light emitted by electronic devices and assess its effect on normal human fibroblasts. The spectral irradiance emitted by electronic devices was optically measured and equipment was developed to accurately reproduce such artificial visible light. Effects on normal human fibroblasts were analyzed on human genome microarray-based gene expression analysis. At cellular level, visualization and image analysis were performed on the mitochondrial network and F-actin cytoskeleton. Cell proliferation, ATP release and type I procollagen secretion were also measured. We developed a device consisting of 36 LEDs simultaneously emitting blue, green and red light at distinct wavelengths (450 nm, 525 nm and 625 nm) with narrow spectra and equivalent radiant power for the three colors. A dose of 99 J/cm 2 artificial visible light was selected so as not to induce cell mortality following exposure. Microarray analysis revealed 2984 light-modulated transcripts. Functional annotation of light-responsive genes revealed several enriched functions including, amongst others, the "mitochondria" and "integrin signaling" categories. Selected results were confirmed by real-time quantitative PCR, analyzing 24 genes representing these two categories. Analysis of micro-patterned culture plates showed marked fragmentation of the mitochondrial network and disorganization of the F-actin cytoskeleton following exposure. Functionally, there was considerable impairment of cell growth and spread, ATP release and type I procollagen secretion in exposed fibroblasts. Artificial visible light induces drastic molecular and cellular changes in normal human fibroblasts. This may impede normal cellular functions and contribute to premature skin aging. The present results extend our knowledge of the effects of the low-energy wavelengths that are increasingly used to treat skin disorders. Copyright © 2018 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.

Condition monitoring of 3G cellular networks through competitive neural models.

PubMed

Barreto, Guilherme A; Mota, João C M; Souza, Luis G M; Frota, Rewbenio A; Aguayo, Leonardo

2005-09-01

We develop an unsupervised approach to condition monitoring of cellular networks using competitive neural algorithms. Training is carried out with state vectors representing the normal functioning of a simulated CDMA2000 network. Once training is completed, global and local normality profiles (NPs) are built from the distribution of quantization errors of the training state vectors and their components, respectively. The global NP is used to evaluate the overall condition of the cellular system. If abnormal behavior is detected, local NPs are used in a component-wise fashion to find abnormal state variables. Anomaly detection tests are performed via percentile-based confidence intervals computed over the global and local NPs. We compared the performance of four competitive algorithms [winner-take-all (WTA), frequency-sensitive competitive learning (FSCL), self-organizing map (SOM), and neural-gas algorithm (NGA)] and the results suggest that the joint use of global and local NPs is more efficient and more robust than current single-threshold methods.

Oxidative stress, protein modification and Alzheimer disease.

PubMed

Tramutola, A; Lanzillotta, C; Perluigi, M; Butterfield, D Allan

2017-07-01

Alzheimer disease (AD) is a progressive neurodegenerative disease that affects the elderly population with complex etiology. Many hypotheses have been proposed to explain different causes of AD, but the exact mechanisms remain unclear. In this review, we focus attention on the oxidative-stress hypothesis of neurodegeneration and we discuss redox proteomics approaches to analyze post-mortem human brain from AD brain. Collectively, these studies have provided valuable insights into the molecular mechanisms involved both in the pathogenesis and progression of AD, demonstrating the impairment of numerous cellular processes such as energy production, cellular structure, signal transduction, synaptic function, mitochondrial function, cell cycle progression, and degradative systems. Each of these cellular functions normally contributes to maintain healthy neuronal homeostasis, so the deregulation of one or more of these functions could contribute to the pathology and clinical presentation of AD. In particular, we discuss the evidence demonstrating the oxidation/dysfunction of a number of enzymes specifically involved in energy metabolism that support the view that reduced glucose metabolism and loss of ATP are crucial events triggering neurodegeneration and progression of AD. Copyright © 2016 Elsevier Inc. All rights reserved.

Wavelet-based multiscale analysis of bioimpedance data measured by electric cell-substrate impedance sensing for classification of cancerous and normal cells.

PubMed

Das, Debanjan; Shiladitya, Kumar; Biswas, Karabi; Dutta, Pranab Kumar; Parekh, Aditya; Mandal, Mahitosh; Das, Soumen

2015-12-01

The paper presents a study to differentiate normal and cancerous cells using label-free bioimpedance signal measured by electric cell-substrate impedance sensing. The real-time-measured bioimpedance data of human breast cancer cells and human epithelial normal cells employs fluctuations of impedance value due to cellular micromotions resulting from dynamic structural rearrangement of membrane protrusions under nonagitated condition. Here, a wavelet-based multiscale quantitative analysis technique has been applied to analyze the fluctuations in bioimpedance. The study demonstrates a method to classify cancerous and normal cells from the signature of their impedance fluctuations. The fluctuations associated with cellular micromotion are quantified in terms of cellular energy, cellular power dissipation, and cellular moments. The cellular energy and power dissipation are found higher for cancerous cells associated with higher micromotions in cancer cells. The initial study suggests that proposed wavelet-based quantitative technique promises to be an effective method to analyze real-time bioimpedance signal for distinguishing cancer and normal cells.

The role of HFE genotype in macrophage phenotype.

PubMed

Nixon, Anne M; Neely, Elizabeth; Simpson, Ian A; Connor, James R

2018-02-01

Iron regulation is essential for cellular energy production. Loss of cellular iron homeostasis has critical implications for both normal function and disease progression. The H63D variant of the HFE gene is the most common gene variant in Caucasians. The resulting mutant protein alters cellular iron homeostasis and is associated with a number of neurological diseases and cancer. In the brain, microglial and infiltrating macrophages are critical to maintaining iron homeostasis and modulating inflammation associated with the pathogenic process in multiple diseases. This study addresses whether HFE genotype affects macrophage function and the implications of these findings for disease processes. Bone marrow macrophages were isolated from wildtype and H67D HFE knock-in mice. The H67D gene variant in mice is the human equivalent of the H63D variant. Upon differentiation, the macrophages were used to analyze iron regulatory proteins, cellular iron release, migration, phagocytosis, and cytokine expression. The results of this study demonstrate that the H67D HFE genotype significantly impacts a number of critical macrophage functions. Specifically, fundamental activities such as proliferation in response to iron exposure, L-ferritin expression in response to iron loading, secretion of BMP6 and cytokines, and migration and phagocytic activity were all found to be impacted by genotype. Furthermore, we demonstrated that exposure to apo-Tf (iron-poor transferrin) can increase the release of iron from macrophages. In normal conditions, 70% of circulating transferrin is unsaturated. Therefore, the ability of apo-Tf to induce iron release could be a major regulatory mechanism for iron release from macrophages. These studies demonstrate that the HFE genotype impacts fundamental components of macrophage phenotype that could alter their role in degenerative and reparative processes in neurodegenerative disorders.

Simultaneous Multiparameter Cellular Energy Metabolism Profiling of Small Populations of Cells.

PubMed

Kelbauskas, Laimonas; Ashili, Shashaanka P; Lee, Kristen B; Zhu, Haixin; Tian, Yanqing; Meldrum, Deirdre R

2018-03-12

Functional and genomic heterogeneity of individual cells are central players in a broad spectrum of normal and disease states. Our knowledge about the role of cellular heterogeneity in tissue and organism function remains limited due to analytical challenges one encounters when performing single cell studies in the context of cell-cell interactions. Information based on bulk samples represents ensemble averages over populations of cells, while data generated from isolated single cells do not account for intercellular interactions. We describe a new technology and demonstrate two important advantages over existing technologies: first, it enables multiparameter energy metabolism profiling of small cell populations (<100 cells)-a sample size that is at least an order of magnitude smaller than other, commercially available technologies; second, it can perform simultaneous real-time measurements of oxygen consumption rate (OCR), extracellular acidification rate (ECAR), and mitochondrial membrane potential (MMP)-a capability not offered by any other commercially available technology. Our results revealed substantial diversity in response kinetics of the three analytes in dysplastic human epithelial esophageal cells and suggest the existence of varying cellular energy metabolism profiles and their kinetics among small populations of cells. The technology represents a powerful analytical tool for multiparameter studies of cellular function.

Mapping of oxidative stress response elements of the caveolin-1 promoter.

PubMed

Bartholomew, Janine N; Galbiati, Ferruccio

2010-01-01

According to the "free radical theory" of aging, normal aging occurs as the result of tissue damages inflicted by reactive oxygen species (ROS). ROS are known to induce cellular senescence, and senescent cells are believed to contribute to organismal aging. The molecular mechanisms that mediate the cellular response to oxidants remain to be fully identified. We have shown that oxidative stress induces cellular senescence through activation of the caveolin-1 promoter and upregulation of caveolin-1 protein expression. Here, we describe how reactive oxygen species activate the caveolin-1 promoter and how the signaling may be assayed. These approaches provide insight into the functional role of caveolin-1 and potentially allow the identification of novel ROS-regulated genes that are part of the signaling machinery regulating cellular senescence/aging.

The neurology of mTOR.

PubMed

Lipton, Jonathan O; Sahin, Mustafa

2014-10-22

The mechanistic target of rapamycin (mTOR) signaling pathway is a crucial cellular signaling hub that, like the nervous system itself, integrates internal and external cues to elicit critical outputs including growth control, protein synthesis, gene expression, and metabolic balance. The importance of mTOR signaling to brain function is underscored by the myriad disorders in which mTOR pathway dysfunction is implicated, such as autism, epilepsy, and neurodegenerative disorders. Pharmacological manipulation of mTOR signaling holds therapeutic promise and has entered clinical trials for several disorders. Here, we review the functions of mTOR signaling in the normal and pathological brain, highlighting ongoing efforts to translate our understanding of cellular physiology into direct medical benefit for neurological disorders.

Cell biology of spinocerebellar ataxia.

PubMed

Orr, Harry T

2012-04-16

Ataxia is a neurological disorder characterized by loss of control of body movements. Spinocerebellar ataxia (SCA), previously known as autosomal dominant cerebellar ataxia, is a biologically robust group of close to 30 progressive neurodegenerative diseases. Six SCAs, including the more prevalent SCA1, SCA2, SCA3, and SCA6 along with SCA7 and SCA17 are caused by expansion of a CAG repeat that encodes a polyglutamine tract in the affected protein. How the mutated proteins in these polyglutamine SCAs cause disease is highly debated. Recent work suggests that the mutated protein contributes to pathogenesis within the context of its "normal" cellular function. Thus, understanding the cellular function of these proteins could aid in the development of therapeutics.

Nuclear Mechanics in Disease

PubMed Central

Zwerger, Monika; Ho, Chin Yee; Lammerding, Jan

2015-01-01

Over the past two decades, the biomechanical properties of cells have emerged as key players in a broad range of cellular functions, including migration, proliferation, and differentiation. Although much of the attention has focused on the cytoskeletal networks and the cell’s microenvironment, relatively little is known about the contribution of the cell nucleus. Here, we present an overview of the structural elements that determine the physical properties of the nucleus and discuss how changes in the expression of nuclear components or mutations in nuclear proteins can affect not only nuclear mechanics but also modulate cytoskeletal organization and diverse cellular functions. These findings illustrate that the nucleus is tightly integrated into the surrounding cellular structure. Consequently, changes in nuclear structure and composition are highly relevant to normal development and physiology and can contribute to many human diseases, such as muscular dystrophy, dilated cardiomyopathy, (premature) aging, and cancer. PMID:21756143

A role for autophagic protein beclin 1 early in lymphocyte development.

PubMed

Arsov, Ivica; Adebayo, Adeola; Kucerova-Levisohn, Martina; Haye, Joanna; MacNeil, Margaret; Papavasiliou, F Nina; Yue, Zhenyu; Ortiz, Benjamin D

2011-02-15

Autophagy is a highly regulated and evolutionarily conserved process of cellular self-digestion. Recent evidence suggests that this process plays an important role in regulating T cell homeostasis. In this study, we used Rag1(-/-) (recombination activating gene 1(-/-)) blastocyst complementation and in vitro embryonic stem cell differentiation to address the role of Beclin 1, one of the key autophagic proteins, in lymphocyte development. Beclin 1-deficient Rag1(-/-) chimeras displayed a dramatic reduction in thymic cellularity compared with control mice. Using embryonic stem cell differentiation in vitro, we found that the inability to maintain normal thymic cellularity is likely caused by impaired maintenance of thymocyte progenitors. Interestingly, despite drastically reduced thymocyte numbers, the peripheral T cell compartment of Beclin 1-deficient Rag1(-/-) chimeras is largely normal. Peripheral T cells displayed normal in vitro proliferation despite significantly reduced numbers of autophagosomes. In addition, these chimeras had greatly reduced numbers of early B cells in the bone marrow compared with controls. However, the peripheral B cell compartment was not dramatically impacted by Beclin 1 deficiency. Collectively, our results suggest that Beclin 1 is required for maintenance of undifferentiated/early lymphocyte progenitor populations. In contrast, Beclin 1 is largely dispensable for the initial generation and function of the peripheral T and B cell compartments. This indicates that normal lymphocyte development involves Beclin 1-dependent, early-stage and distinct, Beclin 1-independent, late-stage processes.

Nicotinic Acid Receptor Abnormalities in Human Skin Cancer: Implications for a Role in Epidermal Differentiation

PubMed Central

Bermudez, Yira; Benavente, Claudia A.; Meyer, Ralph G.; Coyle, W. Russell; Jacobson, Myron K.; Jacobson, Elaine L.

2011-01-01

Background Chronic UV skin exposure leads to epidermal differentiation defects in humans that can be largely restored by pharmacological doses of nicotinic acid. Nicotinic acid has been identified as a ligand for the human G-protein-coupled receptors GPR109A and GPR109B that signal through Gi-mediated inhibition of adenylyl cyclase. We have examined the expression, cellular distribution, and functionality of GPR109A/B in human skin and skin derived epidermal cells. Results Nicotinic acid increases epidermal differentiation in photodamaged human skin as judged by the terminal differentiation markers caspase 14 and filaggrin. Both GPR109A and GPR109B genes are transcribed in human skin and in epidermal keratinocytes, but expression in dermal fibroblasts is below limits of detection. Receptor transcripts are greatly over-expressed in squamous cell cancers. Receptor protein in normal skin is prominent from the basal through granular layers of the epidermis, with cellular localization more dispersive in the basal layer but predominantly localized at the plasma membrane in more differentiated epidermal layers. In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional Gi-mediated signaling. In contrast, in a squamous cell carcinoma derived cell line, receptor protein shows a more diffuse cellular localization and the receptors are nearly non-functional. Conclusions The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis. PMID:21655214

VIRTUAL LIVER: AN IN SILICO FRAMEWORK FOR ANALYZING CHEMICAL-INDUCED HEPATOTOXICITY

EPA Science Inventory

The US EPA Virtual Liver (v-LiverTM) is an in silico framework for the dose-dependent perturbation of normal hepatic functions by chemicals using in vitro data. The framework consists of a computable knowledge-base (KB) to infer putative pathways in hepatotoxicity and a cellular...

On the Normal Force Mechanotransduction of Human Umbilical Vein Endothelial Cells

NASA Astrophysics Data System (ADS)

Vahabikashi, Amir; Wang, Qiuyun; Wilson, James; Wu, Qianhong; Vucbmss Team

2016-11-01

In this paper, we report a cellular biomechanics study to examine the normal force mechanotransduction of Human Umbilical Vein Endothelial Cells (HUVECs) with their implications on hypertension. Endothelial cells sense mechanical forces and adjust their structure and function accordingly. The mechanotransduction of normal forces plays a vital role in hypertension due to the higher pressure buildup inside blood vessels. Herein, HUVECs were cultured to full confluency and then exposed to different mechanical loadings using a novel microfluidic flow chamber. One various pressure levels while keeps the shear stress constant inside the flow chamber. Three groups of cells were examined, the control group (neither shear nor normal stresses), the normal pressure group (10 dyne/cm2 of shear stress and 95 mmHg of pressure), and the hypertensive group (10 dyne/cm2 of shear stress and 142 mmHg of pressure). Cellular response characterized by RT-PCR method indicates that, COX-2 expressed under normal pressure but not high pressure; Mn-SOD expressed under both normal and high pressure while this response was stronger for normal pressure; FOS and e-NOS did not respond under any condition. The differential behavior of COX-2 and Mn-SOD in response to changes in pressure, is instrumental for better understanding the pathogenesis of hypertensive cardiovascular diseases. This research was supported by the National Science Foundation under Award #1511096.

Cell-to-cell communication and cellular environment alter the somatostatin status of delta cells

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

Kelly, Catriona, E-mail: catriona.kelly@qub.ac.uk; Flatt, Peter R.; McClenaghan, Neville H.

2010-08-20

Research highlights: {yields} TGP52 cells display enhanced functionality in pseudoislet form. {yields} Somatostatin content was reduced, but secretion increased in high glucose conditions. {yields} Cellular interactions and environment alter the somatostatin status of TGP52 cells. -- Abstract: Introduction: Somatostatin, released from pancreatic delta cells, is a potent paracrine inhibitor of insulin and glucagon secretion. Islet cellular interactions and glucose homeostasis are essential to maintain normal patterns of insulin secretion. However, the importance of cell-to-cell communication and cellular environment in the regulation of somatostatin release remains unclear. Methods: This study employed the somatostatin-secreting TGP52 cell line maintained in DMEM:F12 (17.5 mMmore » glucose) or DMEM (25 mM glucose) culture media. The effect of pseudoislet formation and culture medium on somatostatin content and release in response to a variety of stimuli was measured by somatostatin EIA. In addition, the effect of pseudoislet formation on cellular viability (MTT and LDH assays) and proliferation (BrdU ELISA) was determined. Results: TGP52 cells readily formed pseudoislets and showed enhanced functionality in three-dimensional form with increased E-cadherin expression irrespective of the culture environment used. However, culture in DMEM decreased cellular somatostatin content (P < 0.01) and increased somatostatin secretion in response to a variety of stimuli including arginine, calcium and PMA (P < 0.001) when compared with cells grown in DMEM:F12. Configuration of TGP52 cells as pseudoislets reduced the proliferative rate and increased cellular cytotoxicity irrespective of culture medium used. Conclusions: Somatostatin secretion is greatly facilitated by cell-to-cell interactions and E-cadherin expression. Cellular environment and extracellular glucose also significantly influence the function of delta cells.« less

Aquatide Activation of SIRT1 Reduces Cellular Senescence through a SIRT1-FOXO1-Autophagy Axis.

PubMed

Lim, Chae Jin; Lee, Yong-Moon; Kang, Seung Goo; Lim, Hyung W; Shin, Kyong-Oh; Jeong, Se Kyoo; Huh, Yang Hoon; Choi, Suin; Kor, Myungho; Seo, Ho Seong; Park, Byeong Deog; Park, Keedon; Ahn, Jeong Keun; Uchida, Yoshikazu; Park, Kyungho

2017-09-01

Ultraviolet (UV) irradiation is a relevant environment factor to induce cellular senescence and photoaging. Both autophagy- and silent information regulator T1 (SIRT1)-dependent pathways are critical cellular processes of not only maintaining normal cellular functions, but also protecting cellular senescence in skin exposed to UV irradiation. In the present studies, we investigated whether modulation of autophagy induction using a novel synthetic SIRT1 activator, heptasodium hexacarboxymethyl dipeptide-12 (named as Aquatide), suppresses the UVB irradiation-induced skin aging. Treatment with Aquatide directly activates SIRT1 and stimulates autophagy induction in cultured human dermal fibroblasts. Next, we found that Aquatide-mediated activation of SIRT1 increases autophagy induction via deacetylation of forkhead box class O (FOXO) 1. Finally, UVB irradiation-induced cellular senescence measured by SA-β-gal staining was significantly decreased in cells treated with Aquatide in parallel to occurring SIRT1 activation-dependent autophagy. Together, Aquatide modulates autophagy through SIRT1 activation, contributing to suppression of skin aging caused by UV irradiation.

A Comparison of Raman Spectral Features of Frozen and Deparaffinized Tissues in Neuroblastoma and Ganglioneuroma

NASA Astrophysics Data System (ADS)

Devpura, Suneetha; Thakur, Jagdish S.; Poulik, Janet M.; Rabah, Raja; Naik, Vaman M.; Naik, Ratna

2012-02-01

We have investigated the cellular regions in neuroblastoma and ganglioneuroma using Raman spectroscopy and compared their spectral characteristics with those of normal adrenal gland. Thin sections from both frozen and deparaffinized tissues, obtained from the same tissue specimen, were studied in conjunction with the pathological examination of the tissues. We found a significant difference in the spectral features of frozen sections of normal adrenal gland, neuroblastoma, and ganglioneuroma when compared to deparaffinized tissues. The quantitative analysis of the Raman data using chemometric methods of principal component analysis and discriminant function analysis obtained from the frozen tissues show a sensitivity and specificity of 100% each. The biochemical identification based on the spectral differences shows that the normal adrenal gland tissues have higher levels of carotenoids, lipids, and cholesterol compared to the neuroblastoma and ganglioneuroma frozen tissues. However, deparaffinized tissues show complete removal of these biochemicals in adrenal tissues. This study demonstrates that Raman spectroscopy combined with chemometric methods can successfully distinguish neuroblastoma and ganglioneuroma at cellular level.

Lognormal Distribution of Cellular Uptake of Radioactivity: Statistical Analysis of α-Particle Track Autoradiography

PubMed Central

Neti, Prasad V.S.V.; Howell, Roger W.

2010-01-01

Recently, the distribution of radioactivity among a population of cells labeled with 210Po was shown to be well described by a log-normal (LN) distribution function (J Nucl Med. 2006;47:1049–1058) with the aid of autoradiography. To ascertain the influence of Poisson statistics on the interpretation of the autoradiographic data, the present work reports on a detailed statistical analysis of these earlier data. Methods The measured distributions of α-particle tracks per cell were subjected to statistical tests with Poisson, LN, and Poisson-lognormal (P-LN) models. Results The LN distribution function best describes the distribution of radioactivity among cell populations exposed to 0.52 and 3.8 kBq/mL of 210Po-citrate. When cells were exposed to 67 kBq/mL, the P-LN distribution function gave a better fit; however, the underlying activity distribution remained log-normal. Conclusion The present analysis generally provides further support for the use of LN distributions to describe the cellular uptake of radioactivity. Care should be exercised when analyzing autoradiographic data on activity distributions to ensure that Poisson processes do not distort the underlying LN distribution. PMID:18483086

Mitochondria and Iron: Current Questions

PubMed Central

Paul, Bibbin T.; Manz, David H.; Torti, Frank M.; Torti, Suzy V.

2017-01-01

Introduction Mitochondria are cellular organelles that perform numerous bioenergetic, biosynthetic, and regulatory functions and play a central role in iron metabolism. Extracellular iron is taken up by cells and transported to the mitochondria, where it is utilized for synthesis of cofactors essential to the function of enzymes involved in oxidation-reduction reactions, DNA synthesis and repair, and a variety of other cellular processes. Areas Covered This article reviews the trafficking of iron to the mitochondria and normal mitochondrial iron metabolism, including heme synthesis and iron-sulfur cluster biogenesis. Much of our understanding of mitochondrial iron metabolism has been revealed by pathologies that disrupt normal iron metabolism. These conditions affect not only iron metabolism but mitochondrial function and systemic health. Therefore, this article also discusses these pathologies, including conditions of systemic and mitochondrial iron dysregulation as well as cancer. Literature covering these areas was identified via PubMed searches using keywords: Iron, mitochondria, Heme Synthesis, Iron-sulfur Cluster, and Cancer. References cited by publications retrieved using this search strategy were also consulted. Expert Commentary While much has been learned about mitochondrial iron, key questions remain. Developing a better understanding of mitochondrial iron regulation will be paramount in developing therapies for syndromes that affect mitochondrial iron. PMID:27911100

Endoplasmic Reticulum Stress in the Diabetic Kidney, the Good, the Bad and the Ugly.

PubMed

Cunard, Robyn

2015-04-20

Diabetic kidney disease is the leading worldwide cause of end stage kidney disease and a growing public health challenge. The diabetic kidney is exposed to many environmental stressors and each cell type has developed intricate signaling systems designed to restore optimal cellular function. The unfolded protein response (UPR) is a homeostatic pathway that regulates endoplasmic reticulum (ER) membrane structure and secretory function. Studies suggest that the UPR is activated in the diabetic kidney to restore normal ER function and viability. However, when the cell is continuously stressed in an environment that lies outside of its normal physiological range, then the UPR is known as the ER stress response. The UPR reduces protein synthesis, augments the ER folding capacity and downregulates mRNA expression of genes by multiple pathways. Aberrant activation of ER stress can also induce inflammation and cellular apoptosis, and modify signaling of protective processes such as autophagy and mTORC activation. The following review will discuss our current understanding of ER stress in the diabetic kidney and explore novel means of modulating ER stress and its interacting signaling cascades with the overall goal of identifying therapeutic strategies that will improve outcomes in diabetic nephropathy.

The thorny path linking cellular senescence to organismalaging

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

Patil, Christopher K.; Mian, Saira; Campisi, Judith

2005-08-09

Half a century is fast approaching since Hayflick and colleagues formally described the limited ability of normal human cells to proliferate in culture (Hayflick and Moorhead, 1961). This finding--that normal somatic cells, in contrast to cancer cells, cannot divide indefinitely--challenged the prevailing idea that cells from mortal multicellular organisms were intrinsically ''immortal'' (Carrell, 1912). It also spawned two hypotheses, essential elements of which persist today. The first held that the restricted proliferation of normal cells, now termed cellular senescence, suppresses cancer (Hayflick, 1965; Sager, 1991; Campisi, 2001). The second hypothesis, as explained in the article by Lorenzini et al., suggestedmore » that the limited proliferation of cells in culture recapitulated aspects of organismal aging (Hayflick, 1965; Martin, 1993). How well have these hypotheses weathered the ensuing decades? Before answering this question, we first consider current insights into the causes and consequences of cellular senescence. Like Lorenzini et al., we limit our discussion to mammals. We also focus on fibroblasts, the cell type studied by Lorenzini et al., but consider other types as well. We suggest that replicative capacity in culture is not a straightforward assessment, and that it correlates poorly with both longevity and body mass. We speculate this is due to the malleable and variable nature of replicative capacity, which renders it an indirect metric of qualitative and quantitative differences among cells to undergo senescence, a response that directly alters cellular phenotype and might indirectly alter tissue structure and function.« less

Effects of microgravity or simulated launch on testicular function in rats

NASA Technical Reports Server (NTRS)

Amann, R. P.; Deaver, D. R.; Zirkin, B. R.; Grills, G. S.; Sapp, W. J.; Veeramachaneni, D. N. R.; Clemens, J. W.; Banerjee, S. D.; Folmer, J.; Gruppi, C. M.

1992-01-01

Reproductive toxicology and cellular and molecular biology approaches were used to evaluate testicular function in rats from Cosmos 2044. It is found that concentrations of testosterone in testicular tissue or peripheral blood plasma were reduced in flight rates to less than 20 percent of values for simulated-launch or vivarium controls. Spermatogenesis was essentially normal in flight rats, but production of testosterone was severely depressed.

Biological effects of weightlessness and clinostatic conditions registered in cells of root meristem and cap of higher plants

NASA Astrophysics Data System (ADS)

Sytnik, K. M.; Kordyum, E. L.; Belyavskaya, N. A.; Nedukha, E. M.; Tarasenko, V. A.

Research in cellular reproduction, differentiation and vital activity, i.e. processes underlying the development and functioning of organisms, plants included, is essential for solving fundamental and applied problems of space biology. Detailed anatomical analysis of roots of higher plants grown on board the Salyut 6 orbital research station show that under conditions of weightlessness for defined duration mitosis, cytokinesis and tissue differentiation in plant vegetative organs occur essentially normally. At the same time, certain rearrangements in the structural organization of cellular organelles - mainly the plastid apparatus, mitochondria, Golgi apparatus and nucleus - are established in the root meristem and cap of the experimental plants. This is evidence for considerable changes in cellular metabolism. The structural changes in the subcellular level arising under spaceflight conditions are partially absent in clinostat experiments designed to simulate weightlessness. Various clinostatic conditions have different influences on the cell structural and functional organization than does space flight. It is suggested that alterations of cellular metabolism under weightlessness and clinostatic conditions occur within existing genetic programs.

Mesenchymal Stem and Progenitor Cells in Normal and Dysplastic Hematopoiesis—Masters of Survival and Clonality?

PubMed Central

Pleyer, Lisa; Valent, Peter; Greil, Richard

2016-01-01

Myelodysplastic syndromes (MDS) are malignant hematopoietic stem cell disorders that have the capacity to progress to acute myeloid leukemia (AML). Accumulating evidence suggests that the altered bone marrow (BM) microenvironment in general, and in particular the components of the stem cell niche, including mesenchymal stem cells (MSCs) and their progeny, play a pivotal role in the evolution and propagation of MDS. We here present an overview of the role of MSCs in the pathogenesis of MDS, with emphasis on cellular interactions in the BM microenvironment and related stem cell niche concepts. MSCs have potent immunomodulatory capacities and communicate with diverse immune cells, but also interact with various other cellular components of the microenvironment as well as with normal and leukemic stem and progenitor cells. Moreover, compared to normal MSCs, MSCs in MDS and AML often exhibit altered gene expression profiles, an aberrant phenotype, and abnormal functional properties. These alterations supposedly contribute to the “reprogramming” of the stem cell niche into a disease-permissive microenvironment where an altered immune system, abnormal stem cell niche interactions, and an impaired growth control lead to disease progression. The current article also reviews molecular targets that play a role in such cellular interactions and possibilities to interfere with abnormal stem cell niche interactions by using specific targeted drugs. PMID:27355944

Mesenchymal Stem and Progenitor Cells in Normal and Dysplastic Hematopoiesis-Masters of Survival and Clonality?

PubMed

Pleyer, Lisa; Valent, Peter; Greil, Richard

2016-06-27

Myelodysplastic syndromes (MDS) are malignant hematopoietic stem cell disorders that have the capacity to progress to acute myeloid leukemia (AML). Accumulating evidence suggests that the altered bone marrow (BM) microenvironment in general, and in particular the components of the stem cell niche, including mesenchymal stem cells (MSCs) and their progeny, play a pivotal role in the evolution and propagation of MDS. We here present an overview of the role of MSCs in the pathogenesis of MDS, with emphasis on cellular interactions in the BM microenvironment and related stem cell niche concepts. MSCs have potent immunomodulatory capacities and communicate with diverse immune cells, but also interact with various other cellular components of the microenvironment as well as with normal and leukemic stem and progenitor cells. Moreover, compared to normal MSCs, MSCs in MDS and AML often exhibit altered gene expression profiles, an aberrant phenotype, and abnormal functional properties. These alterations supposedly contribute to the "reprogramming" of the stem cell niche into a disease-permissive microenvironment where an altered immune system, abnormal stem cell niche interactions, and an impaired growth control lead to disease progression. The current article also reviews molecular targets that play a role in such cellular interactions and possibilities to interfere with abnormal stem cell niche interactions by using specific targeted drugs.

Plasma and cellular fibronectin: distinct and independent functions during tissue repair

PubMed Central

2011-01-01

Fibronectin (FN) is a ubiquitous extracellular matrix (ECM) glycoprotein that plays vital roles during tissue repair. The plasma form of FN circulates in the blood, and upon tissue injury, is incorporated into fibrin clots to exert effects on platelet function and to mediate hemostasis. Cellular FN is then synthesized and assembled by cells as they migrate into the clot to reconstitute damaged tissue. The assembly of FN into a complex three-dimensional matrix during physiological repair plays a key role not only as a structural scaffold, but also as a regulator of cell function during this stage of tissue repair. FN fibrillogenesis is a complex, stepwise process that is strictly regulated by a multitude of factors. During fibrosis, there is excessive deposition of ECM, of which FN is one of the major components. Aberrant FN-matrix assembly is a major contributing factor to the switch from normal tissue repair to misregulated fibrosis. Understanding the mechanisms involved in FN assembly and how these interplay with cellular, fibrotic and immune responses may reveal targets for the future development of therapies to regulate aberrant tissue-repair processes. PMID:21923916

Functional response of osteoblasts in functionally gradient titanium alloy mesh arrays processed by 3D additive manufacturing.

PubMed

Nune, K C; Kumar, A; Misra, R D K; Li, S J; Hao, Y L; Yang, R

2017-02-01

We elucidate here the osteoblasts functions and cellular activity in 3D printed interconnected porous architecture of functionally gradient Ti-6Al-4V alloy mesh structures in terms of cell proliferation and growth, distribution of cell nuclei, synthesis of proteins (actin, vinculin, and fibronectin), and calcium deposition. Cell culture studies with pre-osteoblasts indicated that the interconnected porous architecture of functionally gradient mesh arrays was conducive to osteoblast functions. However, there were statistically significant differences in the cellular response depending on the pore size in the functionally gradient structure. The interconnected porous architecture contributed to the distribution of cells from the large pore size (G1) to the small pore size (G3), with consequent synthesis of extracellular matrix and calcium precipitation. The gradient mesh structure significantly impacted cell adhesion and influenced the proliferation stage, such that there was high distribution of cells on struts of the gradient mesh structure. Actin and vinculin showed a significant difference in normalized expression level of protein per cell, which was absent in the case of fibronectin. Osteoblasts present on mesh struts formed a confluent sheet, bridging the pores through numerous cytoplasmic extensions. The gradient mesh structure fabricated by electron beam melting was explored to obtain fundamental insights on cellular activity with respect to osteoblast functions. Copyright © 2016 Elsevier B.V. All rights reserved.

Mitochondrial dysfunction in blood cells from amyotrophic lateral sclerosis patients.

PubMed

Ehinger, Johannes K; Morota, Saori; Hansson, Magnus J; Paul, Gesine; Elmér, Eskil

2015-06-01

Mitochondrial dysfunction is implicated in amyotrophic lateral sclerosis, where the progressive degeneration of motor neurons results in muscle atrophy, paralysis and death. Abnormalities in both central nervous system and muscle mitochondria have previously been demonstrated in patient samples, indicating systemic disease. In this case-control study, venous blood samples were acquired from 24 amyotrophic lateral sclerosis patients and 21 age-matched controls. Platelets and peripheral blood mononuclear cells were isolated and mitochondrial oxygen consumption measured in intact and permeabilized cells with additions of mitochondrial substrates, inhibitors and titration of an uncoupler. Respiratory values were normalized to cell count and for two markers of cellular mitochondrial content, citrate synthase activity and mitochondrial DNA, respectively. Mitochondrial function was correlated with clinical staging of disease severity. Complex IV (cytochrome c-oxidase)-activity normalized to mitochondrial content was decreased in platelets from amyotrophic lateral sclerosis patients both when normalized to citrate synthase activity and mitochondrial DNA copy number. In mononuclear cells, complex IV-activity was decreased when normalized to citrate synthase activity. Mitochondrial content was increased in amyotrophic lateral sclerosis patient platelets. In mononuclear cells, complex I activity declined and mitochondrial content increased progressively with advancing disease stage. The findings are, however, based on small subsets of patients and need to be confirmed. We conclude that when normalized to mitochondria-specific content, complex IV-activity is reduced in blood cells from amyotrophic lateral sclerosis patients and that there is an apparent compensatory increase in cellular mitochondrial content. This supports systemic involvement in amyotrophic lateral sclerosis and suggests further study of mitochondrial function in blood cells as a future biomarker for the disease.

Characterization and functional analysis of cellular immunity in mice with biotinidase deficiency.

PubMed

Pindolia, Kirit; Li, Hong; Cardwell, Cisley; Wolf, Barry

2014-05-01

Biotinidase deficiency is an autosomal recessively inherited metabolic disorder that can be easily and effectively treated with pharmacological doses of the vitamin, biotin. Untreated children with profound biotinidase deficiency may exhibit neurological, cutaneous and cellular immunological abnormalities, specifically candida infections. To better understand the immunological dysfunction in some symptomatic individuals with biotinidase deficiency, we studied various aspects of immunological function in a genetically engineered knock-out mouse with biotinidase deficiency. The mouse has no detectable biotinidase activity and develops neurological and cutaneous symptoms similar to those seen in symptomatic children with the disorder. Mice with profound biotinidase deficiency on a biotin-restricted diet had smaller thymuses and spleens than identical mice fed a biotin-replete diet or wildtype mice on either diet; however, the organ to body weight ratios were not significantly different. Thymus histology was normal. Splenocyte subpopulation study showed a significant increase in CD4 positive cells. In addition, in vitro lymphocyte proliferation assays consistently showed diminished proliferation in response to various immunological stimuli. Not all symptomatic individuals with profound biotinidase deficiency develop immunological dysfunction; however, our results do show significant alterations in cellular immunological function that may contribute and/or provide a mechanism(s) for the cellular immunity abnormalities in individuals with biotinidase deficiency. Copyright © 2014 Elsevier Inc. All rights reserved.

Neutrophil degranulation and immunosuppression in patients with GBM: restoration of cellular immune function by targeting arginase I.

PubMed

Sippel, Trisha R; White, Jason; Nag, Kamalika; Tsvankin, Vadim; Klaassen, Marci; Kleinschmidt-DeMasters, B K; Waziri, Allen

2011-11-15

The source of glioblastoma (GBM)-associated immunosuppression remains multifactorial. We sought to clarify and therapeutically target myeloid cell-derived peripheral immunosuppression in patients with GBM. Direct ex vivo T-cell function, serum Arginase I (ArgI) levels, and circulating myeloid lineage populations were compared between patients with GBM and normal donors or patients with other intracranial tumors. Immunofunctional assays were conducted using bulk and sorted cell populations to explore the potential transfer of myeloid cell-mediated immunosuppression and to identify a potential mechanism for these effects. ArgI-mediated immunosuppression was therapeutically targeted in vitro through pharmacologic inhibition or arginine supplementation. We identified a significantly expanded population of circulating, degranulated neutrophils associated with elevated levels of serum ArgI and decreased T-cell CD3ζ expression within peripheral blood from patients with GBM. Sorted CD11b(+) cells from patients with GBM were found to markedly suppress normal donor T-cell function in coculture, and media harvested from mitogen-stimulated GBM peripheral blood mononuclear cell (PBMC) or GBM-associated mixed lymphoid reactions showed ArgI levels that were significantly higher than controls. Critically, T-cell suppression in both settings could be completely reversed through pharmacologic ArgI inhibition or with arginine supplementation. These data indicate that peripheral cellular immunosuppression in patients with GBM is associated with neutrophil degranulation and elevated levels of circulating ArgI, and that T-cell function can be restored in these individuals by targeting ArgI. These data identify a novel pathway of GBM-mediated suppression of cellular immunity and offer a potential therapeutic window for improving antitumor immunity in affected patients.

Lysophosphatidylcholine acyltransferase1 overexpression promotes oral squamous cell carcinoma progression via enhanced biosynthesis of platelet-activating factor.

PubMed

Shida-Sakazume, Tomomi; Endo-Sakamoto, Yosuke; Unozawa, Motoharu; Fukumoto, Chonji; Shimada, Ken; Kasamatsu, Atsushi; Ogawara, Katsunori; Yokoe, Hidetaka; Shiiba, Masashi; Tanzawa, Hideki; Uzawa, Katsuhiro

2015-01-01

The relevance of lysophosphatidylcholine acyltransferase1 (LPCAT1), a cytosolic enzyme in the remodeling pathway of phosphatidylcholine metabolism, in oral squamous cell carcinoma (OSCC) is unknown. We investigated LPCAT1 expression and its functional mechanism in OSCCs. We analyzed LPCAT1 mRNA and protein expression levels in OSCC-derived cell lines. Immunohistochemistry was performed to identify correlations between LPCAT1 expression levels and primary OSCCs clinicopathological status. We established LPCAT1 knockdown models of the OSCC-derived cell lines (SAS, Ca9-22) for functional analysis and examined the association between LPCAT1 expression and the platelet-activating factor (PAF) concentration and PAF-receptor (PAFR) expression. LPCAT1 mRNA and protein were up-regulated significantly (p<0.05) in OSCC-derived cell lines compared with human normal oral keratinocytes. Immunohistochemistry showed significantly (p<0.05) elevated LPCAT1 expression in primary OSCCs compared with normal counterparts and a strong correlation between LPCAT1-positive OSCCs and tumoral size and regional lymph node metastasis. In LPCAT1 knockdown cells, cellular proliferation and invasiveness decreased significantly (p<0.05); cellular migration was inhibited compared with control cells. Down-regulation of LPCAT1 resulted in a decreased intercellular PAF concentration and PAFR expression. LPCAT1 was overexpressed in OSCCs and correlated with cellular invasiveness and migration. LPCAT1 may contribute to tumoral growth and metastasis in oral cancer.

Autophagy in the eye: implications for ocular cell health.

PubMed

Frost, Laura S; Mitchell, Claire H; Boesze-Battaglia, Kathleen

2014-07-01

Autophagy, a catabolic process by which a cell "eats" itself, turning over its own cellular constituents, plays a key role in cellular homeostasis. In an effort to maintain normal cellular function, autophagy is often up-regulated in response to environmental stresses and excessive organelle damage to facilitate aggregated protein removal. In the eye, virtually all cell types from those comprising the cornea in the front of the eye to the retinal pigment epithelium (RPE) providing a protective barrier for the retina at the back of the eye, rely on one or more aspects of autophagy to maintain structure and/or normal physiological function. In the lens autophagy plays a critical role in lens fiber cell maturation and the formation of the organelle free zone. Numerous studies delineating the role of Atg5, Vsp34 as well as FYCO1 in maintenance of lens transparency are discussed. Corneal endothelial dystrophies are also characterized as having elevated levels of autophagic proteins. Therefore, novel modulators of autophagy such as lithium and melatonin are proposed as new therapeutic strategies for this group of dystrophies. In addition, we summarize how corneal Herpes Simplex Virus (HSV-1) infection subverts the cornea's response to infection by inhibiting the normal autophagic response. Using glaucoma models we analyze the relative contribution of autophagy to cell death and cell survival. The cytoprotective role of autophagy is further discussed in an analysis of photoreceptor cell heath and function. We focus our analysis on the current understanding of autophagy in photoreceptor and RPE health, specifically on the diverse role of autophagy in rods and cones as well as its protective role in light induced degeneration. Lastly, in the RPE we highlight hybrid phagocytosis-autophagy pathways. This comprehensive review allows us to speculate on how alterations in various stages of autophagy contribute to glaucoma and retinal degenerations. Copyright © 2014 Elsevier Ltd. All rights reserved.

Follicular localization of growth differentiation factor 8 and its receptors in normal and polycystic ovary syndrome ovaries.

PubMed

Lin, Ting-Ting; Chang, Hsun-Ming; Hu, Xiao-Ling; Leung, Peter C K; Zhu, Yi-Min

2018-05-01

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age and its etiology has not been characterized. Growth differentiation factor 8 (GDF8) is a member of the transforming growth factor-β superfamily that plays a critical role in the regulation of ovarian functions. However, the expression pattern of GDF8 in the human ovary is not yet clear. This study examined the cellular distribution of GDF8 and its putative cellular receptors (ACVR2A, ACVR2B, and ALK5) in a series of normal (n = 34) and PCOS ovaries (n = 14). The immunostaining of GDF8, ACVR2A, ACVR2B, and ALK5 was detected in the oocytes regardless of the developmental stage. All these proteins were localized in antral follicles in normal and PCOS ovaries, and the expression of these proteins increased with increasing follicle diameter. A significantly higher expression of GDF8 was detected in the granulosa cells than in the matched theca cells (TCs). These proteins were also localized in the luteal cells of the corpus luteum. Granulosa cells and TCs of large antral follicles in PCOS ovaries display a higher expression of these proteins. The higher expression levels of GDF8 and its functional receptors (ACVR2A, ACVR2B, and ALK5) in antral follicles of PCOS ovaries than those in normal ovaries suggest the possible involvement of dysregulated GDF8 in the pathogenesis of PCOS.

Role of Vitamin C in the Function of the Vascular Endothelium

PubMed Central

Harrison, Fiona E.

2013-01-01

Abstract Significance: Vitamin C, or ascorbic acid, has long been known to participate in several important functions in the vascular bed in support of endothelial cells. These functions include increasing the synthesis and deposition of type IV collagen in the basement membrane, stimulating endothelial proliferation, inhibiting apoptosis, scavenging radical species, and sparing endothelial cell-derived nitric oxide to help modulate blood flow. Although ascorbate may not be able to reverse inflammatory vascular diseases such as atherosclerosis, it may well play a role in preventing the endothelial dysfunction that is the earliest sign of many such diseases. Recent Advances: Beyond simply preventing scurvy, evidence is mounting that ascorbate is required for optimal function of many dioxygenase enzymes in addition to those involved in collagen synthesis. Several of these enzymes regulate the transcription of proteins involved in endothelial function, proliferation, and survival, including hypoxia-inducible factor-1α and histone and DNA demethylases. More recently, ascorbate has been found to acutely tighten the endothelial permeability barrier and, thus, may modulate access of ascorbate and other molecules into tissues and organs. Critical Issues: The issue of the optimal cellular content of ascorbate remains unresolved, but it appears that low millimolar ascorbate concentrations are normal in most animal tissues, in human leukocytes, and probably in the endothelium. Although there may be little benefit of increasing near maximal cellular ascorbate concentrations in normal people, many diseases and conditions have either systemic or localized cellular ascorbate deficiency as a cause for endothelial dysfunction, including early atherosclerosis, sepsis, smoking, and diabetes. Future Directions: A key focus for future studies of ascorbate and the vascular endothelium will likely be to determine the mechanisms and clinical relevance of ascorbate effects on endothelial function, permeability, and survival in diseases that cause endothelial dysfunction. Antioxid. Redox Signal. 19, 2068–2083. PMID:23581713

Intestinal absorption of water-soluble vitamins in health and disease.

PubMed

Said, Hamid M

2011-08-01

Our knowledge of the mechanisms and regulation of intestinal absorption of water-soluble vitamins under normal physiological conditions, and of the factors/conditions that affect and interfere with theses processes has been significantly expanded in recent years as a result of the availability of a host of valuable molecular/cellular tools. Although structurally and functionally unrelated, the water-soluble vitamins share the feature of being essential for normal cellular functions, growth and development, and that their deficiency leads to a variety of clinical abnormalities that range from anaemia to growth retardation and neurological disorders. Humans cannot synthesize water-soluble vitamins (with the exception of some endogenous synthesis of niacin) and must obtain these micronutrients from exogenous sources. Thus body homoeostasis of these micronutrients depends on their normal absorption in the intestine. Interference with absorption, which occurs in a variety of conditions (e.g. congenital defects in the digestive or absorptive system, intestinal disease/resection, drug interaction and chronic alcohol use), leads to the development of deficiency (and sub-optimal status) and results in clinical abnormalities. It is well established now that intestinal absorption of the water-soluble vitamins ascorbate, biotin, folate, niacin, pantothenic acid, pyridoxine, riboflavin and thiamin is via specific carrier-mediated processes. These processes are regulated by a variety of factors and conditions, and the regulation involves transcriptional and/or post-transcriptional mechanisms. Also well recognized now is the fact that the large intestine possesses specific and efficient uptake systems to absorb a number of water-soluble vitamins that are synthesized by the normal microflora. This source may contribute to total body vitamin nutrition, and especially towards the cellular nutrition and health of the local colonocytes. The present review aims to outline our current understanding of the mechanisms involved in intestinal absorption of water-soluble vitamins, their regulation, the cell biology of the carriers involved and the factors that negatively affect these absorptive events. © The Authors Journal compilation © 2011 Biochemical Society

Intestinal absorption of water-soluble vitamins in health and disease

PubMed Central

Said, Hamid M.

2014-01-01

Our knowledge of the mechanisms and regulation of intestinal absorption of water-soluble vitamins under normal physiological conditions, and of the factors/conditions that affect and interfere with theses processes has been significantly expanded in recent years as a result of the availability of a host of valuable molecular/cellular tools. Although structurally and functionally unrelated, the water-soluble vitamins share the feature of being essential for normal cellular functions, growth and development, and that their deficiency leads to a variety of clinical abnormalities that range from anaemia to growth retardation and neurological disorders. Humans cannot synthesize water-soluble vitamins (with the exception of some endogenous synthesis of niacin) and must obtain these micronutrients from exogenous sources. Thus body homoeostasis of these micronutrients depends on their normal absorption in the intestine. Interference with absorption, which occurs in a variety of conditions (e.g. congenital defects in the digestive or absorptive system, intestinal disease/resection, drug interaction and chronic alcohol use), leads to the development of deficiency (and sub-optimal status) and results in clinical abnormalities. It is well established now that intestinal absorption of the water-soluble vitamins ascorbate, biotin, folate, niacin, pantothenic acid, pyridoxine, riboflavin and thiamin is via specific carrier-mediated processes. These processes are regulated by a variety of factors and conditions, and the regulation involves transcriptional and/or post-transcriptional mechanisms. Also well recognized now is the fact that the large intestine possesses specific and efficient uptake systems to absorb a number of water-soluble vitamins that are synthesized by the normal microflora. This source may contribute to total body vitamin nutrition, and especially towards the cellular nutrition and health of the local colonocytes. The present review aims to outline our current understanding of the mechanisms involved in intestinal absorption of water-soluble vitamins, their regulation, the cell biology of the carriers involved and the factors that negatively affect these absorptive events. PMID:21749321

Inhibition of AMPK catabolic action by GSK3

PubMed Central

Suzuki, Tsukasa; Bridges, Dave; Nakada, Daisuke; Skiniotis, Georgios; Morrison, Sean J.; Lin, Jiandie; Saltiel, Alan R.; Inoki, Ken

2013-01-01

SUMMARY AMP-activated protein kinase (AMPK) regulates cellular energy homeostasis by inhibiting anabolic and activating catabolic processes. While AMPK activation has been extensively studied, mechanisms that inhibit AMPK remain elusive. Here we report that glycogen synthase kinase 3 (GSK3) inhibits AMPK function. GSK3 forms a stable complex with AMPK through interactions with the AMPK β regulatory subunit and phosphorylates the AMPK α catalytic subunit. This phosphorylation enhances the accessibility of the activation loop of the α subunit to phosphatases, thereby inhibiting AMPK kinase activity. Surprisingly, PI3K-Akt signaling, which is a major anabolic signaling and normally inhibits GSK3 activity, promotes GSK3 phosphorylation and inhibition of AMPK, thus revealing how AMPK senses anabolic environments in addition to cellular energy levels. Consistently, disrupting GSK3 function within the AMPK complex sustains higher AMPK activity and cellular catabolic processes even under anabolic conditions, indicating that GSK3 acts as a critical sensor for anabolic signaling to regulate AMPK. PMID:23623684

Long-term dopamine transporter expression and normal cellular distribution of mitochondria in dopaminergic neuron transplants in Parkinson’s disease patients

PubMed Central

Hallett, Penelope J; Cooper, Oliver; Sadi, Damaso; Robertson, Harold; Mendez, Ivar; Isacson, Ole

2014-01-01

Summary To determine the long-term health and function of transplanted dopamine neurons in Parkinson’s disease (PD) patients, the expression of dopamine transporters (DAT) and mitochondrial morphology was examined in human fetal midbrain cellular transplants. DAT was robustly expressed in transplanted dopamine neuron terminals in the reinnervated host putamen and caudate, for at least 14 years after transplantation. The transplanted dopamine neurons showed a healthy and non-atrophied morphology at all time points. Labeling of the mitochondrial outer membrane protein Tom20 and alpha-synuclein showed typical cellular pathology in the patients’ own substantia nigra, which was not observed in transplanted dopamine neurons. These results show that the vast majority of transplanted neurons remain healthy long-term in PD patients, consistent with the clinically maintained function of fetal dopamine neuron transplants for up to 15–18 years in patients. These findings are critically important for the rational development of stem cell-based dopamine neuronal replacement therapies for PD. PMID:24910427

Cellular therapy in bone-tendon interface regeneration

PubMed Central

Rothrauff, Benjamin B; Tuan, Rocky S

2014-01-01

The intrasynovial bone-tendon interface is a gradual transition from soft tissue to bone, with two intervening zones of uncalcified and calcified fibrocartilage. Following injury, the native anatomy is not restored, resulting in inferior mechanical properties and an increased risk of re-injury. Recent in vivo studies provide evidence of improved healing when surgical repair of the bone-tendon interface is augmented with cells capable of undergoing chondrogenesis. In particular, cellular therapy in bone-tendon healing can promote fibrocartilage formation and associated improvements in mechanical properties. Despite these promising results in animal models, cellular therapy in human patients remains largely unexplored. This review highlights the development and structure-function relationship of normal bone-tendon insertions. The natural healing response to injury is discussed, with subsequent review of recent research on cellular approaches for improved healing. Finally, opportunities for translating in vivo findings into clinical practice are identified. PMID:24326955

Rapid computation of photoacoustic fields from normal and pathological red blood cells using a Green's function method

NASA Astrophysics Data System (ADS)

Saha, Ratan K.; Fadhel, Muhannad N.; Lawrence, Aamna; Karmakar, Subhajit; Adhikari, Arunabha; Kolios, Michael C.

2017-03-01

Photoacoustic (PA) field calculations using a Green's function approach is presented. The method has been applied to predict PA spectra generated by normal (discocyte) and pathological (stomatocyte) red blood cells (RBCs). The contours of normal and pathological RBCs were generated by employing a popular parametric model and accordingly, fitted with the Legendre polynomial expansions for surface parametrization. The first frequency minimum of theoretical PA spectrum approximately appears at 607 MHz for a discocyte and 410 MHz for a stomatocyte when computed from the direction of symmetry axis. The same feature occurs nearly at 247 and 331 MHz, respectively, for those particles when measured along the perpendicular direction. The average experimental spectrum for normal RBCs is found to be flat over a bandwidth of 150-500 MHz when measured along the direction of symmetry axis. For spherical RBCs, both the theoretical and experimental spectra demonstrate negative slope over a bandwidth of 250-500 MHz. Using the Green's function method discussed, it may be possible to rapidly characterize cellular morphology from single-particle PA spectra.

Alternative Conformations of Cytochrome c: Structure, Function, and Detection.

PubMed

Hannibal, Luciana; Tomasina, Florencia; Capdevila, Daiana A; Demicheli, Verónica; Tórtora, Verónica; Alvarez-Paggi, Damián; Jemmerson, Ronald; Murgida, Daniel H; Radi, Rafael

2016-01-26

Cytochrome c (cyt c) is a cationic hemoprotein of ∼100 amino acid residues that exhibits exceptional functional versatility. While its primary function is electron transfer in the respiratory chain, cyt c is also recognized as a key component of the intrinsic apoptotic pathway, the mitochondrial oxidative protein folding machinery, and presumably as a redox sensor in the cytosol, along with other reported functions. Transition to alternative conformations and gain-of-peroxidase activity are thought to further enable the multiple functions of cyt c and its translocation across cellular compartments. In vitro, direct interactions of cyt c with cardiolipin, post-translational modifications such as tyrosine nitration, phosphorylation, methionine sulfoxidation, mutations, and even fine changes in electrical fields lead to a variety of conformational states that may be of biological relevance. The identification of these alternative conformations and the elucidation of their functions in vivo continue to be a major challenge. Here, we unify the knowledge of the structural flexibility of cyt c that supports functional moonlighting and review biochemical and immunochemical evidence confirming that cyt c undergoes conformational changes during normal and altered cellular homeostasis.

A new treatment for human malignant melanoma targeting L-type amino acid transporter 1 (LAT1): A pilot study in a canine model

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

Fukumoto, Shinya; Hanazono, Kiwamu; Fu, Dah-Renn

2013-09-13

Highlights: •LAT1 is highly expressed in tumors but at low levels in normal tissues. •We examine LAT1 expression and function in malignant melanoma (MM). •LAT1 expression in MM tissues and cell lines is higher than those in normal tissues. •LAT1 selective inhibitors inhibit amino acid uptake and cell growth in MM cells. •New chemotherapeutic protocols including LAT1 inhibitors are effective for treatment. -- Abstract: L-type amino acid transporter 1 (LAT1), an isoform of amino acid transport system L, transports branched or aromatic amino acids essential for fundamental cellular activities such as cellular growth, proliferation and maintenance. This amino acid transportermore » recently has received attention because of its preferential and up-regulated expression in a variety of human tumors in contrast to its limited distribution and low-level expression in normal tissues. In this study, we explored the feasibility of using LAT1 inhibitor as a new therapeutic agent for human malignant melanomas (MM) using canine spontaneous MM as a model for human MM. A comparative study of LAT expression was performed in 48 normal tissues, 25 MM tissues and five cell lines established from MM. The study observed LAT1 mRNA levels from MM tissues and cell lines that were significantly (P < 0.01) higher than in normal tissues. Additionally, MM with distant metastasis showed a higher expression than those without distant metastasis. Functional analysis of LAT1 was performed on one of the five cell lines, CMeC-1. [{sup 3}H]L-Leucine uptake and cellular growth activities in CMeC-1 were inhibited in a dose-dependent manner by selective LAT1 inhibitors (2-amino-2-norbornane-carboxylic acid, BCH and melphalan, LPM). Inhibitory growth activities of various conventional anti-cancer drugs, including carboplatin, cyclophosphamide, dacarbazine, doxorubicin, mitoxantrone, nimustine, vinblastine and vincristine, were significantly (P < 0.05) enhanced by combination use with BCH or LPM. These findings suggest that LAT1 could be a new therapeutic target for MM.« less

Beneficial effects of leptin treatment in a setting of cardiac dysfunction induced by transverse aortic constriction in mouse.

PubMed

Gómez-Hurtado, Nieves; Domínguez-Rodríguez, Alejandro; Mateo, Philippe; Fernández-Velasco, María; Val-Blasco, Almudena; Aizpún, Rafael; Sabourin, Jessica; Gómez, Ana María; Benitah, Jean-Pierre; Delgado, Carmen

2017-07-01

Leptin, is a 16 kDa pleiotropic peptide not only primarily secreted by adipocytes, but also produced by other tissues, including the heart. Controversy exists regarding the adverse and beneficial effects of leptin on the heart We analysed the effect of a non-hypertensive dose of leptin on cardiac function, [Ca 2+ ] i handling and cellular electrophysiology, which participate in the genesis of pump failure and related arrhythmias, both in control mice and in mice subjected to chronic pressure-overload by transverse aorta constriction. We find that leptin activates mechanisms that contribute to cardiac dysfunction under physiological conditions. However, after the establishment of pressure overload, an increase in leptin levels has protective cardiac effects with respect to rescuing the cellular heart failure phenotype. These beneficial effects of leptin involve restoration of action potential duration via normalization of transient outward potassium current and sarcoplasmic reticulum Ca 2+ content via rescue of control sarcoplasmic/endoplasmic reticulum Ca 2+ ATPase levels and ryanodine receptor function modulation, leading to normalization of Ca 2+ handling parameters. Leptin, is a 16 kDa pleiotropic peptide not only primary secreted by adipocytes, but also produced by other tissues, including the heart. Evidence indicates that leptin may have either adverse or beneficial effects on the heart. To obtain further insights, in the present study, we analysed the effect of leptin treatment on cardiac function, [Ca 2+ ] i handling and cellular electrophysiology, which participate in the genesis of pump failure and related arrhythmias, both in control mice and in mice subjected to chronic pressure-overload by transverse aorta constriction (TAC). Three weeks after surgery, animals received either leptin (0.36 mg kg -1  day -1 ) or vehicle via osmotic minipumps for 3 weeks. Echocardiographic measurements showed that, although leptin treatment was deleterious on cardiac function in sham, leptin had a cardioprotective effect following TAC. [Ca 2+ ] i transient in cardiomyocytes followed similar pattern. Patch clamp experiments showed prolongation of action potential duration (APD) in TAC and leptin-treated sham animals, whereas, following TAC, leptin reduced the APD towards control values. APD variations were associated with decreased transient outward potassium current and Kv4.2 and KChIP2 protein expression. TAC myocytes showed a higher incidence of triggered activities and spontaneous Ca 2+ waves. These proarrhythmic manifestations, related to Ca 2+ /calmodulin-dependent protein kinase II and ryanodine receptor phosphorylation, were reduced by leptin. The results of the present study demonstrate that, although leptin treatment was deleterious on cardiac function in control animals, leptin had a cardioprotective effect following TAC, normalizing cardiac function and reducing arrhythmogeneity at the cellular level. © 2017 The Authors. The Journal of Physiology © 2017 The Physiological Society.

Matrix Stiffness Corresponding to Strictured Bowel Induces a Fibrogenic Response in Human Colonic Fibroblasts

PubMed Central

Johnson, Laura A.; Rodansky, Eva S.; Sauder, Kay L.; Horowitz, Jeffrey C.; Mih, Justin D.; Tschumperlin, Daniel J.; Higgins, Peter D.

2013-01-01

Background Crohn’s disease is characterized by repeated cycles of inflammation and mucosal healing which ultimately progress to intestinal fibrosis. This inexorable progression towards fibrosis suggests that fibrosis becomes inflammation-independent and auto-propagative. We hypothesized that matrix stiffness regulates this auto-propagation of intestinal fibrosis. Methods The stiffness of fresh ex vivo samples from normal human small intestine, Crohn’s disease strictures, and the unaffected margin were measured with a microelastometer. Normal human colonic fibroblasts were cultured on physiologically normal or pathologically stiff matrices corresponding to the physiological stiffness of normal or fibrotic bowel. Cellular response was assayed for changes in cell morphology, α-smooth muscle actin (αSMA) staining, and gene expression. Results Microelastometer measurements revealed a significant increase in colonic tissue stiffness between normal human colon and Crohn’s strictures as well as between the stricture and adjacent tissue margin. In Ccd-18co cells grown on stiff matrices corresponding to Crohn’s strictures, cellular proliferation increased. Pathologic stiffness induced a marked change in cell morphology and increased αSMA protein expression. Growth on a stiff matrix induced fibrogenic gene expression, decreased matrix metalloproteinase and pro-inflammatory gene expression, and was associated with nuclear localization of the transcriptional cofactor MRTF-A. Conclusions Matrix stiffness, representative of the pathological stiffness of Crohn’s strictures, activates human colonic fibroblasts to a fibrogenic phenotype. Matrix stiffness affects multiple pathways suggesting the mechanical properties of the cellular environment are critical to fibroblast function and may contribute to autopropagation of intestinal fibrosis in the absence of inflammation, thereby contributing to the intractable intestinal fibrosis characteristic of Crohn’s disease. PMID:23502354

Cellular Signaling by Fibroblast Growth Factors (FGFs) and Their Receptors (FGFRs) in Male Reproduction

PubMed Central

Cotton, Leanne M.; O’Bryan, Moira K.; Hinton, Barry T.

2008-01-01

The major function of the reproductive system is to ensure the survival of the species by passing on hereditary traits from one generation to the next. This is accomplished through the production of gametes and the generation of hormones that function in the maturation and regulation of the reproductive system. It is well established that normal development and function of the male reproductive system is mediated by endocrine and paracrine signaling pathways. Fibroblast growth factors (FGFs), their receptors (FGFRs), and signaling cascades have been implicated in a diverse range of cellular processes including: proliferation, apoptosis, cell survival, chemotaxis, cell adhesion, motility, and differentiation. The maintenance and regulation of correct FGF signaling is evident from human and mouse genetic studies which demonstrate that mutations leading to disruption of FGF signaling cause a variety of developmental disorders including dominant skeletal diseases, infertility, and cancer. Over the course of this review, we will provide evidence for differential expression of FGFs/FGFRs in the testis, male germ cells, the epididymis, the seminal vesicle, and the prostate. We will show that this signaling cascade has an important role in sperm development and maturation. Furthermore, we will demonstrate that FGF/FGFR signaling is essential for normal epididymal function and prostate development. To this end, we will provide evidence for the involvement of the FGF signaling system in the regulation and maintenance of the male reproductive system. PMID:18216218

Cellular signaling by fibroblast growth factors (FGFs) and their receptors (FGFRs) in male reproduction.

PubMed

Cotton, Leanne M; O'Bryan, Moira K; Hinton, Barry T

2008-04-01

The major function of the reproductive system is to ensure the survival of the species by passing on hereditary traits from one generation to the next. This is accomplished through the production of gametes and the generation of hormones that function in the maturation and regulation of the reproductive system. It is well established that normal development and function of the male reproductive system is mediated by endocrine and paracrine signaling pathways. Fibroblast growth factors (FGFs), their receptors (FGFRs), and signaling cascades have been implicated in a diverse range of cellular processes including: proliferation, apoptosis, cell survival, chemotaxis, cell adhesion, motility, and differentiation. The maintenance and regulation of correct FGF signaling is evident from human and mouse genetic studies which demonstrate that mutations leading to disruption of FGF signaling cause a variety of developmental disorders including dominant skeletal diseases, infertility, and cancer. Over the course of this review, we will provide evidence for differential expression of FGFs/FGFRs in the testis, male germ cells, the epididymis, the seminal vesicle, and the prostate. We will show that this signaling cascade has an important role in sperm development and maturation. Furthermore, we will demonstrate that FGF/FGFR signaling is essential for normal epididymal function and prostate development. To this end, we will provide evidence for the involvement of the FGF signaling system in the regulation and maintenance of the male reproductive system.

Pirin Inhibits Cellular Senescence in Melanocytic Cells

PubMed Central

Licciulli, Silvia; Luise, Chiara; Scafetta, Gaia; Capra, Maria; Giardina, Giuseppina; Nuciforo, Paolo; Bosari, Silvano; Viale, Giuseppe; Mazzarol, Giovanni; Tonelli, Chiara; Lanfrancone, Luisa; Alcalay, Myriam

2011-01-01

Cellular senescence has been widely recognized as a tumor suppressing mechanism that acts as a barrier to cancer development after oncogenic stimuli. A prominent in vivo model of the senescence barrier is represented by nevi, which are composed of melanocytes that, after an initial phase of proliferation induced by activated oncogenes (most commonly BRAF), are blocked in a state of cellular senescence. Transformation to melanoma occurs when genes involved in controlling senescence are mutated or silenced and cells reacquire the capacity to proliferate. Pirin (PIR) is a highly conserved nuclear protein that likely functions as a transcriptional regulator whose expression levels are altered in different types of tumors. We analyzed the expression pattern of PIR in adult human tissues and found that it is expressed in melanocytes and has a complex pattern of regulation in nevi and melanoma: it is rarely detected in mature nevi, but is expressed at high levels in a subset of melanomas. Loss of function and overexpression experiments in normal and transformed melanocytic cells revealed that PIR is involved in the negative control of cellular senescence and that its expression is necessary to overcome the senescence barrier. Our results suggest that PIR may have a relevant role in melanoma progression. PMID:21514450

Defective granulocyte chemotaxis in the Chediak-Higashi syndrome

PubMed Central

Clark, Robert A.; Kimball, Harry R.

1971-01-01

In vivo and in vitro studies of granulocyte chemotaxis were performed in three patients with the Chediak-Higashi syndrome. Rebuck skin windows showed a decreased accumulation of leukocytes at an inflammatory site. Studies in Boyden chambers documented a cellular defect in granulocyte chemotaxis. The chemotactic response of Chediak-Higashi cells by this technique averaged approximately 40% of normal and was consistently reduced using several different chemotactic stimuli. This deficit was magnified by shortening the chamber incubation time or by decreasing the pore size of the micropore filter and was independent of granulocytopenia. No abnormalities of passive motility, adhesiveness, viability, or pH optimum for migration were found in these cells. Chediak-Higashi serum contained no inhibitors of chemotaxis and was capable of generating normal amounts of chemotactic factors with the exception of one patient with the accelerated phase of the disease. Heterozygotes for the Chediak-Higashi trait had normal chemotactic function. This cellular defect in chemotaxis may contribute to the marked susceptibility to pyogenic infections which is so characteristic of patients with the Chediak-Higashi syndrome. Images PMID:4942966

Endogenous sterol biosynthesis is important for mitochondrial function and cell morphology in procyclic forms of Trypanosoma brucei.

PubMed

Pérez-Moreno, Guiomar; Sealey-Cardona, Marco; Rodrigues-Poveda, Carlos; Gelb, Michael H; Ruiz-Pérez, Luis Miguel; Castillo-Acosta, Víctor; Urbina, Julio A; González-Pacanowska, Dolores

2012-10-01

Sterol biosynthesis inhibitors are promising entities for the treatment of trypanosomal diseases. Insect forms of Trypanosoma brucei, the causative agent of sleeping sickness, synthesize ergosterol and other 24-alkylated sterols, yet also incorporate cholesterol from the medium. While sterol function has been investigated by pharmacological manipulation of sterol biosynthesis, molecular mechanisms by which endogenous sterols influence cellular processes remain largely unknown in trypanosomes. Here we analyse by RNA interference, the effects of a perturbation of three specific steps of endogenous sterol biosynthesis in order to dissect the role of specific intermediates in proliferation, mitochondrial function and cellular morphology in procyclic cells. A decrease in the levels of squalene synthase and squalene epoxidase resulted in a depletion of cellular sterol intermediates and end products, impaired cell growth and led to aberrant morphologies, DNA fragmentation and a profound modification of mitochondrial structure and function. In contrast, cells deficient in sterol methyl transferase, the enzyme involved in 24-alkylation, exhibited a normal growth phenotype in spite of a complete abolition of the synthesis and content of 24-alkyl sterols. Thus, the data provided indicates that while the depletion of squalene and post-squalene endogenous sterol metabolites results in profound cellular defects, bulk 24-alkyl sterols are not strictly required to support growth in insect forms of T. brucei in vitro. Copyright © 2012 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Comparative studies of cellular viability levels on 2D and 3D in vitro culture matrices.

PubMed

Gargotti, M; Lopez-Gonzalez, U; Byrne, H J; Casey, A

2018-02-01

In this study, the cellular viability and function of immortalized human cervical and dermal cells are monitored and compared in conventional 2D and two commercial 3D membranes, Collagen and Geltrex, of varying working concentration and volume. Viability was monitored with the aid of the Alamar Blue assay, cellular morphology was monitored with confocal microscopy, and cell cycle studies and cell death mechanism studies were performed with flow cytometry. The viability studies showed apparent differences between the 2D and 3D culture systems, the differences attributed in part to the physical transition from 2D to 3D environment causing alterations to effective resazurin concentration, uptake and conversion rates, which was dependent on exposure time, but also due to the effect of the membrane itself on cellular function. These effects were verified by flow cytometry, in which no significant differences in viable cell numbers between 2D and 3D systems were observed after 24 h culture. The results showed the observed effect was different after shorter exposure periods, was also dependent on working concentration of the 3D system and could be mediated by altering the culture vessel size. Cell cycle analysis revealed cellular function could be altered by growth on the 3D substrates and the alterations were noted to be dependent on 3D membrane concentration. The use of 3D culture matrices has been widely interpreted to result in "improved viability levels" or "reduced" toxicity or cellular "resistance" compared to cells cultured on traditional 2D systems. The results of this study show that cellular health and viability levels are not altered by culture in 3D environments, but their normal cycle can be altered as indicated in the cell cycle studies performed and such variations must be accounted for in studies employing 3D membranes for in vitro cellular screening.

The inhibitory effect of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G) and its family members on the activity of cellular microRNAs.

PubMed

Zhang, Hui

2010-01-01

The apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G or APOBEC3G) and its fellow cytidine deaminase family members are potent restrictive factors for human immunodeficiency virus type 1 (HIV-1) and many other retroviruses. However, the cellular function of APOBEC3G remains to be further clarified. It has been reported that APOBEC3s can restrict the mobility of endogenous retroviruses and LTR-retrotransposons, suggesting that they can maintain stability in host genomes. However, APOBEC3G is normally cytoplasmic. Further studies have demonstrated that it is associated with an RNase-sensitive high molecular mass (HMM) and located in processing bodies (P-bodies) of replicating T-cells, indicating that the major cellular function of APOBEC3G seems to be related to P-body-related RNA processing and metabolism. As the function of P-body is closely related to miRNA activity, APOBEC3G could affect the miRNA function. Recent studies have demonstrated that APOBEC3G and its family members counteract miRNA-mediated repression of protein translation. Further, APOBEC3G enhances the association of miRNA-targeted mRNA with polysomes, and facilitates the dissociation of miRNA-targeted mRNA from P-bodies. As such, APOBEC3G regulate the activity of cellular miRNAs. Whether this function is related to its potent antiviral activity remains to be further determined.

A Decade of Boon or Burden: What Has the CHIP Ever Done for Cellular Protein Quality Control Mechanism Implicated in Neurodegeneration and Aging?

PubMed Central

Joshi, Vibhuti; Amanullah, Ayeman; Upadhyay, Arun; Mishra, Ribhav; Kumar, Amit; Mishra, Amit

2016-01-01

Cells regularly synthesize new proteins to replace old and abnormal proteins for normal cellular functions. Two significant protein quality control pathways inside the cellular milieu are ubiquitin proteasome system (UPS) and autophagy. Autophagy is known for bulk clearance of cytoplasmic aggregated proteins, whereas the specificity of protein degradation by UPS comes from E3 ubiquitin ligases. Few E3 ubiquitin ligases, like C-terminus of Hsc70-interacting protein (CHIP) not only take part in protein quality control pathways, but also plays a key regulatory role in other cellular processes like signaling, development, DNA damage repair, immunity and aging. CHIP targets misfolded proteins for their degradation through proteasome, as well as autophagy; simultaneously, with the help of chaperones, it also regulates folding attempts for misfolded proteins. The broad range of CHIP substrates and their associations with multiple pathologies make it a key molecule to work upon and focus for future therapeutic interventions. E3 ubiquitin ligase CHIP interacts and degrades many protein inclusions formed in neurodegenerative diseases. The presence of CHIP at various nodes of cellular protein-protein interaction network presents this molecule as a potential candidate for further research. In this review, we have explored a wide range of functionality of CHIP inside cells by a detailed presentation of its co-chaperone, E3 and E4 enzyme like functions, with central focus on its protein quality control roles in neurodegenerative diseases. We have also raised many unexplored but expected fundamental questions regarding CHIP functions, which generate hopes for its future applications in research, as well as drug discovery. PMID:27757073

AAA+ Machines of Protein Destruction in Mycobacteria.

PubMed

Alhuwaider, Adnan Ali H; Dougan, David A

2017-01-01

The bacterial cytosol is a complex mixture of macromolecules (proteins, DNA, and RNA), which collectively are responsible for an enormous array of cellular tasks. Proteins are central to most, if not all, of these tasks and as such their maintenance (commonly referred to as protein homeostasis or proteostasis) is vital for cell survival during normal and stressful conditions. The two key aspects of protein homeostasis are, (i) the correct folding and assembly of proteins (coupled with their delivery to the correct cellular location) and (ii) the timely removal of unwanted or damaged proteins from the cell, which are performed by molecular chaperones and proteases, respectively. A major class of proteins that contribute to both of these tasks are the AAA+ (ATPases associated with a variety of cellular activities) protein superfamily. Although much is known about the structure of these machines and how they function in the model Gram-negative bacterium Escherichia coli , we are only just beginning to discover the molecular details of these machines and how they function in mycobacteria. Here we review the different AAA+ machines, that contribute to proteostasis in mycobacteria. Primarily we will focus on the recent advances in the structure and function of AAA+ proteases, the substrates they recognize and the cellular pathways they control. Finally, we will discuss the recent developments related to these machines as novel drug targets.

An imidazole functionalized pentameric thiophene displays different staining patterns in normal and malignant cells

NASA Astrophysics Data System (ADS)

Nilsson, Peter; Magnusson, Karin; Appelqvist, Hanna; Cieslar-Pobuda, Artur; Bäck, Marcus; Kågedal, Bertil; Jonasson, Jon; Los, Marek

2015-10-01

Molecular tools for fluorescent imaging of cells and their components are vital for understanding the function and activity of cells. Here, we report an imidazole functionalized pentameric oligothiophene, p-HTIm, that can be utilized for fluorescent imaging of cells. p-HTIm fluorescence in normal cells appeared in a peripheral punctate pattern partially co-localized with lysosomes, whereas a one-sided perinuclear Golgi associated localization of the dye was observed in malignant cells. The uptake of p-HTIm was temperature dependent and the intracellular target was reached within 1 h after staining. The ability of p-HTIm to stain cells was reduced when the imidazole side chain was chemically altered, verifying that specific imidazole side-chain functionalities are necessary for achieving the observed cellular staining. Our findings confirm that properly functionalized oligothiophenes can be utilized as fluorescent tools for vital staining of cells and that the selectivity towards distinct intracellular targets are highly dependent on the side-chain functionalities along the conjugated thiophene backbone.

Are Diadenosine Polyphosphates and/or FHIT Involved in Anoikis?

DTIC Science & Technology

2002-06-01

Ap4A ) in anoikis. These molecules occur in all organisms, accumulate in response to cellular stress, and have quite recently been implicated in...and Ap4A hydrolase, connecting these dinucleotides with cancer. ApnAs probably act as cofactors for Fhit’s effector function (analogous to the function...breast cancer, the purpose of this IDEA project is to determine whether Ap3A/ Ap4A and/or FHIT can regulate anoikis in normal and transformed mammary

Mapping Cellular Polarity Networks Using Mass Spectrometry-based Strategies.

PubMed

Daulat, Avais M; Puvirajesinghe, Tania M; Camoin, Luc; Borg, Jean-Paul

2018-05-18

Cell polarity is a vital biological process involved in the building, maintenance and normal functioning of tissues in invertebrates and vertebrates. Unsurprisingly, molecular defects affecting polarity organization and functions have a strong impact on tissue homeostasis, embryonic development and adult life, and may directly or indirectly lead to diseases. Genetic studies have demonstrated the causative effect of several polarity genes in diseases; however, much remains to be clarified before a comprehensive view of the molecular organization and regulation of the protein networks associated with polarity proteins is obtained. This challenge can be approached head-on using proteomics to identify protein complexes involved in cell polarity and their modifications in a spatio-temporal manner. We review the fundamental basics of mass spectrometry techniques and provide an in-depth analysis of how mass spectrometry has been instrumental in understanding the complex and dynamic nature of some cell polarity networks at the tissue (apico-basal and planar cell polarities) and cellular (cell migration, ciliogenesis) levels, with the fine dissection of the interconnections between prototypic cell polarity proteins and signal transduction cascades in normal and pathological situations. This review primarily focuses on epithelial structures which are the fundamental building blocks for most metazoan tissues, used as the archetypal model to study cellular polarity. This field offers broad perspectives thanks to the ever-increasing sensitivity of mass spectrometry and its use in combination with recently developed molecular strategies able to probe in situ proteomic networks. Copyright © 2018 Elsevier Ltd. All rights reserved.

[Morphology of basement membrane and associated matrix proteins in normal and pathological tissues].

PubMed

Nerlich, A

1995-01-01

Basement membranes (BM) are specialized structures of the extracellular matrix. Their composition is of particular importance for the maintenance of normal morphological and functional properties of a multitude of organs and tissue systems and it is thus required for regular homeostasis of body function. Generally, they possess three main functions, i.e. participation in the maintenance of tissue structure, control of fluid and substrate exchange, and regulation of cell growth and differentiation. BMs are made up by various components which are in part specifically localized within the BM zone, or which represent ubiquitous matrix constituents with specific quantitative and/or qualitative differences in their localization. On the basis of a thorough immunohistochemical analysis of normal and diseased tissues, we provide here a concept of "functional morphology/pathomorphology" of the different BM components analyzed: 1.) The ubiquitous BM-constituent collagen IV primarily stabilizes the BM-zone and thus represents the "backbone" of the BM providing mechanical strength. Its loss leads to cystic tissue transformation as it is evidenced from the analysis of polycystic nephropathies. Thus, in other cystic tissue transformations a similar formal pathogenesis may be present. 2.) The specific localization of collagen VII as the main structural component of anchoring fibrils underlines the mechanical anchoring function of this collagenous protein. Defects in this protein lead to hereditary epidermolysis. The rapid re-occurrence of epidermal collagen VII during normal human wound healing indicates a quick reconstitution of the mechanical tensile strength of healing wounds. 3.) The BM-specific heparan sulfate proteoglycan (HSPG, Perlecan) with its highly negative anionic charge can be assumed to exert filter control. This assumption is corroborated by the localizatory findings of a preferential deposition of HSPG in endothelial and particularly in glomerular BM. Similarly, the lack of HSPG in the BM of lymph capillaries can be regarded as the correlate for a free fluid influx into lymphatic capillaries. The relative reduction in HSPG-staining in the developing glomerular BM also explains the still immature filter function. Furthermore, the low content of HSPG in placental chorionic capillaries can be regarded as morphological correlate for the required free fluid exchange between maternal and fetal blood systems. In diabetic glomerulopathy, the loss of HSPG coincides with a reduced filter function providing further support for the function of the HSPG. In further analyses of diabetic glomerulopathy, we provide evidence for an extensive matrix dysregulation resulting in either the overexpression of certain BM-components (diffuse glomerulosclerosis) or microfibrillar collagen VI (nodular glomerulosclerosis) indicating changes in cell function and possibly also cellular "differentiation". The analysis of congenital nephropathies additionally indicates that also the HSPG side chains with their negative charges may be involved in certain diseases with filter impairment. 4.) Furthermore, HSPG serves as a binding site for growth factors, particularly for the basic fibroblast growth factor (bFGF). It is of particular interest that the localization of HSPG and bFGF is not completely identical indicating some tissue specific differences in the receptor-ligand interaction. The functional importance of the bFGF-HSPG-interaction is exemplified by arteriosclerotic intima lesions where in highly cellular lesions high amounts of bFGF and HSPG coincide and low levels of both appear in poorly cellular lesions. Similarly, the granulation tissue in wound healing contains large amounts of bFGF-positive mesenchymal cells. 5.) The role of individual matrix components can be deduced from the normal human wound healing process, where epithelial cells migrate on a fibronectin matrix without complete BM.

Effects of aging and sensory loss on glial cells in mouse visual and auditory cortices.

PubMed

Tremblay, Marie-Ève; Zettel, Martha L; Ison, James R; Allen, Paul D; Majewska, Ania K

2012-04-01

Normal aging is often accompanied by a progressive loss of receptor sensitivity in hearing and vision, whose consequences on cellular function in cortical sensory areas have remained largely unknown. By examining the primary auditory (A1) and visual (V1) cortices in two inbred strains of mice undergoing either age-related loss of audition (C57BL/6J) or vision (CBA/CaJ), we were able to describe cellular and subcellular changes that were associated with normal aging (occurring in A1 and V1 of both strains) or specifically with age-related sensory loss (only in A1 of C57BL/6J or V1 of CBA/CaJ), using immunocytochemical electron microscopy and light microscopy. While the changes were subtle in neurons, glial cells and especially microglia were transformed in aged animals. Microglia became more numerous and irregularly distributed, displayed more variable cell body and process morphologies, occupied smaller territories, and accumulated phagocytic inclusions that often displayed ultrastructural features of synaptic elements. Additionally, evidence of myelination defects were observed, and aged oligodendrocytes became more numerous and were more often encountered in contiguous pairs. Most of these effects were profoundly exacerbated by age-related sensory loss. Together, our results suggest that the age-related alteration of glial cells in sensory cortical areas can be accelerated by activity-driven central mechanisms that result from an age-related loss of peripheral sensitivity. In light of our observations, these age-related changes in sensory function should be considered when investigating cellular, cortical, and behavioral functions throughout the lifespan in these commonly used C57BL/6J and CBA/CaJ mouse models. Copyright © 2012 Wiley Periodicals, Inc.

Effects of aging and sensory loss on glial cells in mouse visual and auditory cortices

PubMed Central

Tremblay, Marie-Ève; Zettel, Martha L.; Ison, James R.; Allen, Paul D.; Majewska, Ania K.

2011-01-01

Normal aging is often accompanied by a progressive loss of receptor sensitivity in hearing and vision, whose consequences on cellular function in cortical sensory areas have remained largely unknown. By examining the primary auditory (A1) and visual (V1) cortices in two inbred strains of mice undergoing either age-related loss of audition (C57BL/6J) or vision (CBA/CaJ), we were able to describe cellular and subcellular changes that were associated with normal aging (occurring in A1 and V1 of both strains) or specifically with age-related sensory loss (only in A1 of C57BL/6J or V1 of CBA/CaJ), using immunocytochemical electron microscopy and light microscopy. While the changes were subtle in neurons, glial cells and especially microglia were transformed in aged animals. Microglia became more numerous and irregularly distributed, displayed more variable cell body and process morphologies, occupied smaller territories, and accumulated phagocytic inclusions that often displayed ultrastructural features of synaptic elements. Additionally, evidence of myelination defects were observed, and aged oligodendrocytes became more numerous and were more often encountered in contiguous pairs. Most of these effects were profoundly exacerbated by age-related sensory loss. Together, our results suggest that the age-related alteration of glial cells in sensory cortical areas can be accelerated by activity-driven central mechanisms that result from an age-related loss of peripheral sensitivity. In light of our observations, these age-related changes in sensory function should be considered when investigating cellular, cortical and behavioral functions throughout the lifespan in these commonly used C57BL/6J and CBA/CaJ mouse models. PMID:22223464

Role of microtubules in the contractile dysfunction of hypertrophied myocardium

NASA Technical Reports Server (NTRS)

Zile, M. R.; Koide, M.; Sato, H.; Ishiguro, Y.; Conrad, C. H.; Buckley, J. M.; Morgan, J. P.; Cooper, G. 4th

1999-01-01

OBJECTIVES: We sought to determine whether the ameliorative effects of microtubule depolymerization on cellular contractile dysfunction in pressure overload cardiac hypertrophy apply at the tissue level. BACKGROUND: A selective and persistent increase in microtubule density causes decreased contractile function of cardiocytes from cats with hypertrophy produced by chronic right ventricular (RV) pressure overloading. Microtubule depolymerization by colchicine normalizes contractility in these isolated cardiocytes. However, whether these changes in cellular function might contribute to changes in function at the more highly integrated and complex cardiac tissue level was unknown. METHODS: Accordingly, RV papillary muscles were isolated from 25 cats with RV pressure overload hypertrophy induced by pulmonary artery banding (PAB) for 4 weeks and 25 control cats. Contractile state was measured using physiologically sequenced contractions before and 90 min after treatment with 10(-5) mol/liter colchicine. RESULTS: The PAB significantly increased RV systolic pressure and the RV weight/body weight ratio in PAB; it significantly decreased developed tension from 59+/-3 mN/mm2 in control to 25+/-4 mN/mm2 in PAB, shortening extent from 0.21+/-0.01 muscle lengths (ML) in control to 0.12+/-0.01 ML in PAB, and shortening rate from 1.12+/-0.07 ML/s in control to 0.55+/-0.03 ML/s in PAB. Indirect immunofluorescence confocal microscopy showed that PAB muscles had a selective increase in microtubule density and that colchicine caused complete microtubule depolymerization in both control and PAB papillary muscles. Microtubule depolymerization normalized myocardial contractility in papillary muscles of PAB cats but did not alter contractility in control muscles. CONCLUSIONS: Excess microtubule density, therefore, is equally important to both cellular and to myocardial contractile dysfunction caused by chronic, severe pressure-overload cardiac hypertrophy.

Cellular metabolism and disease: what do metabolic outliers teach us?

PubMed Central

DeBerardinis, Ralph J.; Thompson, Craig B.

2012-01-01

An understanding of metabolic pathways based solely on biochemistry textbooks would underestimate the pervasive role of metabolism in essentially every aspect of biology. It is evident from recent work that many human diseases involve abnormal metabolic states – often genetically programmed – that perturb normal physiology and lead to severe tissue dysfunction. Understanding these metabolic outliers is now a crucial frontier in disease-oriented research. This review discusses the broad impact of metabolism in cellular function, how modern concepts of metabolism can inform our understanding of common diseases like cancer, and considers the prospects of developing new metabolic approaches to disease treatment. PMID:22424225

Managing Hyperkalemia: Stepping Into a New Frontier.

PubMed

Pham, Antony Q; Sexton, Jessica; Wimer, Dexter; Rana, Isha; Nguyen, Timothy

2017-10-01

Maintaining potassium balance in the body is essential for cellular function. Even a slight increase in normal serum potassium levels (3.5-5.0 mEq/L) can interfere with metabolism, electrical action potentials, and cellular processes. Hyperkalemia is commonly seen in patients with chronic kidney disease (CKD) and in patients on renin-angiotensin-aldosterone system (RAAS) inhibitors. Sodium polystyrene sulfonate (SPS), diuretics, and hemodialysis are currently available methods for removing potassium from the body; however, these options have their limitations. Patiromer (Veltassa) and sodium zirconium cyclosilicate are 2 new therapeutic options that can potentially lead a new frontier in the management of hyperkalemia. This article will review these novel treatments.

A specific, nonproliferative role for E2F-5 in choroid plexus function revealed by gene targeting

PubMed Central

Lindeman, Geoffrey J.; Dagnino, Lina; Gaubatz, Stefan; Xu, Yuhui; Bronson, Roderick T.; Warren, Henry B.; Livingston, David M.

1998-01-01

Homozygous E2F-5 knockout embryos and mice have been generated. Although embryonic development appeared normal, newborn mice developed nonobstructive hydrocephalus, suggesting excessive cerebrospinal fluid (CSF) production. Although the CSF-producing choroid plexus displayed normal cellular organization, it contained abundant electron-lucent epithelial cells, consistent with excessive CSF secretory activity. Moreover, E2F-5 CNS expression in normal animals was largely confined to the choroid plexus. Cell cycle kinetics were not perturbed in homozygous knockout embryo fibroblasts. Thus, E2F-5 is not essential for cell proliferation. Rather, it affects the secretory behavior of a differentiated neural tissue. PMID:9553039

Cystic Fibrosis, Cystic Fibrosis Transmembrane Conductance Regulator and Drugs: Insights from Cellular Trafficking.

PubMed

Bridges, Robert J; Bradbury, Neil A

2018-01-01

The eukaryotic cell is organized into membrane-delineated compartments that are characterized by specific cadres of proteins sustaining biochemically distinct cellular processes. The appropriate subcellular localization of proteins is key to proper organelle function and provides a physiological context for cellular processes. Disruption of normal trafficking pathways for proteins is seen in several genetic diseases, where a protein's absence for a specific subcellular compartment leads to organelle disruption, and in the context of an individual, a disruption of normal physiology. Importantly, several drug therapies can also alter protein trafficking, causing unwanted side effects. Thus, a deeper understanding of trafficking pathways needs to be appreciated as novel therapeutic modalities are proposed. Despite the promising efficacy of novel therapeutic agents, the intracellular bioavailability of these compounds has proved to be a potential barrier, leading to failures in treatments for various diseases and disorders. While endocytosis of drug moieties provides an efficient means of getting material into cells, the subsequent release and endosomal escape of materials into the cytosol where they need to act has been a barrier. An understanding of cellular protein/lipid trafficking pathways has opened up strategies for increasing drug bioavailability. Approaches to enhance endosomal exit have greatly increased the cytosolic bioavailability of drugs and will provide a means of investigating previous drugs that may have been shelved due to their low cytosolic concentration.

Cellular Automata Generalized To An Inferential System

NASA Astrophysics Data System (ADS)

Blower, David J.

2007-11-01

Stephen Wolfram popularized elementary one-dimensional cellular automata in his book, A New Kind of Science. Among many remarkable things, he proved that one of these cellular automata was a Universal Turing Machine. Such cellular automata can be interpreted in a different way by viewing them within the context of the formal manipulation rules from probability theory. Bayes's Theorem is the most famous of such formal rules. As a prelude, we recapitulate Jaynes's presentation of how probability theory generalizes classical logic using modus ponens as the canonical example. We emphasize the important conceptual standing of Boolean Algebra for the formal rules of probability manipulation and give an alternative demonstration augmenting and complementing Jaynes's derivation. We show the complementary roles played in arguments of this kind by Bayes's Theorem and joint probability tables. A good explanation for all of this is afforded by the expansion of any particular logic function via the disjunctive normal form (DNF). The DNF expansion is a useful heuristic emphasized in this exposition because such expansions point out where relevant 0s should be placed in the joint probability tables for logic functions involving any number of variables. It then becomes a straightforward exercise to rely on Boolean Algebra, Bayes's Theorem, and joint probability tables in extrapolating to Wolfram's cellular automata. Cellular automata are seen as purely deductive systems, just like classical logic, which probability theory is then able to generalize. Thus, any uncertainties which we might like to introduce into the discussion about cellular automata are handled with ease via the familiar inferential path. Most importantly, the difficult problem of predicting what cellular automata will do in the far future is treated like any inferential prediction problem.

A new twist on an old idea part 2: cyclosporine preserves normal mitochondrial but not cardiomyocyte function in mini‐swine with compensated heart failure

PubMed Central

Hiemstra, Jessica A.; Gutiérrez‐Aguilar, Manuel; Marshall, Kurt D.; McCommis, Kyle S.; Zgoda, Pamela J.; Cruz‐Rivera, Noelany; Jenkins, Nathan T.; Krenz, Maike; Domeier, Timothy L.; Baines, Christopher P.; Emter, Craig A.

2014-01-01

Abstract We recently developed a clinically relevant mini‐swine model of heart failure with preserved ejection fraction (HFpEF), in which diastolic dysfunction was associated with increased mitochondrial permeability transition (MPT). Early diastolic function is ATP and Ca2+‐dependent, thus, we hypothesized chronic low doses of cyclosporine (CsA) would preserve mitochondrial function via inhibition of MPT and subsequently maintain normal cardiomyocyte Ca2+ handling and contractile characteristics. Left ventricular cardiomyocytes were isolated from aortic‐banded Yucatan mini‐swine divided into three groups; control nonbanded (CON), HFpEF nontreated (HF), and HFpEF treated with CsA (HF‐CsA). CsA mitigated the deterioration of mitochondrial function observed in HF animals, including functional uncoupling of Complex I‐dependent mitochondrial respiration and increased susceptibility to MPT. Attenuation of mitochondrial dysfunction in the HF‐CsA group was not associated with commensurate improvement in cardiomyocyte Ca2+ handling or contractility. Ca2+ transient amplitude was reduced and transient time to peak and recovery (tau) prolonged in HF and HF‐CsA groups compared to CON. Alterations in Ca2+ transient parameters observed in the HF and HF‐CsA groups were associated with decreased cardiomyocyte shortening and shortening rate. Cellular function was consistent with impaired in vivo systolic and diastolic whole heart function. A significant systemic hypertensive response to CsA was observed in HF‐CsA animals, and may have played a role in the accelerated the development of heart failure at both the whole heart and cellular levels. Given the significant detriment to cardiac function observed in response to CsA, our findings suggest chronic CsA treatment is not a viable therapeutic option for HFpEF. PMID:24963034

Atomic force microscopy studies on cellular elastic and viscoelastic properties.

PubMed

Li, Mi; Liu, Lianqing; Xi, Ning; Wang, Yuechao

2018-01-01

In this work, a method based on atomic force microscopy (AFM) approach-reside-retract experiments was established to simultaneously quantify the elastic and viscoelastic properties of single cells. First, the elastic and viscoelastic properties of normal breast cells and cancerous breast cells were measured, showing significant differences in Young's modulus and relaxation times between normal and cancerous breast cells. Remarkable differences in cellular topography between normal and cancerous breast cells were also revealed by AFM imaging. Next, the elastic and viscoelasitc properties of three other types of cell lines and primary normal B lymphocytes were measured; results demonstrated the potential of cellular viscoelastic properties in complementing cellular Young's modulus for discerning different states of cells. This research provides a novel way to quantify the mechanical properties of cells by AFM, which allows investigation of the biomechanical behaviors of single cells from multiple aspects.

The ins and outs of intracellular ion homeostasis: NHX-type cation/H(+) transporters.

PubMed

Bassil, Elias; Blumwald, Eduardo

2014-12-01

The biochemical characterization of cation/H(+) exchange has been known since 1985 [1], yet only recently have we begun to understand the contribution of individual exchangers to ion homeostasis in plants. One particularly important class of exchangers is the NHX-type that is associated with Na(+) transport and therefore salinity tolerance. New evidence suggests that under normal growth conditions NHXs are critical regulators of K(+) and pH homeostasis and have important roles, depending on their cellular localization, in the generation of turgor as well as in vesicular trafficking. Recent advances highlight novel and exciting functions of intracellular NHXs in growth and development, stress adaptation and osmotic adjustment. Here, we elaborate on new and emerging cellular and physiological functions of this group of H(+)-coupled cation exchangers. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cell Biology and Pathophysiology of α-Synuclein

PubMed Central

Burré, Jacqueline; Sharma, Manu; Südhof, Thomas C.

2017-01-01

α-Synuclein is an abundant neuronal protein that is highly enriched in presynaptic nerve terminals. Genetics and neuropathology studies link α-synuclein to Parkinson’s disease (PD) and other neurodegenerative disorders. Accumulation of misfolded oligomers and larger aggregates of α-synuclein defines multiple neurodegenerative diseases called synucleino-pathies, but the mechanisms by which α-synuclein acts in neurodegeneration are unknown. Moreover, the normal cellular function of α-synuclein remains debated. In this perspective, we review the structural characteristics of α-synuclein, its developmental expression pattern, its cellular and subcellular localization, and its function in neurons. We also discuss recent progress on secretion of α-synuclein, which may contribute to its interneuronal spread in a prion-like fashion, and describe the neurotoxic effects of α-synuclein that are thought to be responsible for its role in neurodegeneration. PMID:28108534

Relative Expression Levels Rather Than Specific Activity Plays the Major Role in Determining In Vivo AKT Isoform Substrate Specificity

PubMed Central

Lee, Rachel S.; House, Colin M.; Cristiano, Briony E.; Hannan, Ross D.; Pearson, Richard B.; Hannan, Katherine M.

2011-01-01

The AKT protooncogene mediates many cellular processes involved in normal development and disease states such as cancer. The three structurally similar isoforms: AKT1, AKT2, and AKT3 exhibit both functional redundancy and isoform-specific functions; however the basis for their differential signalling remains unclear. Here we show that in vitro, purified AKT3 is ∼47-fold more active than AKT1 at phosphorylating peptide and protein substrates. Despite these marked variations in specific activity between the individual isoforms, a comprehensive analysis of phosphorylation of validated AKT substrates indicated only subtle differences in signalling via individual isoforms in vivo. Therefore, we hypothesise, at least in this model system, that relative tissue/cellular abundance, rather than specific activity, plays the dominant role in determining AKT substrate specificity in situ. PMID:21869924

Immunological Development and Cardiovascular Function Are Normal in Annexin VI Null Mutant Mice

PubMed Central

Hawkins, Tim E.; Roes, Jürgen; Rees, Daryl; Monkhouse, Jayne; Moss, Stephen E.

1999-01-01

Annexins are calcium-binding proteins of unknown function but which are implicated in important cellular processes, including anticoagulation, ion flux regulation, calcium homeostasis, and endocytosis. To gain insight into the function of annexin VI, we performed targeted disruption of its gene in mice. Matings between heterozygous mice produced offspring with a normal Mendelian pattern of inheritance, indicating that the loss of annexin VI did not interfere with viability in utero. Mice lacking annexin VI reached sexual maturity at the same age as their normal littermates, and both males and females were fertile. Because of interest in the role of annexin VI in cardiovascular function, we examined heart rate and blood pressure in knockout and wild-type mice and found these to be identical in the two groups. Similarly, the cardiovascular responses of both sets of mice to septic shock were indistinguishable. We also examined components of the immune system and found no differences in thymic, splenic, or bone marrow lymphocyte levels between knockout and wild-type mice. This is the first study of annexin knockout mice, and the lack of a clear phenotype has broad implications for current views of annexin function. PMID:10567528

Deep sequencing of the small RNA transcriptome of normal and malignant human B cells identifies hundreds of novel microRNAs

PubMed Central

Jima, Dereje D.; Zhang, Jenny; Jacobs, Cassandra; Richards, Kristy L.; Dunphy, Cherie H.; Choi, William W. L.; Yan Au, Wing; Srivastava, Gopesh; Czader, Magdalena B.; Rizzieri, David A.; Lagoo, Anand S.; Lugar, Patricia L.; Mann, Karen P.; Flowers, Christopher R.; Bernal-Mizrachi, Leon; Naresh, Kikkeri N.; Evens, Andrew M.; Gordon, Leo I.; Luftig, Micah; Friedman, Daphne R.; Weinberg, J. Brice; Thompson, Michael A.; Gill, Javed I.; Liu, Qingquan; How, Tam; Grubor, Vladimir; Gao, Yuan; Patel, Amee; Wu, Han; Zhu, Jun; Blobe, Gerard C.; Lipsky, Peter E.; Chadburn, Amy

2010-01-01

A role for microRNA (miRNA) has been recognized in nearly every biologic system examined thus far. A complete delineation of their role must be preceded by the identification of all miRNAs present in any system. We elucidated the complete small RNA transcriptome of normal and malignant B cells through deep sequencing of 31 normal and malignant human B-cell samples that comprise the spectrum of B-cell differentiation and common malignant phenotypes. We identified the expression of 333 known miRNAs, which is more than twice the number previously recognized in any tissue type. We further identified the expression of 286 candidate novel miRNAs in normal and malignant B cells. These miRNAs were validated at a high rate (92%) using quantitative polymerase chain reaction, and we demonstrated their application in the distinction of clinically relevant subgroups of lymphoma. We further demonstrated that a novel miRNA cluster, previously annotated as a hypothetical gene LOC100130622, contains 6 novel miRNAs that regulate the transforming growth factor-β pathway. Thus, our work suggests that more than a third of the miRNAs present in most cellular types are currently unknown and that these miRNAs may regulate important cellular functions. PMID:20733160

Harnessing the Power of Metabolism for Seizure Prevention: Focus on Dietary Treatments

PubMed Central

Hartman, Adam L.; Stafstrom, Carl E.

2012-01-01

The continued occurrence of refractory seizures in at least one-third of children and adults with epilepsy, despite the availability of almost 15 conventional and novel anticonvulsant drugs, speaks to a dire need to develop novel therapeutic approaches. Cellular metabolism, the critical pathways by which cells access and utilize energy, is critical for normal neuronal function. Furthermore, mounting evidence suggests direct links between energy metabolism and cellular excitability. The high-fat, low-carbohydrate ketogenic diet has been used as a treatment for drug-refractory epilepsy for almost a century. Yet, the multitude of alternative therapies to target aspects of cellular metabolism and hyperexcitability is almost untapped. Approaches discussed in this review offer a wide diversity of therapeutic targets that might be exploited by investigators in the search for safer and more effective epilepsy treatments. PMID:23110824

NADPH oxidases of the brain: distribution, regulation, and function.

PubMed

Infanger, David W; Sharma, Ram V; Davisson, Robin L

2006-01-01

The NADPH oxidase is a multi-subunit enzyme that catalyzes the reduction of molecular oxygen to form superoxide (O(2)(-)). While classically linked to the respiratory burst in neutrophils, recent evidence now shows that O(2)(-) (and associated reactive oxygen species, ROS) generated by NADPH oxidase in nonphagocytic cells serves myriad functions in health and disease. An entire new family of NADPH Oxidase (Nox) homologues has emerged, which vary widely in cell and tissue distribution, as well as in function and regulation. A major concept in redox signaling is that while NADPH oxidase-derived ROS are necessary for normal cellular function, excessive oxidative stress can contribute to pathological disease. This certainly is true in the central nervous system (CNS), where normal NADPH oxidase function appears to be required for processes such as neuronal signaling, memory, and central cardiovascular homeostasis, but overproduction of ROS contributes to neurotoxicity, neurodegeneration, and cardiovascular diseases. Despite implications of NADPH oxidase in normal and pathological CNS processes, still relatively little is known about the mechanisms involved. This paper summarizes the evidence for NADPH oxidase distribution, regulation, and function in the CNS, emphasizing the diversity of Nox isoforms and their new and emerging role in neuro-cardiovascular function. In addition, perspectives for future research and novel therapeutic targets are offered.

Yeast prions are useful for studying protein chaperones and protein quality control.

PubMed

Masison, Daniel C; Reidy, Michael

2015-01-01

Protein chaperones help proteins adopt and maintain native conformations and play vital roles in cellular processes where proteins are partially folded. They comprise a major part of the cellular protein quality control system that protects the integrity of the proteome. Many disorders are caused when proteins misfold despite this protection. Yeast prions are fibrous amyloid aggregates of misfolded proteins. The normal action of chaperones on yeast prions breaks the fibers into pieces, which results in prion replication. Because this process is necessary for propagation of yeast prions, even small differences in activity of many chaperones noticeably affect prion phenotypes. Several other factors involved in protein processing also influence formation, propagation or elimination of prions in yeast. Thus, in much the same way that the dependency of viruses on cellular functions has allowed us to learn much about cell biology, the dependency of yeast prions on chaperones presents a unique and sensitive way to monitor the functions and interactions of many components of the cell's protein quality control system. Our recent work illustrates the utility of this system for identifying and defining chaperone machinery interactions.

Somatostatin-Positive Gamma-Aminobutyric Acid Interneuron Deficits in Depression: Cortical Microcircuit and Therapeutic Perspectives.

PubMed

Fee, Corey; Banasr, Mounira; Sibille, Etienne

2017-10-15

The functional integration of external and internal signals forms the basis of information processing and is essential for higher cognitive functions. This occurs in finely tuned cortical microcircuits whose functions are balanced at the cellular level by excitatory glutamatergic pyramidal neurons and inhibitory gamma-aminobutyric acidergic (GABAergic) interneurons. The balance of excitation and inhibition, from cellular processes to neural network activity, is characteristically disrupted in multiple neuropsychiatric disorders, including major depressive disorder (MDD), bipolar disorder, anxiety disorders, and schizophrenia. Specifically, nearly 3 decades of research demonstrate a role for reduced inhibitory GABA level and function across disorders. In MDD, recent evidence from human postmortem and animal studies suggests a selective vulnerability of GABAergic interneurons that coexpress the neuropeptide somatostatin (SST). Advances in cell type-specific molecular genetics have now helped to elucidate several important roles for SST interneurons in cortical processing (regulation of pyramidal cell excitatory input) and behavioral control (mood and cognition). Here, we review evidence for altered inhibitory function arising from GABAergic deficits across disorders and specifically in MDD. We then focus on properties of the cortical microcircuit, where SST-positive GABAergic interneuron deficits may disrupt functioning in several ways. Finally, we discuss the putative origins of SST cell deficits, as informed by recent research, and implications for therapeutic approaches. We conclude that deficits in SST interneurons represent a contributing cellular pathology and therefore a promising target for normalizing altered inhibitory function in MDD and other disorders with reduced SST cell and GABA functions. Copyright © 2017 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

The relationship between the nucleolus and cancer: Current evidence and emerging paradigms.

PubMed

Orsolic, Ines; Jurada, Deana; Pullen, Nick; Oren, Moshe; Eliopoulos, Aristides G; Volarevic, Sinisa

2016-06-01

The nucleolus is the most prominent nuclear substructure assigned to produce ribosomes; molecular machines that are responsible for carrying out protein synthesis. To meet the increased demand for proteins during cell growth and proliferation the cell must increase protein synthetic capacity by upregulating ribosome biogenesis. While larger nucleolar size and number have been recognized as hallmark features of many tumor types, recent evidence has suggested that, in addition to overproduction of ribosomes, decreased ribosome biogenesis as well as qualitative changes in this process could also contribute to tumor initiation and cancer progression. Furthermore, the nucleolus has become the focus of intense attention for its involvement in processes that are clearly unrelated to ribosome biogenesis such as sensing and responding to endogenous and exogenous stressors, maintenance of genome stability, regulation of cell-cycle progression, cellular senescence, telomere function, chromatin structure, establishment of nuclear architecture, global regulation of gene expression and biogenesis of multiple ribonucleoprotein particles. The fact that dysregulation of many of these fundamental cellular processes may contribute to the malignant phenotype suggests that normal functioning of the nucleolus safeguards against the development of cancer and indicates its potential as a therapeutic approach. Here we review the recent advances made toward understanding these newly-recognized nucleolar functions and their roles in normal and cancer cells, and discuss possible future research directions. Copyright © 2015 Elsevier Ltd. All rights reserved.

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

PubMed Central

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

2007-01-01

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

Sub-cellular force microscopy in single normal and cancer cells

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

Babahosseini, H.; Carmichael, B.; Strobl, J.S.

2015-08-07

This work investigates the biomechanical properties of sub-cellular structures of breast cells using atomic force microscopy (AFM). The cells are modeled as a triple-layered structure where the Generalized Maxwell model is applied to experimental data from AFM stress-relaxation tests to extract the elastic modulus, the apparent viscosity, and the relaxation time of sub-cellular structures. The triple-layered modeling results allow for determination and comparison of the biomechanical properties of the three major sub-cellular structures between normal and cancerous cells: the up plasma membrane/actin cortex, the mid cytoplasm/nucleus, and the low nuclear/integrin sub-domains. The results reveal that the sub-domains become stiffer andmore » significantly more viscous with depth, regardless of cell type. In addition, there is a decreasing trend in the average elastic modulus and apparent viscosity of the all corresponding sub-cellular structures from normal to cancerous cells, which becomes most remarkable in the deeper sub-domain. The presented modeling in this work constitutes a unique AFM-based experimental framework to study the biomechanics of sub-cellular structures. - Highlights: • The cells are modeled as a triple-layered structure using Generalized Maxwell model. • The sub-domains include membrane/cortex, cytoplasm/nucleus, and nuclear/integrin. • Biomechanics of corresponding sub-domains are compared among normal and cancer cells. • Viscoelasticity of sub-domains show a decreasing trend from normal to cancer cells. • The decreasing trend becomes most significant in the deeper sub-domain.« less

The role of MDM2 and MDM4 in breast cancer development and prevention.

PubMed

Haupt, Sue; Vijayakumaran, Reshma; Miranda, Panimaya Jeffreena; Burgess, Andrew; Lim, Elgene; Haupt, Ygal

2017-02-01

The major cause of death from breast cancer is not the primary tumour, but relapsing, drug-resistant, metastatic disease. Identifying factors that contribute to aggressive cancer offers important leads for therapy. Inherent defence against carcinogens depends on the individual molecular make-up of each person. Important molecular determinants of these responses are under the control of the mouse double minute (MDM) family: comprised of the proteins MDM2 and MDM4. In normal, healthy adult cells, the MDM family functions to critically regulate measured, cellular responses to stress and subsequent recovery. Proper function of the MDM family is vital for normal breast development, but also for preserving genomic fidelity. The MDM family members are best characterized for their negative regulation of the major tumour suppressor p53 to modulate stress responses. Their impact on other cellular regulators is emerging. Inappropriately elevated protein levels of the MDM family are highly associated with an increased risk of cancer incidence. Exploration of the MDM family members as cancer therapeutic targets is relevant for designing tailored anti-cancer treatments, but successful approaches must strategically consider the impact on both the target cancer and adjacent healthy cells and tissues. This review focuses on recent findings pertaining to the role of the MDM family in normal and malignant breast cells. © The Author (2017). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS.

Synthetic biology meets tissue engineering

PubMed Central

Davies, Jamie A.; Cachat, Elise

2016-01-01

Classical tissue engineering is aimed mainly at producing anatomically and physiologically realistic replacements for normal human tissues. It is done either by encouraging cellular colonization of manufactured matrices or cellular recolonization of decellularized natural extracellular matrices from donor organs, or by allowing cells to self-organize into organs as they do during fetal life. For repair of normal bodies, this will be adequate but there are reasons for making unusual, non-evolved tissues (repair of unusual bodies, interface to electromechanical prostheses, incorporating living cells into life-support machines). Synthetic biology is aimed mainly at engineering cells so that they can perform custom functions: applying synthetic biological approaches to tissue engineering may be one way of engineering custom structures. In this article, we outline the ‘embryological cycle’ of patterning, differentiation and morphogenesis and review progress that has been made in constructing synthetic biological systems to reproduce these processes in new ways. The state-of-the-art remains a long way from making truly synthetic tissues, but there are now at least foundations for future work. PMID:27284030

Mammary Gland Involution Provides a Unique Model to Study the TGF-β Cancer Paradox

PubMed Central

Guo, Qiuchen; Betts, Courtney; Pennock, Nathan; Mitchell, Elizabeth; Schedin, Pepper

2017-01-01

Transforming Growth Factor-β (TGF-β) signaling in cancer has been termed the “TGF-β paradox”, acting as both a tumor suppresser and promoter. The complexity of TGF-β signaling within the tumor is context dependent, and greatly impacted by cellular crosstalk between TGF-β responsive cells in the microenvironment including adjacent epithelial, endothelial, mesenchymal, and hematopoietic cells. Here we utilize normal, weaning-induced mammary gland involution as a tissue microenvironment model to study the complexity of TGF-β function. This article reviews facets of mammary gland involution that are TGF-β regulated, namely mammary epithelial cell death, immune activation, and extracellular matrix remodeling. We outline how distinct cellular responses and crosstalk between cell types during physiologically normal mammary gland involution contribute to simultaneous tumor suppressive and promotional microenvironments. We also highlight alternatives to direct TGF-β blocking anti-cancer therapies with an emphasis on eliciting concerted microenvironmental-mediated tumor suppression. PMID:28098775

PRUNE is crucial for normal brain development and mutated in microcephaly with neurodevelopmental impairment.

PubMed

Zollo, Massimo; Ahmed, Mustafa; Ferrucci, Veronica; Salpietro, Vincenzo; Asadzadeh, Fatemeh; Carotenuto, Marianeve; Maroofian, Reza; Al-Amri, Ahmed; Singh, Royana; Scognamiglio, Iolanda; Mojarrad, Majid; Musella, Luca; Duilio, Angela; Di Somma, Angela; Karaca, Ender; Rajab, Anna; Al-Khayat, Aisha; Mohan Mohapatra, Tribhuvan; Eslahi, Atieh; Ashrafzadeh, Farah; Rawlins, Lettie E; Prasad, Rajniti; Gupta, Rashmi; Kumari, Preeti; Srivastava, Mona; Cozzolino, Flora; Kumar Rai, Sunil; Monti, Maria; Harlalka, Gaurav V; Simpson, Michael A; Rich, Philip; Al-Salmi, Fatema; Patton, Michael A; Chioza, Barry A; Efthymiou, Stephanie; Granata, Francesca; Di Rosa, Gabriella; Wiethoff, Sarah; Borgione, Eugenia; Scuderi, Carmela; Mankad, Kshitij; Hanna, Michael G; Pucci, Piero; Houlden, Henry; Lupski, James R; Crosby, Andrew H; Baple, Emma L

2017-04-01

PRUNE is a member of the DHH (Asp-His-His) phosphoesterase protein superfamily of molecules important for cell motility, and implicated in cancer progression. Here we investigated multiple families from Oman, India, Iran and Italy with individuals affected by a new autosomal recessive neurodevelopmental and degenerative disorder in which the cardinal features include primary microcephaly and profound global developmental delay. Our genetic studies identified biallelic mutations of PRUNE1 as responsible. Our functional assays of disease-associated variant alleles revealed impaired microtubule polymerization, as well as cell migration and proliferation properties, of mutant PRUNE. Additionally, our studies also highlight a potential new role for PRUNE during microtubule polymerization, which is essential for the cytoskeletal rearrangements that occur during cellular division and proliferation. Together these studies define PRUNE as a molecule fundamental for normal human cortical development and define cellular and clinical consequences associated with PRUNE mutation. © The Author (2017). Published by Oxford University Press on behalf of the Guarantors of Brain.

Imaging of normal and pathologic joint synovium using nonlinear optical microscopy as a potential diagnostic tool

NASA Astrophysics Data System (ADS)

Tiwari, Nivedan; Chabra, Sanjay; Mehdi, Sheherbano; Sweet, Paula; Krasieva, Tatiana B.; Pool, Roy; Andrews, Brian; Peavy, George M.

2010-09-01

An estimated 1.3 million people in the United States suffer from rheumatoid arthritis (RA). RA causes profound changes in the synovial membrane of joints, and without early diagnosis and intervention, progresses to permanent alterations in joint structure and function. The purpose of this study is to determine if nonlinear optical microscopy (NLOM) can utilize the natural intrinsic fluorescence properties of tissue to generate images that would allow visualization of the structural and cellular composition of fresh, unfixed normal and pathologic synovial tissue. NLOM is performed on rabbit knee joint synovial samples using 730- and 800-nm excitation wavelengths. Less than 30 mW of excitation power delivered with a 40×, 0.8-NA water immersion objective is sufficient for the visualization of synovial structures to a maximum depth of 70 μm without tissue damage. NLOM imaging of normal and pathologic synovial tissue reveals the cellular structure, synoviocytes, adipocytes, collagen, vascular structures, and differential characteristics of inflammatory infiltrates without requiring tissue processing or staining. Further study to evaluate the ability of NLOM to assess the characteristics of pathologic synovial tissue and its potential role for the management of disease is warranted.

[Effect of polysaccharides in processed Sibiraea on immunologic function of immunosuppression mice].

PubMed

Duan, Bowen; Li, Yun; Liu, Xin; Yang, Yongjian

2010-06-01

To study the effect of polysaccharides in processed Sibiraea on the immunologic function of immunosuppression mice. The immunosuppressed mice were induced by cyclophosphamide. After the treatment, the organ weight index and the delayed type hypersensitivity of the mice were investigated. The humoral immune function was determined by serum hemolysin assay. Non-specific immune function was determined by carbon clearance method. Cellular immune function was determined by spleen lymphocyte proliferation test. Two hundred kunming mice were randomly divided into five groups: normal controls, model group, low-dose group (110 mg x kg(-1)), middle-dose group (220 mg x kg(-1)), high-dose group (440 mg x kg(-1)). Drugs were given to the mice by oral gavage every day. The immunosuppressed mice treated with Sibiraea polysibcharide at intragastrica dose of 110-440 mg x kg(-1) have increased weight of the immune organs, increased content of DTH and content in serum hemolysin lgG and lgM. Mean while the rate of carbon clearance was enhanced and the proliferation of spleen lymphocyte was increased. Polysaccharides in processed Sibiraea can increase the weight of the immune organs. At the same time, non-specific immune, DTH, humoral immune and cellular immune function were enhanced significantly.

Prion Protein Regulates Iron Transport by Functioning as a Ferrireductase

PubMed Central

Singh, Ajay; Haldar, Swati; Horback, Katharine; Tom, Cynthia; Zhou, Lan; Meyerson, Howard; Singh, Neena

2017-01-01

Prion protein (PrPC) is implicated in the pathogenesis of prion disorders, but its normal function is unclear. We demonstrate that PrPC is a ferrireductase (FR), and its absence causes systemic iron deficiency in PrP knock-out mice (PrP−/−). When exposed to non-transferrin-bound (NTB) radioactive-iron (59FeCl3) by gastric-gavage, PrP−/− mice absorb significantly more 59Fe from the intestinal lumen relative to controls, indicating appropriate systemic response to the iron deficiency. Chronic exposure to excess dietary iron corrects this deficiency, but unlike wild-type (PrP+/+) controls that remain iron over-loaded, PrP−/− mice revert back to the iron deficient phenotype after 5 months of chase on normal diet. Bone marrow (BM) preparations of PrP−/− mice on normal diet show relatively less stainable iron, and this phenotype is only partially corrected by intraperitoneal administration of excess iron-dextran. Cultured PrP−/− BM-macrophages incorporate significantly less NTB-59Fe in the absence or presence of excess extracellular iron, indicating reduced uptake and/or storage of available iron in the absence of PrPC. When expressed in neuroblastoma cells, PrPC exhibits NAD(P)H-dependent cell-surface and intracellular FR activity that requires the copper-binding octa-peptide-repeat region and linkage to the plasma membrane for optimal function. Incorporation of NTB-59Fe by neuroblastoma cells correlates with FR activity of PrPC, implicating PrPC in cellular iron uptake and metabolism. These observations explain the correlation between PrPC expression and cellular iron levels, and the cause of iron imbalance in sporadic-Creutzfeldt-Jakob-disease brains where PrPC accumulates as insoluble aggregates. PMID:23478311

Galactose-depleted xyloglucan is dysfunctional and leads to dwarfism in Arabidopsis.

PubMed

Kong, Yingzhen; Peña, Maria J; Renna, Luciana; Avci, Utku; Pattathil, Sivakumar; Tuomivaara, Sami T; Li, Xuemei; Reiter, Wolf-Dieter; Brandizzi, Federica; Hahn, Michael G; Darvill, Alan G; York, William S; O'Neill, Malcolm A

2015-04-01

Xyloglucan is a polysaccharide that has important roles in the formation and function of the walls that surround growing land plant cells. Many of these plants synthesize xyloglucan that contains galactose in two different side chains (L and F), which exist in distinct molecular environments. However, little is known about the contribution of these side chains to xyloglucan function. Here, we show that Arabidopsis (Arabidopsis thaliana) mutants devoid of the F side chain galactosyltransferase MURUS3 (MUR3) form xyloglucan that lacks F side chains and contains much less galactosylated xylose than its wild-type counterpart. The galactose-depleted xyloglucan is dysfunctional, as it leads to mutants that are dwarfed with curled rosette leaves, short petioles, and short inflorescence stems. Moreover, cell wall matrix polysaccharides, including xyloglucan and pectin, are not properly secreted and instead accumulate within intracellular aggregates. Near-normal growth is restored by generating mur3 mutants that produce no detectable amounts of xyloglucan. Thus, cellular processes are affected more by the presence of the dysfunctional xyloglucan than by eliminating xyloglucan altogether. To identify structural features responsible for xyloglucan dysfunction, xyloglucan structure was modified in situ by generating mur3 mutants that lack specific xyloglucan xylosyltransferases (XXTs) or that overexpress the XYLOGLUCAN L-SIDE CHAIN GALACTOSYLTRANSFERASE2 (XLT2) gene. Normal growth was restored in the mur3-3 mutant overexpressing XLT2 and in mur3-3 xxt double mutants when the dysfunctional xyloglucan was modified by doubling the amounts of galactosylated side chains. Our study assigns a role for galactosylation in normal xyloglucan function and demonstrates that altering xyloglucan side chain structure disturbs diverse cellular and physiological processes. © 2015 American Society of Plant Biologists. All Rights Reserved.

Controlled expression of nif and isc iron-sulfur protein maturation components reveals target specificity and limited functional replacement between the two systems.

PubMed

Dos Santos, Patricia C; Johnson, Deborah C; Ragle, Brook E; Unciuleac, Mihaela-Carmen; Dean, Dennis R

2007-04-01

The nitrogen-fixing organism Azotobacter vinelandii contains at least two systems that catalyze formation of [Fe-S] clusters. One of these systems is encoded by nif genes, whose products supply [Fe-S] clusters required for maturation of nitrogenase. The other system is encoded by isc genes, whose products are required for maturation of [Fe-S] proteins that participate in general metabolic processes. The two systems are similar in that they include an enzyme for the mobilization of sulfur (NifS or IscS) and an assembly scaffold (NifU or IscU) upon which [Fe-S] clusters are formed. Normal cellular levels of the Nif system, which supplies [Fe-S] clusters for the maturation of nitrogenase, cannot also supply [Fe-S] clusters for the maturation of other cellular [Fe-S] proteins. Conversely, when produced at the normal physiological levels, the Isc system cannot supply [Fe-S] clusters for the maturation of nitrogenase. In the present work we found that such target specificity for IscU can be overcome by elevated production of NifU. We also found that NifU, when expressed at normal levels, is able to partially replace the function of IscU if cells are cultured under low-oxygen-availability conditions. In contrast to the situation with IscU, we could not establish conditions in which the function of IscS could be replaced by NifS. We also found that elevated expression of the Isc components, as a result of deletion of the regulatory iscR gene, improved the capacity for nitrogen-fixing growth of strains deficient in either NifU or NifS.

Functional Dynamics of PDZ Binding Domains: A Normal-Mode Analysis

PubMed Central

De Los Rios, Paolo; Cecconi, Fabio; Pretre, Anna; Dietler, Giovanni; Michielin, Olivier; Piazza, Francesco; Juanico, Brice

2005-01-01

Postsynaptic density-95/disks large/zonula occludens-1 (PDZ) domains are relatively small (80–120 residues) protein binding modules central in the organization of receptor clusters and in the association of cellular proteins. Their main function is to bind C-terminals of selected proteins that are recognized through specific amino acids in their carboxyl end. Binding is associated with a deformation of the PDZ native structure and is responsible for dynamical changes in regions not in direct contact with the target. We investigate how this deformation is related to the harmonic dynamics of the PDZ structure and show that one low-frequency collective normal mode, characterized by the concerted movements of different secondary structures, is involved in the binding process. Our results suggest that even minimal structural changes are responsible for communication between distant regions of the protein, in agreement with recent NMR experiments. Thus, PDZ domains are a very clear example of how collective normal modes are able to characterize the relation between function and dynamics of proteins, and to provide indications on the precursors of binding/unbinding events. PMID:15821164

Monocyte function in infectious mononucleosis: evidence for a reversible cellular defect.

PubMed

Britton, S

1976-10-01

Migration of blood monocytes from patients with acute infectious mononucleosis and from normal controls was measured against chemotactic factors in serum. Moncytes from patients with acute infectious mononucleosis showed decreased migration as compared with that of control monocytes. However, serum from patients with infectious mononucleosis contained normal or above normal amounts of chemotaxins for monocytes. The migratory defect of monocytes from patients with infectious mononucleosis was reversible within three months after the onset of diesease. The cause of this monocyte migration defect in infectious mononucleosis is though to be an in vivo blockade of receptors on monocytes for chemotaxins, and it is speculated that this defect can partially explain the explain the ablated delayed-hypersensitivity skin reactions in this disease.

Correlating two-photon excited fluorescence imaging of breast cancer cellular redox state with seahorse flux analysis of normalized cellular oxygen consumption

NASA Astrophysics Data System (ADS)

Hou, Jue; Wright, Heather J.; Chan, Nicole; Tran, Richard; Razorenova, Olga V.; Potma, Eric O.; Tromberg, Bruce J.

2016-06-01

Two-photon excited fluorescence (TPEF) imaging of the cellular cofactors nicotinamide adenine dinucleotide and oxidized flavin adenine dinucleotide is widely used to measure cellular metabolism, both in normal and pathological cells and tissues. When dual-wavelength excitation is used, ratiometric TPEF imaging of the intrinsic cofactor fluorescence provides a metabolic index of cells-the "optical redox ratio" (ORR). With increased interest in understanding and controlling cellular metabolism in cancer, there is a need to evaluate the performance of ORR in malignant cells. We compare TPEF metabolic imaging with seahorse flux analysis of cellular oxygen consumption in two different breast cancer cell lines (MCF-7 and MDA-MB-231). We monitor metabolic index in living cells under both normal culture conditions and, for MCF-7, in response to cell respiration inhibitors and uncouplers. We observe a significant correlation between the TPEF-derived ORR and the flux analyzer measurements (R=0.7901, p<0.001). Our results confirm that the ORR is a valid dynamic index of cell metabolism under a range of oxygen consumption conditions relevant for cancer imaging.

Sub-cellular force microscopy in single normal and cancer cells.

PubMed

Babahosseini, H; Carmichael, B; Strobl, J S; Mahmoodi, S N; Agah, M

2015-08-07

This work investigates the biomechanical properties of sub-cellular structures of breast cells using atomic force microscopy (AFM). The cells are modeled as a triple-layered structure where the Generalized Maxwell model is applied to experimental data from AFM stress-relaxation tests to extract the elastic modulus, the apparent viscosity, and the relaxation time of sub-cellular structures. The triple-layered modeling results allow for determination and comparison of the biomechanical properties of the three major sub-cellular structures between normal and cancerous cells: the up plasma membrane/actin cortex, the mid cytoplasm/nucleus, and the low nuclear/integrin sub-domains. The results reveal that the sub-domains become stiffer and significantly more viscous with depth, regardless of cell type. In addition, there is a decreasing trend in the average elastic modulus and apparent viscosity of the all corresponding sub-cellular structures from normal to cancerous cells, which becomes most remarkable in the deeper sub-domain. The presented modeling in this work constitutes a unique AFM-based experimental framework to study the biomechanics of sub-cellular structures. Copyright © 2015 Elsevier Inc. All rights reserved.

Neutrophil dysregulation during sepsis: an overview and update.

PubMed

Shen, Xiao-Fei; Cao, Ke; Jiang, Jin-Peng; Guan, Wen-Xian; Du, Jun-Feng

2017-09-01

Sepsis remains a leading cause of death worldwide, despite advances in critical care, and understanding of the pathophysiology and treatment strategies. No specific therapy or drugs are available for sepsis. Neutrophils play a critical role in controlling infection under normal conditions, and it is suggested that their migration and antimicrobial activity are impaired during sepsis which contribute to the dysregulation of immune responses. Recent studies further demonstrated that interruption or reversal of the impaired migration and antimicrobial function of neutrophils improves the outcome of sepsis in animal models. In this review, we provide an overview of the associated mediators and signal pathways involved which govern the survival, migration and antimicrobial function of neutrophils in sepsis, and discuss the potential of neutrophils as a target to specifically diagnose and/or predict the outcome of sepsis. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

RORα, a Potential Tumor Suppressor and Therapeutic Target of Breast Cancer

PubMed Central

Du, Jun; Xu, Ren

2012-01-01

The function of the nuclear receptor (NR) in breast cancer progression has been investigated for decades. The majority of the nuclear receptors have well characterized natural ligands, but a few of them are orphan receptors for which no ligand has been identified. RORα, one member of the retinoid orphan nuclear receptor (ROR) subfamily of orphan receptors, regulates various cellular and pathological activities. RORα is commonly down-regulated and/or hypoactivated in breast cancer compared to normal mammary tissue. Expression of RORα suppresses malignant phenotypes in breast cancer cells, in vitro and in vivo. Activity of RORα can be categorized into the canonical and non-canonical nuclear receptor pathways, which in turn regulate various breast cancer cellular function, including cell proliferation, apoptosis and invasion. This information suggests that RORα is a potent tumor suppressor and a potential therapeutic target for breast cancer. PMID:23443091

Restoring functional neurofibromin by protein transduction.

PubMed

Mellert, K; Lechner, S; Lüdeke, M; Lamla, M; Möller, P; Kemkemer, R; Scheffzek, K; Kaufmann, D

2018-04-18

In Neurofibromatosis 1 (NF1) germ line loss of function mutations result in reduction of cellular neurofibromin content (NF1+/-, NF1 haploinsufficiency). The Ras-GAP neurofibromin is a very large cytoplasmic protein (2818 AA, 319 kDa) involved in the RAS-MAPK pathway. Aside from regulation of proliferation, it is involved in mechanosensoric of cells. We investigated neurofibromin replacement in cultured human fibroblasts showing reduced amount of neurofibromin. Full length neurofibromin was produced recombinantly in insect cells and purified. Protein transduction into cultured fibroblasts was performed employing cell penetrating peptides along with photochemical internalization. This combination of transduction strategies ensures the intracellular uptake and the translocation to the cytoplasm of neurofibromin. The transduced neurofibromin is functional, indicated by functional rescue of reduced mechanosensoric blindness and reduced RasGAP activity in cultured fibroblasts of NF1 patients or normal fibroblasts treated by NF1 siRNA. Our study shows that recombinant neurofibromin is able to revert cellular effects of NF1 haploinsuffiency in vitro, indicating a use of protein transduction into cells as a potential treatment strategy for the monogenic disease NF1.

Hyaluronan – A Functional and Structural Sweet Spot in the Tissue Microenvironment

PubMed Central

Monslow, James; Govindaraju, Priya; Puré, Ellen

2015-01-01

Transition from homeostatic to reactive matrix remodeling is a fundamental adaptive tissue response to injury, inflammatory disease, fibrosis, and cancer. Alterations in architecture, physical properties, and matrix composition result in changes in biomechanical and biochemical cellular signaling. The dynamics of pericellular and extracellular matrices, including matrix protein, proteoglycan, and glycosaminoglycan modification are continually emerging as essential regulatory mechanisms underlying cellular and tissue function. Nevertheless, the impact of matrix organization on inflammation and immunity in particular and the consequent effects on tissue healing and disease outcome are arguably under-studied aspects of adaptive stress responses. Herein, we review how the predominant glycosaminoglycan hyaluronan (HA) contributes to the structure and function of the tissue microenvironment. Specifically, we examine the evidence of HA degradation and the generation of biologically active smaller HA fragments in pathological settings in vivo. We discuss how HA fragments versus nascent HA via alternate receptor-mediated signaling influence inflammatory cell recruitment and differentiation, resident cell activation, as well as tumor growth, survival, and metastasis. Finally, we discuss how HA fragmentation impacts restoration of normal tissue function and pathological outcomes in disease. PMID:26029216

The Function of V-ATPases in Cancer

PubMed Central

Stransky, Laura; Cotter, Kristina

2016-01-01

The vacuolar ATPases (V-ATPases) are a family of proton pumps that couple ATP hydrolysis to proton transport into intracellular compartments and across the plasma membrane. They function in a wide array of normal cellular processes, including membrane traffic, protein processing and degradation, and the coupled transport of small molecules, as well as such physiological processes as urinary acidification and bone resorption. The V-ATPases have also been implicated in a number of disease processes, including viral infection, renal disease, and bone resorption defects. This review is focused on the growing evidence for the important role of V-ATPases in cancer. This includes functions in cellular signaling (particularly Wnt, Notch, and mTOR signaling), cancer cell survival in the highly acidic environment of tumors, aiding the development of drug resistance, as well as crucial roles in tumor cell invasion, migration, and metastasis. Of greatest excitement is evidence that at least some tumors express isoforms of V-ATPase subunits whose disruption is not lethal, leading to the possibility of developing anti-cancer therapeutics that selectively target V-ATPases that function in cancer cells. PMID:27335445

The Impact of Sleep Deprivation on the Brain

PubMed

Trošt Bobić, Tatjana; Šečić, Ana; Zavoreo, Iris; Matijević, Valentina; Filipović, Branimir; Kolak, Željka; Bašić Kes, Vanja; Ciliga, Dubravka; Sajković, Dubravka

2016-09-01

Each sleep phase is characterized by specific chemical, cellular and anatomic events of vital importance for normal neural functioning. Different forms of sleep deprivation may lead to a decline of cognitive functions in individuals. Studies in this field make a distinction between total sleep deprivation, chronic sleep restriction, and the situation of sleep disruption. Investigations covering the acute effects of sleep deprivation on the brain show that the discovered behavioral deficits in most cases regenerate after two nights of complete sleep. However, some studies done on mice emphasize the possible chronic effects of long-term sleep deprivation or chronic restriction on the occurrence of neurodegenerative diseases such as Alzheimer’s disease and dementia. In order to better understand the acute and chronic effects of sleep loss, the mechanisms of neural adaptation in the situations of insufficient sleep need to be further investigated. Future integrative research on the impact of sleep deprivation on neural functioning measured through the macro level of cognitive functions and the micro molecular and cell level could contribute to more accurate conclusions about the basic cellular mechanisms responsible for the detected behavioral deficits occurring due to sleep deprivation.

The emerging role of nuclear viral DNA sensors.

PubMed

Diner, Benjamin A; Lum, Krystal K; Cristea, Ileana M

2015-10-30

Detecting pathogenic DNA by intracellular receptors termed "sensors" is critical toward galvanizing host immune responses and eliminating microbial infections. Emerging evidence has challenged the dogma that sensing of viral DNA occurs exclusively in sub-cellular compartments normally devoid of cellular DNA. The interferon-inducible protein IFI16 was shown to bind nuclear viral DNA and initiate immune signaling, culminating in antiviral cytokine secretion. Here, we review the newly characterized nucleus-originating immune signaling pathways, their links to other crucial host defenses, and unique mechanisms by which viruses suppress their functions. We frame these findings in the context of human pathologies associated with nuclear replicating DNA viruses. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Endoplasmic Reticulum Stress and Unfolded Protein Response Pathways: Potential for Treating Age-related Retinal Degeneration

PubMed Central

Haeri, Mohammad; Knox, Barry E

2012-01-01

Accumulation of misfolded proteins in the endoplasmic reticulum (ER) and their aggregation impair normal cellular function and can be toxic, leading to cell death. Prolonged expression of misfolded proteins triggers ER stress, which initiates a cascade of reactions called the unfolded protein response (UPR). Protein misfolding is the basis for a variety of disorders known as ER storage or conformational diseases. There are an increasing number of eye disorders associated with misfolded proteins and pathologic ER responses, including retinitis pigmentosa (RP). Herein we review the basic cellular and molecular biology of UPR with focus on pathways that could be potential targets for treating retinal degenerative diseases. PMID:22737387

Animal models to study microRNA function

PubMed Central

Pal, Arpita S.; Kasinski, Andrea L.

2018-01-01

The discovery of the microRNAs, lin-4 and let-7 as critical mediators of normal development in Caenorhabditis elegans and their conservation throughout evolution has spearheaded research towards identifying novel roles of microRNAs in other cellular processes. To accurately elucidate these fundamental functions, especially in the context of an intact organism various microRNA transgenic models have been generated and evaluated. Transgenic C. elegans (worms), Drosophila melanogaster (flies), Danio rerio (zebrafish), and Mus musculus (mouse) have contributed immensely towards uncovering the roles of multiple microRNAs in cellular processes such as proliferation, differentiation, and apoptosis, pathways that are severely altered in human diseases such as cancer. The simple model organisms, C. elegans, D. melanogaster and D. rerio do not develop cancers, but have proved to be convenient systesm in microRNA research, especially in characterizing the microRNA biogenesis machinery which is often dysregulated during human tumorigenesis. The microRNA-dependent events delineated via these simple in vivo systems have been further verified in vitro, and in more complex models of cancers, such as M. musculus. The focus of this review is to provide an overview of the important contributions made in the microRNA field using model organisms. The simple model systems provided the basis for the importance of microRNAs in normal cellular physiology, while the more complex animal systems provided evidence for the role of microRNAs dysregulation in cancers. Highlights include an overview of the various strategies used to generate transgenic organisms and a review of the use of transgenic mice for evaluating pre-clinical efficacy of microRNA-based cancer therapeutics. PMID:28882225

Integrated biocircuits: engineering functional multicellular circuits and devices.

PubMed

Prox, Jordan; Smith, Tory; Holl, Chad; Chehade, Nick; Guo, Liang

2018-04-01

Implantable neurotechnologies have revolutionized neuromodulatory medicine for treating the dysfunction of diseased neural circuitry. However, challenges with biocompatibility and lack of full control over neural network communication and function limits the potential to create more stable and robust neuromodulation devices. Thus, we propose a platform technology of implantable and programmable cellular systems, namely Integrated Biocircuits, which use only cells as the functional components of the device. We envision the foundational principles for this concept begins with novel in vitro platforms used for the study and reconstruction of cellular circuitry. Additionally, recent advancements in organoid and 3D culture systems account for microenvironment factors of cytoarchitecture to construct multicellular circuits as they are normally formed in the brain. We explore the current state of the art of these platforms to provide knowledge of their advancements in circuit fabrication and identify the current biological principles that could be applied in designing integrated biocircuit devices. We have highlighted the exemplary methodologies and techniques of in vitro circuit fabrication and propose the integration of selected controllable parameters, which would be required in creating suitable biodevices. We provide our perspective and propose new insights into the future of neuromodulaion devices within the scope of living cellular systems that can be applied in designing more reliable and biocompatible stimulation-based neuroprosthetics.

Integrated biocircuits: engineering functional multicellular circuits and devices

NASA Astrophysics Data System (ADS)

Prox, Jordan; Smith, Tory; Holl, Chad; Chehade, Nick; Guo, Liang

2018-04-01

Objective. Implantable neurotechnologies have revolutionized neuromodulatory medicine for treating the dysfunction of diseased neural circuitry. However, challenges with biocompatibility and lack of full control over neural network communication and function limits the potential to create more stable and robust neuromodulation devices. Thus, we propose a platform technology of implantable and programmable cellular systems, namely Integrated Biocircuits, which use only cells as the functional components of the device. Approach. We envision the foundational principles for this concept begins with novel in vitro platforms used for the study and reconstruction of cellular circuitry. Additionally, recent advancements in organoid and 3D culture systems account for microenvironment factors of cytoarchitecture to construct multicellular circuits as they are normally formed in the brain. We explore the current state of the art of these platforms to provide knowledge of their advancements in circuit fabrication and identify the current biological principles that could be applied in designing integrated biocircuit devices. Main results. We have highlighted the exemplary methodologies and techniques of in vitro circuit fabrication and propose the integration of selected controllable parameters, which would be required in creating suitable biodevices. Significance. We provide our perspective and propose new insights into the future of neuromodulaion devices within the scope of living cellular systems that can be applied in designing more reliable and biocompatible stimulation-based neuroprosthetics.

Fumarate hydratase is a critical metabolic regulator of hematopoietic stem cell functions.

PubMed

Guitart, Amelie V; Panagopoulou, Theano I; Villacreces, Arnaud; Vukovic, Milica; Sepulveda, Catarina; Allen, Lewis; Carter, Roderick N; van de Lagemaat, Louie N; Morgan, Marcos; Giles, Peter; Sas, Zuzanna; Gonzalez, Marta Vila; Lawson, Hannah; Paris, Jasmin; Edwards-Hicks, Joy; Schaak, Katrin; Subramani, Chithra; Gezer, Deniz; Armesilla-Diaz, Alejandro; Wills, Jimi; Easterbrook, Aaron; Coman, David; So, Chi Wai Eric; O'Carroll, Donal; Vernimmen, Douglas; Rodrigues, Neil P; Pollard, Patrick J; Morton, Nicholas M; Finch, Andrew; Kranc, Kamil R

2017-03-06

Strict regulation of stem cell metabolism is essential for tissue functions and tumor suppression. In this study, we investigated the role of fumarate hydratase (Fh1), a key component of the mitochondrial tricarboxylic acid (TCA) cycle and cytosolic fumarate metabolism, in normal and leukemic hematopoiesis. Hematopoiesis-specific Fh1 deletion (resulting in endogenous fumarate accumulation and a genetic TCA cycle block reflected by decreased maximal mitochondrial respiration) caused lethal fetal liver hematopoietic defects and hematopoietic stem cell (HSC) failure. Reexpression of extramitochondrial Fh1 (which normalized fumarate levels but not maximal mitochondrial respiration) rescued these phenotypes, indicating the causal role of cellular fumarate accumulation. However, HSCs lacking mitochondrial Fh1 (which had normal fumarate levels but defective maximal mitochondrial respiration) failed to self-renew and displayed lymphoid differentiation defects. In contrast, leukemia-initiating cells lacking mitochondrial Fh1 efficiently propagated Meis1 / Hoxa9 -driven leukemia. Thus, we identify novel roles for fumarate metabolism in HSC maintenance and hematopoietic differentiation and reveal a differential requirement for mitochondrial Fh1 in normal hematopoiesis and leukemia propagation. © 2017 Guitart et al.

Lab-On-Chip Clinorotation System for Live-Cell Microscopy Under Simulated Microgravity

NASA Technical Reports Server (NTRS)

Yew, Alvin G.; Atencia, Javier; Chinn, Ben; Hsieh, Adam H.

2013-01-01

Cells in microgravity are subject to mechanical unloading and changes to the surrounding chemical environment. How these factors jointly influence cellular function is not well understood. We can investigate their role using ground-based analogues to spaceflight, where mechanical unloading is simulated through the time-averaged nullification of gravity. The prevailing method for cellular microgravity simulation is to use fluid-filled containers called clinostats. However, conventional clinostats are not designed for temporally tracking cell response, nor are they able to establish dynamic fluid environments. To address these needs, we developed a Clinorotation Time-lapse Microscopy (CTM) system that accommodates lab-on- chip cell culture devices for visualizing time-dependent alterations to cellular behavior. For the purpose of demonstrating CTM, we present preliminary results showing time-dependent differences in cell area between human mesenchymal stem cells (hMSCs) under modeled microgravity and normal gravity.

Lab-On-Chip Clinorotation System for Live-Cell Microscopy Under Simulated Microgravity

NASA Technical Reports Server (NTRS)

Yew, Alvin G.; Atencia, Javier; Chinn, Ben; Hsieh, Adam H.

1980-01-01

Cells in microgravity are subject to mechanical unloading and changes to the surrounding chemical environment. How these factors jointly influence cellular function is not well understood. We can investigate their role using ground-based analogues to spaceflight, where mechanical unloading is simulated through the time-averaged nullification of gravity. The prevailing method for cellular microgravity simulation is to use fluid-filled containers called clinostats. However, conventional clinostats are not designed for temporally tracking cell response, nor are they able to establish dynamic fluid environments. To address these needs, we developed a Clinorotation Time-lapse Microscopy (CTM) system that accommodates lab-on- chip cell culture devices for visualizing time-dependent alterations to cellular behavior. For the purpose of demonstrating CTM, we present preliminary results showing time-dependent differences in cell area between human mesenchymal stem cells (hMSCs) under modeled microgravity and normal gravity.

Simulating large-scale pedestrian movement using CA and event driven model: Methodology and case study

NASA Astrophysics Data System (ADS)

Li, Jun; Fu, Siyao; He, Haibo; Jia, Hongfei; Li, Yanzhong; Guo, Yi

2015-11-01

Large-scale regional evacuation is an important part of national security emergency response plan. Large commercial shopping area, as the typical service system, its emergency evacuation is one of the hot research topics. A systematic methodology based on Cellular Automata with the Dynamic Floor Field and event driven model has been proposed, and the methodology has been examined within context of a case study involving the evacuation within a commercial shopping mall. Pedestrians walking is based on Cellular Automata and event driven model. In this paper, the event driven model is adopted to simulate the pedestrian movement patterns, the simulation process is divided into normal situation and emergency evacuation. The model is composed of four layers: environment layer, customer layer, clerk layer and trajectory layer. For the simulation of movement route of pedestrians, the model takes into account purchase intention of customers and density of pedestrians. Based on evacuation model of Cellular Automata with Dynamic Floor Field and event driven model, we can reflect behavior characteristics of customers and clerks at the situations of normal and emergency evacuation. The distribution of individual evacuation time as a function of initial positions and the dynamics of the evacuation process is studied. Our results indicate that the evacuation model using the combination of Cellular Automata with Dynamic Floor Field and event driven scheduling can be used to simulate the evacuation of pedestrian flows in indoor areas with complicated surroundings and to investigate the layout of shopping mall.

Rewiring of cellular membrane homeostasis by picornaviruses.

PubMed

Belov, George A; Sztul, Elizabeth

2014-09-01

Viruses are obligatory intracellular parasites and utilize host elements to support key viral processes, including penetration of the plasma membrane, initiation of infection, replication, and suppression of the host's antiviral defenses. In this review, we focus on picornaviruses, a family of positive-strand RNA viruses, and discuss the mechanisms by which these viruses hijack the cellular machinery to form and operate membranous replication complexes. Studies aimed at revealing factors required for the establishment of viral replication structures identified several cellular-membrane-remodeling proteins and led to the development of models in which the virus used a preexisting cellular-membrane-shaping pathway "as is" for generating its replication organelles. However, as more data accumulate, this view is being increasingly questioned, and it is becoming clearer that viruses may utilize cellular factors in ways that are distinct from the normal functions of these proteins in uninfected cells. In addition, the proteincentric view is being supplemented by important new studies showing a previously unappreciated deep remodeling of lipid homeostasis, including extreme changes to phospholipid biosynthesis and cholesterol trafficking. The data on viral modifications of lipid biosynthetic pathways are still rudimentary, but it appears once again that the viruses may rewire existing pathways to generate novel functions. Despite remarkable progress, our understanding of how a handful of viral proteins can completely overrun the multilayered, complex mechanisms that control the membrane organization of a eukaryotic cell remains very limited. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Emerging roles of microRNAs as molecular switches in the integrated circuit of the cancer cell

PubMed Central

Sotiropoulou, Georgia; Pampalakis, Georgios; Lianidou, Evi; Mourelatos, Zissimos

2009-01-01

Transformation of normal cells into malignant tumors requires the acquisition of six hallmark traits, e.g., self-sufficiency in growth signals, insensitivity to antigrowth signals and self-renewal, evasion of apoptosis, limitless replication potential, angiogenesis, invasion, and metastasis, which are common to all cancers (Hanahan and Weinberg 2000). These new cellular traits evolve from defects in major regulatory microcircuits that are fundamental for normal homeostasis. The discovery of microRNAs (miRNAs) as a new class of small non-protein-coding RNAs that control gene expression post-transcriptionally by binding to various mRNA targets suggests that these tiny RNA molecules likely act as molecular switches in the extensive regulatory web that involves thousands of transcripts. Most importantly, accumulating evidence suggests that numerous microRNAs are aberrantly expressed in human cancers. In this review, we discuss the emergent roles of microRNAs as switches that function to turn on/off known cellular microcircuits. We outline recent compelling evidence that deregulated microRNA-mediated control of cellular microcircuits cooperates with other well-established regulatory mechanisms to confer the hallmark traits of the cancer cell. Furthermore, these exciting insights into aberrant microRNA control in cancer-associated circuits may be exploited for cancer therapies that will target deregulated miRNA switches. PMID:19561119

In vitro evaluation of low-intensity light radiation on murine melanoma (B16F10) cells.

PubMed

Peidaee, P; Almansour, N M; Pirogova, E

2016-03-01

Changes in the energy state of biomolecules induced by electromagnetic radiation lead to changes in biological functions of irradiated biomolecules. Using the RRM approach, it was computationally predicted that far-infrared light irradiation in the range of 3500-6000 nm affects biological activity of proto-oncogene proteins. This in vitro study evaluates quantitatively and qualitatively the effects of selected far-infrared exposures in the computationally determined wavelengths on mouse melanoma B16F10 cells and Chinese hamster ovarian (CHO) cells by MTT (thiazolyl blue tetrazolium bromide) cell proliferation assay and confocal laser-scanning microscopy (CLSM). This paper also presents the findings obtained from irradiating B16F10 and CHO cells by the selected wavelengths in visible and near-infrared range. The MTT results show that far-infrared wavelength irradiation induces detrimental effect on cellular viability of B16F10 cells, while that of normal CHO cells is not affected considerably. Moreover, CLSM images demonstrate visible cellular detachment of cancer cells. The observed effects support the hypothesis that far-infrared light irradiation within the computationally determined wavelength range induces biological effect on cancer cells. From irradiation of selected visible and near-infrared wavelengths, no visible changes were detected in cellular viability of either normal or cancer cells.

Hydroxylamine derivatives for regulation of spermine and spermidine metabolism.

PubMed

Khomutov, M A; Weisell, J; Hyvönen, M; Keinänen, T A; Vepsäläinen, J; Alhonen, L; Khomutov, A R; Kochetkov, S N

2013-12-01

The biogenic polyamines spermine, spermidine, and their precursor putrescine are present in micro-to-millimolar concentrations in all cell types and are vitally important for their normal growth. High intracellular content of spermine and spermidine determines the multiplicity of the cellular functions of the polyamines. Many of these functions are not well characterized at the molecular level, ensuring the ongoing development of this field of biochemistry. Tumor cells have elevated polyamine level if compared with normal cells, and this greatly stimulates the search for new opportunities to deplete the intracellular pool of spermine and spermidine resulting in decrease in cell growth and even cell death. O-Substituted hydroxylamines occupy their own place among chemical regulators of the activity of the enzymes of polyamine metabolism. Varying the structure of the alkyl substituent made it possible to obtain within one class of chemical compounds highly effective inhibitors and regulators of the activity of all the enzymes of putrescine, spermine and spermidine metabolism (with the exception of FAD-dependent spermine oxidase and acetylpolyamine oxidase), effectors of the polyamine transport system, and even actively transported in cells "proinhibitor" of ornithine decarboxylase. Some principles for the design of specific inhibitors of these enzymes as well as the peculiarities of cellular effects of corresponding O-substituted hydroxylamines are discussed.

Separation of metabolic supply and demand: aerobic glycolysis as a normal physiological response to fluctuating energetic demands in the membrane.

PubMed

Epstein, Tamir; Xu, Liping; Gillies, Robert J; Gatenby, Robert A

2014-01-01

Cancer cells, and a variety of normal cells, exhibit aerobic glycolysis, high rates of glucose fermentation in the presence of normal oxygen concentrations, also known as the Warburg effect. This metabolism is considered abnormal because it violates the standard model of cellular energy production that assumes glucose metabolism is predominantly governed by oxygen concentrations and, therefore, fermentative glycolysis is an emergency back-up for periods of hypoxia. Though several hypotheses have been proposed for the origin of aerobic glycolysis, its biological basis in cancer and normal cells is still not well understood. We examined changes in glucose metabolism following perturbations in membrane activity in different normal and tumor cell lines and found that inhibition or activation of pumps on the cell membrane led to reduction or increase in glycolysis, respectively, while oxidative phosphorylation remained unchanged. Computational simulations demonstrated that these findings are consistent with a new model of normal physiological cellular metabolism in which efficient mitochondrial oxidative phosphorylation supplies chronic energy demand primarily for macromolecule synthesis and glycolysis is necessary to supply rapid energy demands primarily to support membrane pumps. A specific model prediction was that the spatial distribution of ATP-producing enzymes in the glycolytic pathway must be primarily localized adjacent to the cell membrane, while mitochondria should be predominantly peri-nuclear. The predictions were confirmed experimentally. Our results show that glycolytic metabolism serves a critical physiological function under normoxic conditions by responding to rapid energetic demand, mainly from membrane transport activities, even in the presence of oxygen. This supports a new model for glucose metabolism in which glycolysis and oxidative phosphorylation supply different types of energy demand. Cells use efficient but slow-responding aerobic metabolism to meet baseline, steady energy demand and glycolytic metabolism, which is inefficient but can rapidly increase adenosine triphosphate (ATP) production, to meet short-timescale energy demands, mainly from membrane transport activities. In this model, the origin of the Warburg effect in cancer cells and aerobic glycolysis in general represents a normal physiological function due to enhanced energy demand for membrane transporters activity required for cell division, growth, and migration.

Confocal Microscopy and Molecular-Specific Optical Contrast Agents for the Detection of Oral Neoplasia

PubMed Central

Carlson, Alicia L.; Gillenwater, Ann M.; Williams, Michelle D.; El-Naggar, Adel K.; Richards-Kortum, R. R.

2009-01-01

Using current clinical diagnostic techniques, it is difficult to visualize tumor morphology and architecture at the cellular level, which is necessary for diagnostic localization of pathologic lesions. Optical imaging techniques have the potential to address this clinical need by providing real-time, sub-cellular resolution images. This paper describes the use of dual mode confocal microscopy and optical molecular-specific contrast agents to image tissue architecture, cellular morphology, and sub-cellular molecular features of normal and neoplastic oral tissues. Fresh tissue slices were prepared from 33 biopsies of clinically normal and abnormal oral mucosa obtained from 14 patients. Reflectance confocal images were acquired after the application of 6% acetic acid, and fluorescence confocal images were acquired after the application of a fluorescence contrast agent targeting the epidermal growth factor receptor (EGFR). The dual imaging modes provided images similar to light microscopy of hematoxylin and eosin and immunohistochemistry staining, but from thick fresh tissue slices. Reflectance images provided information on the architecture of the tissue and the cellular morphology. The nuclear-to-cytoplasmic (N/C) ratio from the reflectance images was at least 7.5 times greater for the carcinoma than the corresponding normal samples, except for one case of highly keratinized carcinoma. Separation of carcinoma from normal and mild dysplasia was achieved using this ratio (p<0.01). Fluorescence images of EGFR expression yielded a mean fluorescence labeling intensity (FLI) that was at least 2.7 times higher for severe dysplasia and carcinoma samples than for the corresponding normal sample, and could be used to distinguish carcinoma from normal and mild dysplasia (p<0.01). Analyzed together, the N/C ratio and the mean FLI may improve the ability to distinguish carcinoma from normal squamous epithelium. PMID:17877424

Surface modification of microparticles causes differential uptake responses in normal and tumoral human breast epithelial cells

PubMed Central

Patiño, Tania; Soriano, Jorge; Barrios, Lleonard; Ibáñez, Elena; Nogués, Carme

2015-01-01

The use of micro- and nanodevices as multifunctional systems for biomedical applications has experienced an exponential growth during the past decades. Although a large number of studies have focused on the design and fabrication of new micro- and nanosystems capable of developing multiple functions, a deeper understanding of their interaction with cells is required. In the present study, we evaluated the effect of different microparticle surfaces on their interaction with normal and tumoral human breast epithelial cell lines. For this, AlexaFluor488 IgG functionalized polystyrene microparticles (3 μm) were coated with Polyethyleneimine (PEI) at two different molecular weights, 25 and 750 kDa. The effect of microparticle surface properties on cytotoxicity, cellular uptake and endocytic pathways were assessed for both normal and tumoral cell lines. Results showed a differential response between the two cell lines regarding uptake efficiency and mechanisms of endocytosis, highlighting the potential role of microparticle surface tunning for specific cell targeting. PMID:26068810

Surface modification of microparticles causes differential uptake responses in normal and tumoral human breast epithelial cells

NASA Astrophysics Data System (ADS)

Patiño, Tania; Soriano, Jorge; Barrios, Lleonard; Ibáñez, Elena; Nogués, Carme

2015-06-01

The use of micro- and nanodevices as multifunctional systems for biomedical applications has experienced an exponential growth during the past decades. Although a large number of studies have focused on the design and fabrication of new micro- and nanosystems capable of developing multiple functions, a deeper understanding of their interaction with cells is required. In the present study, we evaluated the effect of different microparticle surfaces on their interaction with normal and tumoral human breast epithelial cell lines. For this, AlexaFluor488 IgG functionalized polystyrene microparticles (3 μm) were coated with Polyethyleneimine (PEI) at two different molecular weights, 25 and 750 kDa. The effect of microparticle surface properties on cytotoxicity, cellular uptake and endocytic pathways were assessed for both normal and tumoral cell lines. Results showed a differential response between the two cell lines regarding uptake efficiency and mechanisms of endocytosis, highlighting the potential role of microparticle surface tunning for specific cell targeting.

Surface modification of microparticles causes differential uptake responses in normal and tumoral human breast epithelial cells.

PubMed

Patiño, Tania; Soriano, Jorge; Barrios, Lleonard; Ibáñez, Elena; Nogués, Carme

2015-06-12

The use of micro- and nanodevices as multifunctional systems for biomedical applications has experienced an exponential growth during the past decades. Although a large number of studies have focused on the design and fabrication of new micro- and nanosystems capable of developing multiple functions, a deeper understanding of their interaction with cells is required. In the present study, we evaluated the effect of different microparticle surfaces on their interaction with normal and tumoral human breast epithelial cell lines. For this, AlexaFluor488 IgG functionalized polystyrene microparticles (3 μm) were coated with Polyethyleneimine (PEI) at two different molecular weights, 25 and 750 kDa. The effect of microparticle surface properties on cytotoxicity, cellular uptake and endocytic pathways were assessed for both normal and tumoral cell lines. Results showed a differential response between the two cell lines regarding uptake efficiency and mechanisms of endocytosis, highlighting the potential role of microparticle surface tunning for specific cell targeting.

Cellular membrane trafficking of mesoporous silica nanoparticles

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

Fang, I-Ju

This dissertation mainly focuses on the investigation of the cellular membrane trafficking of mesoporous silica nanoparticles. We are interested in the study of endocytosis and exocytosis behaviors of mesoporous silica nanoparticles with desired surface functionality. The relationship between mesoporous silica nanoparticles and membrane trafficking of cells, either cancerous cells or normal cells was examined. Since mesoporous silica nanoparticles were applied in many drug delivery cases, the endocytotic efficiency of mesoporous silica nanoparticles needs to be investigated in more details in order to design the cellular drug delivery system in the controlled way. It is well known that cells can engulfmore » some molecules outside of the cells through a receptor-ligand associated endocytosis. We are interested to determine if those biomolecules binding to cell surface receptors can be utilized on mesoporous silica nanoparticle materials to improve the uptake efficiency or govern the mechanism of endocytosis of mesoporous silica nanoparticles. Arginine-glycine-aspartate (RGD) is a small peptide recognized by cell integrin receptors and it was reported that avidin internalization was highly promoted by tumor lectin. Both RGD and avidin were linked to the surface of mesoporous silica nanoparticle materials to investigate the effect of receptor-associated biomolecule on cellular endocytosis efficiency. The effect of ligand types, ligand conformation and ligand density were discussed in Chapter 2 and 3. Furthermore, the exocytosis of mesoporous silica nanoparticles is very attractive for biological applications. The cellular protein sequestration study of mesoporous silica nanoparticles was examined for further information of the intracellular pathway of endocytosed mesoporous silica nanoparticle materials. The surface functionality of mesoporous silica nanoparticle materials demonstrated selectivity among the materials and cancer and normal cell lines. We aimed to determine the specific organelle that mesoporous silica nanoparticles could approach via the identification of harvested proteins from exocytosis process. Based on the study of endo- and exocytosis behavior of mesoporous silica nanoparticle materials, we can design smarter drug delivery vehicles for cancer therapy that can be effectively controlled. The destination, uptake efficiency and the cellular distribution of mesoporous silica nanoparticle materials can be programmable. As a result, release mechanism and release rate of drug delivery systems can be a well-controlled process. The deep investigation of an endo- and exocytosis study of mesoporous silica nanoparticle materials promotes the development of drug delivery applications.« less

Membrane Lipid Replacement for chronic illnesses, aging and cancer using oral glycerolphospholipid formulations with fructooligosaccharides to restore phospholipid function in cellular membranes, organelles, cells and tissues.

PubMed

Nicolson, Garth L; Ash, Michael E

2017-09-01

Membrane Lipid Replacement is the use of functional, oral supplements containing mixtures of cell membrane glycerolphospholipids, plus fructooligosaccharides (for protection against oxidative, bile acid and enzymatic damage) and antioxidants, in order to safely replace damaged, oxidized, membrane phospholipids and restore membrane, organelle, cellular and organ function. Defects in cellular and intracellular membranes are characteristic of all chronic medical conditions, including cancer, and normal processes, such as aging. Once the replacement glycerolphospholipids have been ingested, dispersed, complexed and transported, while being protected by fructooligosaccharides and several natural mechanisms, they can be inserted into cell membranes, lipoproteins, lipid globules, lipid droplets, liposomes and other carriers. They are conveyed by the lymphatics and blood circulation to cellular sites where they are endocytosed or incorporated into or transported by cell membranes. Inside cells the glycerolphospholipids can be transferred to various intracellular membranes by lipid globules, liposomes, membrane-membrane contact or by lipid carrier transfer. Eventually they arrive at their membrane destinations due to 'bulk flow' principles, and there they can stimulate the natural removal and replacement of damaged membrane lipids while undergoing further enzymatic alterations. Clinical trials have shown the benefits of Membrane Lipid Replacement in restoring mitochondrial function and reducing fatigue in aged subjects and chronically ill patients. Recently Membrane Lipid Replacement has been used to reduce pain and other symptoms as well as removing hydrophobic chemical contaminants, suggesting that there are additional new uses for this safe, natural medicine supplement. This article is part of a Special Issue entitled: Membrane Lipid Therapy: Drugs Targeting Biomembranes edited by Pablo V. Escribá. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

Molecular pathophysiology of SLC4 bicarbonate transporters.

PubMed

Romero, Michael F

2005-09-01

Acid-base (H and HCO3) transport in the kidney is crucial for maintaining blood pH, cellular pH and excreting metabolic acid. HCO3 transport in the kidney is mediated by HCO3 transporter proteins which occur in two gene families in humans, vertebrates and invertebrates (SLC4 and SLC26). Since SLC26 transporters have other, non-HCO3 transport functions, this review highlights the history and recent advances in the SLC4 transporters in the kidney. The SLC4 gene and protein family (10 genes) contains three types of HCO3 transporters: Cl-HCO3 exchangers, Na/HCO3 cotransporters and Na-driven Cl-HCO3 exchangers. Function and human chromosomal location have been determined for most members. Human mutations in AE1 (SLC4A1) and NBCe1 (SLC4A4) are associated with distal and proximal renal tubular acidosis, respectively. Recent advances include the cellular and biophysical mechanisms by which AE1 and NBCe1 mutations lead to renal disease. Mutational and cellular trafficking studies have begun to elucidate the membrane topology and functional domains of AE1 and NBCe1. Knockout mice for AE2 and NBCn1 do not have obvious renal phenotypes. Recently, SLC4A11 (bicarbonate transporter 1) was shown to function as an electrogenic Na/borate cotransporter unable to transport HCO3 but involved in cell cycle control. SLC4 HCO3 transporters play critical roles in systemic and cellular pH homeostasis. Most of the SLC4 members are present at some level in the kidney. Future studies will likely continue to make use of knockout animals, for example mice and zebrafish, human mutations or polymorphisms to elucidate the normal and pathophysiologic roles of these proteins.

The cell biology of aging.

PubMed

Hayflick, L

1985-02-01

It is only within the past ten years that biogerontology has become attractive to a sufficient number of biologists so that the field can be regarded as a seriously studied discipline. Cytogerontology, or the study of aging at the cellular level, had its genesis about 20 years ago when the dogma that maintained that cultured normal cells could replicate forever was overturned. Normal human and animal cells have a finite capacity to replicate and function whether they are cultured in vitro or transplanted as grafts in vivo. This phenomenon has been interpreted to be aging at the cellular level. Only abnormal somatic cells are capable of immortality. In recent years it has been found that the number of population doublings of which cultured normal cells are capable is inversely proportional to donor age. There is also good evidence that the number of population doublings of cultured normal fibroblasts is directly proportional to the maximum lifespan of ten species that have been studied. Cultures prepared from patients with accelerated aging syndromes (progeria and Werner's syndrome) undergo far fewer doublings than do those of age-matched controls. The normal human fibroblast cell strain WI-38 was established in 1962 from fetal lung, and several hundred ampules of these cells were frozen in liquid nitrogen at that time. These ampules have been reconstituted periodically and shown to be capable of replication. This represents the longest period of time that a normal human cell has ever been frozen. Normal human fetal cell strains such as WI-38 have the capacity to double only about 50 times. If cultures are frozen at various population doublings, the number of doublings remaining after reconstitution is equal to 50 minus the number of doublings that occurred prior to freezing. The memory of the cells has been found to be accurate after 23 years of preservation in liquid nitrogen. Normal human cells incur many physiologic decrements that herald the approach of their failure to divide. Many of these functional decrements are identical to decrements found in humans as they age. Thus it is likely that these decrements are also the precursors of age changes in vivo. The finite replicative capacity of normal cells is never seen to occur in vivo because aging and death of the individual occurs well before the doubling limit is reached.

STAT proteins: from normal control of cellular events to tumorigenesis.

PubMed

Calò, Valentina; Migliavacca, Manuela; Bazan, Viviana; Macaluso, Marcella; Buscemi, Maria; Gebbia, Nicola; Russo, Antonio

2003-11-01

Signal transducers and activators of transcription (STAT) proteins comprise a family of transcription factors latent in the cytoplasm that participate in normal cellular events, such as differentiation, proliferation, cell survival, apoptosis, and angiogenesis following cytokine, growth factor, and hormone signaling. STATs are activated by tyrosine phosphorylation, which is normally a transient and tightly regulates process. Nevertheless, several constitutively activated STATs have been observed in a wide number of human cancer cell lines and primary tumors, including blood malignancies and solid neoplasias. STATs can be divided into two groups according to their specific functions. One is made up of STAT2, STAT4, and STAT6, which are activated by a small number of cytokines and play a distinct role in the development of T-cells and in IFNgamma signaling. The other group includes STAT1, STAT3, and STAT5, activated in different tissues by means of a series of ligands and involved in IFN signaling, development of the mammary gland, response to GH, and embriogenesis. This latter group of STATS plays an important role in controlling cell-cycle progression and apoptosis and thus contributes to oncogenesis. Although an increased expression of STAT1 has been observed in many human neoplasias, this molecule can be considered a potential tumor suppressor, since it plays an important role in growth arrest and in promoting apoptosis. On the other hand, STAT3 and 5 are considered as oncogenes, since they bring about the activation of cyclin D1, c-Myc, and bcl-xl expression, and are involved in promoting cell-cycle progression, cellular transformation, and in preventing apoptosis.

Phosphate toxicity: new insights into an old problem

PubMed Central

RAZZAQUE, M. Shawkat

2011-01-01

Phosphorus is an essential nutrient required for critical biological reactions that maintain the normal homoeostatic control of the cell. This element is an important component of different cellular structures, including nucleic acids and cell membranes. Adequate phosphorus balance is vital for maintaining basic cellular functions, ranging from energy metabolism to cell signalling. In addition, many intracellular pathways utilize phosphate ions for important cellular reactions; therefore, homoeostatic control of phosphate is one of the most delicate biological regulations. Impaired phosphorus balance can affect the functionality of almost every human system, including musculoskeletal and cardiovascular systems, ultimately leading to an increase in morbidity and mortality of the affected patients. Human and experimental studies have found that delicate balance among circulating factors, like vitamin D, PTH (parathyroid hormone) and FGF23 (fibroblast growth factor 23), are essential for regulation of physiological phosphate balance. Dysregulation of these factors, either alone or in combination, can induce phosphorus imbalance. Recent studies have shown that suppression of the FGF23–klotho system can lead to hyperphosphataemia with extensive tissue damage caused by phosphate toxicity. The cause and consequences of phosphate toxicity will be briefly summarized in the present review. PMID:20958267

Phosphate toxicity: new insights into an old problem.

PubMed

Razzaque, M Shawkat

2011-02-01

Phosphorus is an essential nutrient required for critical biological reactions that maintain the normal homoeostatic control of the cell. This element is an important component of different cellular structures, including nucleic acids and cell membranes. Adequate phosphorus balance is vital for maintaining basic cellular functions, ranging from energy metabolism to cell signalling. In addition, many intracellular pathways utilize phosphate ions for important cellular reactions; therefore, homoeostatic control of phosphate is one of the most delicate biological regulations. Impaired phosphorus balance can affect the functionality of almost every human system, including musculoskeletal and cardiovascular systems, ultimately leading to an increase in morbidity and mortality of the affected patients. Human and experimental studies have found that delicate balance among circulating factors, like vitamin D, PTH (parathyroid hormone) and FGF23 (fibroblast growth factor 23), are essential for regulation of physiological phosphate balance. Dysregulation of these factors, either alone or in combination, can induce phosphorus imbalance. Recent studies have shown that suppression of the FGF23-klotho system can lead to hyperphosphataemia with extensive tissue damage caused by phosphate toxicity. The cause and consequences of phosphate toxicity will be briefly summarized in the present review.

An enhanced functional interrogation/manipulation of intracellular signaling pathways with the peptide 'stapling' technology.

PubMed

He, Y; Chen, D; Zheng, W

2015-11-12

Specific protein-protein interactions (PPIs) constitute a key underlying mechanism for the presence of a multitude of intracellular signaling pathways, which are essential for the survival of normal and cancer cells. Specific molecular blockers for a crucial PPI would therefore be invaluable tools for an enhanced functional interrogation of the signaling pathway harboring this particular PPI. On the other hand, if a particular PPI is essential for the survival of cancer cells but is absent in or dispensable for the survival of normal cells, its specific molecular blockers could potentially be developed into effective anticancer therapeutics. Due to the flat and extended PPI interface, it would be conceivably difficult for small molecules to achieve an effective blockade, a problem which could be potentially circumvented with peptides or proteins. However, the well-documented proteolytic instability and cellular impermeability of peptides and proteins in general would make their developing into effective intracellular PPI blockers quite a challenge. With the advent of the peptide 'stapling' technology which was demonstrated to be able to stabilize the α-helical conformation of a peptide via bridging two neighboring amino-acid side chains with a 'molecular staple', a linear parent peptide could be transformed into a stronger PPI blocker with enhanced proteolytic stability and cellular permeability. This review will furnish an account on the peptide 'stapling' technology and its exploitation in efforts to achieve an enhanced functional interrogation or manipulation of intracellular signaling pathways especially those that are cancer relevant.

Effects of ozone in normal human epidermal keratinocytes.

PubMed

McCarthy, James T; Pelle, Edward; Dong, Kelly; Brahmbhatt, Krupa; Yarosh, Dan; Pernodet, Nadine

2013-05-01

Ozone is a tropospheric pollutant that can form at ground level as a result of an interaction between sunlight and hydrocarbon engine emissions. As ozone is an extremely oxidative reaction product, epidermal cells are in the outer layer of defense against ozone. We exposed normal human epidermal keratinocytes (NHEK) to concentrations of ozone that have been measured in cities and assayed for its effects. Hydrogen peroxide and IL-1α levels both increased while ATP levels decreased. We found a decrease in the NAD-dependent histone deacetylase, sirtuin 3. Lastly, we found that ozone increased DNA damage as evaluated by Comet assay. Taken together, our results show increased damage to NHEK that will ultimately impair normal cellular function as a result of an environmentally relevant ozone exposure. © 2013 John Wiley & Sons A/S.

Hepatocyte growth factor activator inhibitors (HAI-1 and HAI-2): Emerging key players in epithelial integrity and cancer.

PubMed

Kataoka, Hiroaki; Kawaguchi, Makiko; Fukushima, Tsuyoshi; Shimomura, Takeshi

2018-03-01

The growth, survival, and metabolic activities of multicellular organisms at the cellular level are regulated by intracellular signaling, systemic homeostasis and the pericellular microenvironment. Pericellular proteolysis has a crucial role in processing bioactive molecules in the microenvironment and thereby has profound effects on cellular functions. Hepatocyte growth factor activator inhibitor type 1 (HAI-1) and HAI-2 are type I transmembrane serine protease inhibitors expressed by most epithelial cells. They regulate the pericellular activities of circulating hepatocyte growth factor activator and cellular type II transmembrane serine proteases (TTSPs), proteases required for the activation of hepatocyte growth factor (HGF)/scatter factor (SF). Activated HGF/SF transduces pleiotropic signals through its receptor tyrosine kinase, MET (coded by the proto-oncogene MET), which are necessary for cellular migration, survival, growth and triggering stem cells for accelerated healing. HAI-1 and HAI-2 are also required for normal epithelial functions through regulation of TTSP-mediated activation of other proteases and protease-activated receptor 2, and also through suppressing excess degradation of epithelial junctional proteins. This review summarizes current knowledge regarding the mechanism of pericellular HGF/SF activation and highlights emerging roles of HAIs in epithelial development and integrity, as well as tumorigenesis and progression of transformed epithelial cells. © 2018 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.

Robust imaging and gene delivery to study human lymphoblastoid cell lines.

PubMed

Jolly, Lachlan A; Sun, Ying; Carroll, Renée; Homan, Claire C; Gecz, Jozef

2018-06-20

Lymphoblastoid cell lines (LCLs) have been by far the most prevalent cell type used to study the genetics underlying normal and disease-relevant human phenotypic variation, across personal to epidemiological scales. In contrast, only few studies have explored the use of LCLs in functional genomics and mechanistic studies. Two major reasons are technical, as (1) interrogating the sub-cellular spatial information of LCLs is challenged by their non-adherent nature, and (2) LCLs are refractory to gene transfection. Methodological details relating to techniques that overcome these limitations are scarce, largely inadequate (without additional knowledge and expertise), and optimisation has never been described. Here we compare, optimise, and convey such methods in-depth. We provide a robust method to adhere LCLs to coverslips, which maintained cellular integrity, morphology, and permitted visualisation of sub-cellular structures and protein localisation. Next, we developed the use of lentiviral-based gene delivery to LCLs. Through empirical and combinatorial testing of multiple transduction conditions, we improved transduction efficiency from 3% up to 48%. Furthermore, we established strategies to purify transduced cells, to achieve sustainable cultures containing >85% transduced cells. Collectively, our methodologies provide a vital resource that enables the use of LCLs in functional cell and molecular biology experiments. Potential applications include the characterisation of genetic variants of unknown significance, the interrogation of cellular disease pathways and mechanisms, and high-throughput discovery of genetic modifiers of disease states among others.

Fuzzy cellular automata models in immunology

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

Ahmed, E.

1996-10-01

The self-nonself character of antigens is considered to be fuzzy. The Chowdhury et al. cellular automata model is generalized accordingly. New steady states are found. The first corresponds to a below-normal help and suppression and is proposed to be related to autoimmune diseases. The second corresponds to a below-normal B-cell level.

[Computer simulation of thyroid regulatory mechanisms in health and malignancy].

PubMed

Abduvaliev, A A; Gil'dieva, M S; Khidirov, B N; Saĭdalieva, M; Saatov, T S

2010-07-01

The paper describes a computer model for regulation of the number of thyroid follicular cells in health and malignancy. The authors'computer program for mathematical simulation of the regulatory mechanisms of a thyroid follicular cellular community cannot be now referred to as good commercial products. For commercialization of this product, it is necessary to draw up a direct relation of the introduced corrected values from the actually existing normal values, such as the peripheral blood concentrations of thyroid hormones or the mean values of endocrine tissue mitotic activity. However, the described computer program has been also used in researches by our scientific group in the study of thyroid cancer. The available biological experimental data and theoretical provisions on thyroid structural and functional organization at the cellular level allow one to construct mathematical models for quantitative analysis of the regulation of the size of a cellular community of a thyroid follicle in health and abnormalities, by using the method for simulation of the regulatory mechanisms of living systems and the equations of cellular community regulatory communities.

Yeast aquaporin regulation by 4-hydroxynonenal is implicated in oxidative stress response.

PubMed

Rodrigues, Claudia; Tartaro Bujak, Ivana; Mihaljević, Branka; Soveral, Graça; Cipak Gasparovic, Ana

2017-05-01

Reactive oxygen species, especially hydrogen peroxide (H 2 O 2 ), contribute to functional molecular impairment and cellular damage, but also are necessary in normal cellular metabolism, and in low doses play stimulatory role in cell proliferation and stress resistance. In parallel, reactive aldehydes such as 4-hydroxynonenal (HNE), are lipid peroxidation breakdown products which also contribute to regulation of numerous cellular processes. Recently, channeling of H 2 O 2 by some mammalian aquaporin isoforms has been reported and suggested to contribute to aquaporin involvement in cancer malignancies, although the mechanism by which these membrane water channels are implicated in oxidative stress is not clear. In this study, two yeast models with increased levels of membrane polyunsaturated fatty acids (PUFAs) and aquaporin AQY1 overexpression, respectively, were used to evaluate their interplay in cell's oxidative status. In particular, the aim of the study was to investigate if HNE accumulation could affect aquaporin function with an outcome in oxidative stress response. The data showed that induction of aquaporin expression by PUFAs results in increased water permeability in yeast membranes and that AQY1 activity is impaired by HNE. Moreover, AQY1 expression increases cellular sensitivity to oxidative stress by facilitating H 2 O 2 influx. On the other hand, AQY1 expression has no influence on the cellular antioxidant GSH levels and catalase activity. These results strongly suggest that aquaporins are important players in oxidative stress response and could contribute to regulation of cellular processes by regulation of H 2 O 2 influx. © 2017 IUBMB Life, 69(5):355-362, 2017. © 2017 International Union of Biochemistry and Molecular Biology.

Role of cellular oxalate in oxalate clearance of patients with calcium oxalate monohydrate stone formation and normal controls.

PubMed

Oehlschläger, Sven; Fuessel, Susanne; Meye, Axel; Herrmann, Jana; Froehner, Michael; Albrecht, Steffen; Wirth, Manfred P

2009-03-01

To examine the cellular, plasma, and urinary oxalate and erythrocyte oxalate flux in patients with calcium oxalate monohydrate (COM) stone formation vs normal controls. Pathologic oxalate clearance in humans is mostly integrated in calcium oxalate stone formation. An underlying cause of deficient oxalate clearance could be defective transmembrane oxalate transport, which, in many tissues, is regulated by an anion exchanger (SLC26). We studied 2 groups: 40 normal controls and 41 patients with COM stone formation. Red blood cells were divided for cellular oxalate measurement and for resuspension in a buffered solution (pH 7.40); 0.1 mmol/L oxalate was added. The supernatant was measured for oxalate immediately and 1 hour after incubation. The plasma and urinary oxalate were analyzed in parallel. The mean cellular oxalate concentrations were significantly greater in the normal controls (5.25 +/- 0.47 micromol/L) than in those with COM stone formation (2.36 +/- 0.28 micromol/L; P < .01). The mean urinary oxalate concentrations were significantly greater in those with COM stone formation (0.31 +/- 0.02 mmol/L) than in the controls (0.24 +/- 0.02 mmol/L; P < .01). The cellular oxalate concentrations correlated significantly with the plasma (r = 0.49-0.63; P < .01) and urinary oxalate (r = -0.29-0.41; P < .03) concentrations in both groups. The plasma oxalate concentrations correlated significantly with the urinary oxalate concentrations (r = -0.30; P < .03) in the controls and with the erythrocyte oxalate flux (r = 0.25; P < .05) in those with COM stone formation. Our data implicate the presence of a cellular oxalate buffer to stabilize plasma and urinary oxalate concentrations in normal controls.

EMPOWERING ADULT STEM CELLS FOR MYOCARDIAL REGENERATION

PubMed Central

Mohsin, Sadia; Siddiqi, Sailay; Collins, Brett; Sussman, Mark A.

2012-01-01

Treatment strategies for heart failure remain a high priority for ongoing research due to the profound unmet need in clinical disease coupled with lack of significant translational progress. The underlying issue is the same whether the cause is acute damage, chronic stress from disease, or aging: progressive loss of functional cardiomyocytes and diminished hemodynamic output. To stave off cardiomyocyte losses, a number of strategic approaches have been embraced in recent years involving both molecular and cellular approaches to augment myocardial structure and performance. Resultant excitement surrounding regenerative medicine in the heart has been tempered by realizations that reparative processes in the heart are insufficient to restore damaged myocardium to normal functional capacity and that cellular cardiomyoplasty is hampered by poor survival, proliferation, engraftment and differentiation of the donated population. To overcome these limitations, a combination of molecular and cellular approaches needs to be adopted involving use of genetic engineering to enhance resistance to cell death and increase regenerative capacity. This review will highlight biological properties of approached to potentiate stem cell-mediated regeneration to promote enhanced myocardial regeneration, persistence of donated cells, and long lasting tissue repair. Optimizing cell delivery and harnessing the power of survival signaling cascades for ex vivo genetic modification of stem cells prior to reintroduction into the patient will be critical to enhance the efficacy of cellular cardiomyoplasty. Once this goal is achieved, then cell-based therapy has great promise for treatment of heart failure to combat the loss of cardiac structure and function associated with acute damage, chronic disease or aging. PMID:22158649

Safety Implications of High-Field MRI: Actuation of Endogenous Magnetic Iron Oxides in the Human Body

PubMed Central

Dobson, Jon; Bowtell, Richard; Garcia-Prieto, Ana; Pankhurst, Quentin

2009-01-01

Background Magnetic Resonance Imaging scanners have become ubiquitous in hospitals and high-field systems (greater than 3 Tesla) are becoming increasingly common. In light of recent European Union moves to limit high-field exposure for those working with MRI scanners, we have evaluated the potential for detrimental cellular effects via nanomagnetic actuation of endogenous iron oxides in the body. Methodology Theoretical models and experimental data on the composition and magnetic properties of endogenous iron oxides in human tissue were used to analyze the forces on iron oxide particles. Principal Finding and Conclusions Results show that, even at 9.4 Tesla, forces on these particles are unlikely to disrupt normal cellular function via nanomagnetic actuation. PMID:19412550

Endoreplication and polyploidy: insights into development and disease

PubMed Central

Fox, Donald T.; Duronio, Robert J.

2013-01-01

Polyploid cells have genomes that contain multiples of the typical diploid chromosome number and are found in many different organisms. Studies in a variety of animal and plant developmental systems have revealed evolutionarily conserved mechanisms that control the generation of polyploidy and have recently begun to provide clues to its physiological function. These studies demonstrate that cellular polyploidy plays important roles during normal development and also contributes to human disease, particularly cancer. PMID:23222436

Repression of anti-proliferative factor Tob1 in osteoarthritic cartilage

PubMed Central

Gebauer, Mathias; Saas, Joachim; Haag, Jochen; Dietz, Uwe; Takigawa, Masaharu; Bartnik, Eckart; Aigner, Thomas

2005-01-01

Osteoarthritis is the most common degenerative disorder of the modern world. However, many basic cellular features and molecular processes of the disease are poorly understood. In the present study we used oligonucleotide-based microarray analysis of genes of known or assumed relevance to the cellular phenotype to screen for relevant differences in gene expression between normal and osteoarthritic chondrocytes. Custom made oligonucleotide DNA arrays were used to screen for differentially expressed genes in normal (n = 9) and osteoarthritic (n = 10) cartilage samples. Real-time polymerase chain reaction (PCR) with gene-specific primers was used for quantification. Primary human adult articular chondrocytes and chondrosarcoma cell line HCS-2/8 were used to study changes in gene expression levels after stimulation with interleukin-1β and bone morphogenetic protein, as well as the dependence on cell differentiation. In situ hybridization with a gene-specific probe was applied to detect mRNA expression levels in fetal growth plate cartilage. Overall, more than 200 significantly regulated genes were detected between normal and osteoarthritic cartilage (P < 0.01). One of the significantly repressed genes, Tob1, encodes a protein belonging to a family involved in silencing cells in terms of proliferation and functional activity. The repression of Tob1 was confirmed by quantitative PCR and correlated to markers of chondrocyte activity and proliferation in vivo. Tob1 expression was also detected at a decreased level in isolated chondrocytes and in the chondrosarcoma cell line HCS-2/8. Again, in these cells it was negatively correlated with proliferative activity and positively with cellular differentiation. Altogether, the downregulation of the expression of Tob1 in osteoarthritic chondrocytes might be an important aspect of the cellular processes taking place during osteoarthritic cartilage degeneration. Activation, the reinitiation of proliferative activity and the loss of a stable phenotype are three major changes in osteoarthritic chondrocytes that are highly significantly correlated with the repression of Tob1 expression. PMID:15743474

Cellular, synaptic and biochemical features of resilient cognition in Alzheimer’s disease

PubMed Central

Arnold, Steven. E.; Louneva, Natalia; Cao, Kajia; Wang, Li-San; Han, Li-Ying; Wolk, David A.; Negash, Selamawit; Leurgans, Sue E.; Schneider, Julie A.; Buchman, Aron S.; Wilson, Robert S.; Bennett, David A.

2012-01-01

While neuritic plaques and neurofibrillary tangles in older adults are correlated with cognitive impairment and severity of dementia, it has long been recognized that the relationship is imperfect as some people exhibit normal cognition despite high levels of AD pathology. We compared the cellular, synaptic and biochemical composition of midfrontal cortices in female subjects from the Religious Orders Study who were stratified into three subgroups: 1) pathological AD with normal cognition (“AD-Resilient”), 2) pathological AD with AD-typical dementia (“AD-Dementia)” and 3) pathologically normal with normal cognition (“Normal Comparison”). The AD-Resilient group exhibited preserved densities of synaptophysin-labeled presynaptic terminals and synaptopodin-labeled dendritic spines compared to the AD-Dementia group, and increased densities of GFAP astrocytes compared to both the AD-Dementia and Normal Comparison group. Further, in a discovery antibody microarray protein analysis we identified a number of candidate protein abnormalities that were associated with diagnostic group. These data characterize cellular and synaptic features and identify novel biochemical targets that may be associated with resilient cognitive brain aging in the setting of pathological AD. PMID:22554416

Current state of cartilage tissue engineering

PubMed Central

Tuli, Richard; Li, Wan-Ju; Tuan, Rocky S

2003-01-01

Damage to cartilage is of great clinical consequence given the tissue's limited intrinsic potential for healing. Current treatments for cartilage repair are less than satisfactory, and rarely restore full function or return the tissue to its native normal state. The rapidly emerging field of tissue engineering holds great promise for the generation of functional cartilage tissue substitutes. The general approach involves a biocompatible, structurally and mechanically sound scaffold, with an appropriate cell source, which is loaded with bioactive molecules that promote cellular differentiation and/or maturation. This review highlights aspects of current progress in cartilage tissue engineering. PMID:12932283

Reduced Abd-B Hox function during kidney development results in lineage infidelity.

PubMed

Magella, Bliss; Mahoney, Robert; Adam, Mike; Potter, S Steven

2018-06-15

Hox genes can function as key drivers of segment identity, with Hox mutations in Drosophila often resulting in dramatic homeotic transformations. In addition, however, they can serve other essential functions. In mammals, the study of Hox gene roles in development is complicated by the presence of four Hox clusters with a total of 39 genes showing extensive functional overlap. In this study, in order to better understand shared core Hox functions, we examined kidney development in mice with frameshift mutations of multiple Abd-B type Hox genes. The resulting phenotypes included dramatically reduced branching morphogenesis of the ureteric bud, premature depletion of nephron progenitors and abnormal development of the stromal compartment. Most unexpected, however, we also observed a cellular level lineage infidelity in nephron segments. Scattered cells within the proximal tubules, for example, expressed genes normally expressed only in collecting ducts. Multiple combinations of inappropriate nephron segment specific marker expression were found. In some cases, cells within a tubule showed incorrect identity, while in other cases cells showed ambiguous character, with simultaneous expression of genes associated with more than one nephron segment. These results give evidence that Hox genes have an overlapping core function at the cellular level in driving and/or maintaining correct differentiation decisions. Copyright © 2018 Elsevier Inc. All rights reserved.

Human immunodeficiency virus (HIV) type 1 Vpr induces differential regulation of T cell costimulatory molecules: Direct effect of Vpr on T cell activation and immune function

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

Venkatachari, Narasimhan J.; Majumder, Biswanath; Ayyavoo, Velpandi

2007-02-20

Human immunodeficiency virus type 1 (HIV-1) viral proteins disrupt the normal host cellular immune pathways thus exploiting the cellular machinery for replication, survival and to escape host immune attack. Here we evaluated the direct effects of HIV-1 Vpr-mediated immune modulation of infected T cells. Vpr specifically downregulated the expression of CD28 and increased the expression of CTLA-4, whereas no significant difference in the expression of CD25 and HLA-DR was observed. Interferon gamma (IFN-{gamma}) production in T cells was evaluated as a measure of the downstream effector functions. Results indicate that Vpr significantly inhibited IFN-{gamma} production and this may, in part,more » due to Vpr's ability to inhibit the nuclear translocation of NF-{kappa}B, and its transcriptional regulation. Together these results support that HIV-1 Vpr selectively dysregulates the immune functions at multiple levels and exerts its inhibitory effects in the presence of other viral proteins.« less

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

Influence of local anesthetics upon human polymorphonuclear leukocyte function in vitro. Reduction of lysosomal enzyme release and superoxide anion production

PubMed Central

1977-01-01

Cationic local anesthetics have been reported to influence cellular responses to surface stimuli by interfering with the function of microtubules and microfilaments. Since unimpaired microtubule and microfilament functions are required by human polymorphonuclear leukocytes in order to respond normally to surface stimulation, we have studied effects of the local anesthetic, tetracaine on the function and morphology of these cells in vitro. Tetracaine (0.25--1.0 mM) significantly reduced extracellular release of the lysosomal enzymes, beta-glucuronidase and lysozyme from polymorphonuclear leukocytes exposed to serum-treated zymosan (a particulate stimulus), zymosan- treated serum (a soluble stimulus), and to the surface-active lectin, concanavalin A. Tetracaine also significantly reduced superoixde anion production (superoxide dismutase-inhibitable cytochrome c reduction) by these cells. Tetrancaine was not cytotoxic and its effects could be reversed completely by washing cells once with buffer. Electron microscope examination of tetracaine-treated cells revealed marked alterations of surface membranes. Microtubules and microfilaments appeared normal in "resting" polymorphonuclear leukocytes, but the increase in microtubules normally observed in stimulated cells was not seen after tetracaine treatment. These results suggest that tetracaine interferes with those interactions between immune reactants and the polymorphonuclear leukocyte cell surface which provoke exocytosis and increased oxidative metabolism. PMID:195003

Rap G protein signal in normal and disordered lymphohematopoiesis

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

Minato, Nagahiro, E-mail: minato@imm.med.kyoto-u.ac.jp

2013-09-10

Rap proteins (Rap1, Rap2a, b, c) are small molecular weight GTPases of the Ras family. Rap G proteins mediate diverse cellular events such as cell adhesion, proliferation, and gene activation through various signaling pathways. Activation of Rap signal is regulated tightly by several specific regulatory proteins including guanine nucleotide exchange factors and GTPase-activating proteins. Beyond cell biological studies, increasing attempts have been made in the past decade to define the roles of Rap signal in specific functions of normal tissue systems as well as in cancer. In the immune and hematopoietic systems, Rap signal plays crucial roles in the developmentmore » and function of essentially all lineages of lymphocytes and hematopoietic cells, and importantly, deregulated Rap signal may lead to unique pathological conditions depending on the affected cell types, including various types of leukemia and autoimmunity. The phenotypical studies have unveiled novel, even unexpected functional aspects of Rap signal in cells from a variety of tissues, providing potentially important clues for controlling human diseases, including malignancy.« less

Synthetic biology meets tissue engineering.

PubMed

Davies, Jamie A; Cachat, Elise

2016-06-15

Classical tissue engineering is aimed mainly at producing anatomically and physiologically realistic replacements for normal human tissues. It is done either by encouraging cellular colonization of manufactured matrices or cellular recolonization of decellularized natural extracellular matrices from donor organs, or by allowing cells to self-organize into organs as they do during fetal life. For repair of normal bodies, this will be adequate but there are reasons for making unusual, non-evolved tissues (repair of unusual bodies, interface to electromechanical prostheses, incorporating living cells into life-support machines). Synthetic biology is aimed mainly at engineering cells so that they can perform custom functions: applying synthetic biological approaches to tissue engineering may be one way of engineering custom structures. In this article, we outline the 'embryological cycle' of patterning, differentiation and morphogenesis and review progress that has been made in constructing synthetic biological systems to reproduce these processes in new ways. The state-of-the-art remains a long way from making truly synthetic tissues, but there are now at least foundations for future work. © 2016 Authors; published by Portland Press Limited.

Structure of the vault, a ubiquitous celular component.

PubMed

Kong, L B; Siva, A C; Rome, L H; Stewart, P L

1999-04-15

The vault is a ubiquitous and highly conserved ribonucleoprotein particle of approximately 13 MDa. This particle has been shown to be upregulated in certain multidrug-resistant cancer cell lines and to share a protein component with the telomerase complex. Determination of the structure of the vault was undertaken to provide a first step towards understanding the role of this cellular component in normal metabolism and perhaps to shed some light on its role in mediating drug resistance. Over 1300 particle images were combined to calculate an approximately 31 A resolution structure of the vault. Rotational power spectra did not yield a clear symmetry peak, either because of the thin, smooth walls or inherent flexibility of the vault. Although cyclic eightfold (C8) symmetry was imposed, the resulting reconstruction may be partially cylindrically averaged about the eightfold axis. Our results reveal the vault to be a hollow, barrel-like structure with two protruding caps and an invaginated waist. Although the normal cellular function of the vault is as yet undetermined, the structure of the vault is consistent with either a role in subcellular transport, as previously suggested, or in sequestering macromolecular assemblies.

An extended cost potential field cellular automata model considering behavior variation of pedestrian flow

NASA Astrophysics Data System (ADS)

Guo, Fang; Li, Xingli; Kuang, Hua; Bai, Yang; Zhou, Huaguo

2016-11-01

The original cost potential field cellular automata describing normal pedestrian evacuation is extended to study more general evacuation scenarios. Based on the cost potential field function, through considering the psychological characteristics of crowd under emergencies, the quantitative formula of behavior variation is introduced to reflect behavioral changes caused by psychology tension. The numerical simulations are performed to investigate the effects of the magnitude of behavior variation, the different pedestrian proportions with different behavior variation and other factors on the evacuation efficiency and process in a room. The spatiotemporal dynamic characteristic during the evacuation process is also discussed. The results show that compared with the normal evacuation, the behavior variation under an emergency does not necessarily lead to the decrease of the evacuation efficiency. At low density, the increase of the behavior variation can improve the evacuation efficiency, while at high density, the evacuation efficiency drops significantly with the increasing amplitude of the behavior variation. In addition, the larger proportion of pedestrian affected by the behavior variation will prolong the evacuation time.

Regulated production of free radicals by the mitochondrial electron transport chain: Cardiac ischemic preconditioning.

PubMed

Matsuzaki, Satoshi; Szweda, Pamela A; Szweda, Luke I; Humphries, Kenneth M

2009-11-30

Excessive production of free radicals by mitochondria is associated with, and likely contributes to, the progression of numerous pathological conditions. Nevertheless, the production of free radicals by the mitochondria may have important biological functions under normal or stressed conditions by activating or modulating redox-sensitive cellular signaling pathways. This raises the intriguing possibility that regulated mitochondrial free radical production occurs via mechanisms that are distinct from pathologies associated with oxidative damage. Indeed, the capacity of mitochondria to produce free radicals in a limited manner may play a role in ischemic preconditioning, the phenomenon whereby short bouts of ischemia protect from subsequent prolonged ischemia and reperfusion. Ischemic preconditioning can thus serve as an important model system for defining regulatory mechanisms that allow for transient, signal-inducing, production of free radicals by mitochondria. Defining how these mechanism(s) occur will provide insight into therapeutic approaches that minimize oxidative damage without altering normal cellular redox biology. The aim of this review is to present and discuss evidence for the regulated production of superoxide by the electron transport chain within the ischemic preconditioning paradigm of redox regulation.

Natural killing and antibody-dependent cellular cytotoxicity are independent immune functions in the Minnesota miniature swine.

PubMed

Koren, H S; Amos, D B; Kim, Y B

1978-10-01

Peripheral blood lymphocytes from Minnesota miniature pigs were tested for natural killing (NK) and antibody-dependent cellular cytotoxicity (ADCC) in a 2- to 4-hr 51Cr release assay against human myeloid and lymphoid tumor target cells. Adult specific pathogen-free and germfree animals exhibited normal levels of activity in both assays. In addition, the NK and ADCC activities of peripheral blood lymphocytes from colostrum-deprived newborn piglets were examined. These animals were obtained by hysterectomy and previously shown to be immunologically "virgin." We found that these newborn piglets exhibited normal ADCC but lacked NK activity. The differences in the ontogeny of the two activities suggest that they are distinct. Preliminary effector cell characterization studies suggest that: (i) NK and ADCC in the pig are physically not separable; (ii) the majority of the cytotoxic activity on a cell-per-cell basis is mediated by the non-T lymphocyte fraction; and (iii) the rosetted T cells, which account for about 60% of the total pig peripheral blood lymphocytes, have low but demonstrable cytotoxic activity as well.

Dysfunction of protein kinase FA/GSK-3 alpha in lymphocytes of patients with schizophrenic disorder.

PubMed

Yang, S D; Yu, J S; Lee, T T; Yang, C C; Ni, M H; Yang, Y Y

1995-09-01

As compared to normal people, the lymphocytes of patients with schizophrenia were found to have an impairment of ATP.Mg-dependent protein phosphatase activation. More importantly, the impaired protein phosphatase activation in the lymphocytes of schizophrenic patients could be consistently and completely restored to normal by exogenous pure protein kinase FA/glycogen synthase kinase-3 alpha (kinase FA/GSK-3 alpha) (the activating factor of ATP.Mg-dependent protein phosphatase), indicating that the molecular mechanism for the impaired protein phosphatase activation in schizophrenic patients may be due to a functional loss of kinase FA/GSK-3 alpha. Immunoblotting and kinase activity analysis in an anti-kinase FA/GSK-3 alpha immunoprecipitate further demonstrate that both cellular activities and protein levels of kinase FA/GSK-3 alpha in the lymphocytes of schizophrenic patients were greatly impared as compared to normal controls. Statistical analysis revealed that the lymphocytes isolated from 37 normal people contain kinase FA/GSK-3 alpha activity in the high levels of 14.8 +/- 2.4 units/mg of cell protein, whereas the lymphocytes of 48 patients with schizophrenic disorder contain kinase FA/GSK-3 alpha activity in the low levels of 2.8 +/- 1.6 units/mg, indicating that the different levels of kinase FA/GSK-3 alpha activity between schizophrenic patients and normal people are statistically significant. Taken together, the results provide initial evidence that patients with schizophrenic disorder may have a common impairment in the protein levels and cellular activities of kinase FA/GSK-3 alpha, a multisubstrate protein kinase and a multisubstrate protein phosphatase activator in their lymphocytes.

Differential expression of lymphocyte function-associated antigen (LFA-1) on peripheral blood leucocytes from individuals with Down's syndrome.

PubMed Central

Barrena, M J; Echaniz, P; Garcia-Serrano, C; Zubillaga, P; Cuadrado, E

1992-01-01

We analysed the expression of lymphocyte function-associated antigen LFA-1 on the cell surface of peripheral blood lymphocytes, monocytes and granulocytes from 20 children with Down's syndrome. No differences in LFA-1 expression was found within monocytes or granulocytes from either normal or Down's syndrome children; however, a clear-cut difference was observed on lymphoid cells. Both normal and Down's syndrome lymphocytes displayed a bimodal pattern of LFA-1 staining by flow cytometry, with a predominance of cells with low expression in normal population, and an increased proportion of lymphocytes with high level of LFA-1 expression in Down's syndrome children. This difference correlates well with the abnormal proportion of T cell subsets and inversion of CD4/CD8 observed in a majority of our cases, and therefore, it could merely reflect the increase of certain T cell subsets normally expressing higher number of LFA-1 molecules. Taken together, our results do not support an abnormally increased expression of leucocytes integrins in trisomy 21 cells, and raise some doubt about the suggested role of the abnormal cellular expression of LFA-1 in the pathogensis of secondary immunodeficiency associated to Down's syndrome. PMID:1348667

Keeping It All Going-Complement Meets Metabolism.

PubMed

Kolev, Martin; Kemper, Claudia

2017-01-01

The complement system is an evolutionary old and crucial component of innate immunity, which is key to the detection and removal of invading pathogens. It was initially discovered as a liver-derived sentinel system circulating in serum, the lymph, and interstitial fluids that mediate the opsonization and lytic killing of bacteria, fungi, and viruses and the initiation of the general inflammatory responses. Although work performed specifically in the last five decades identified complement also as a critical instructor of adaptive immunity-indicating that complement's function is likely broader than initially anticipated-the dominant opinion among researchers and clinicians was that the key complement functions were in principle defined. However, there is now a growing realization that complement activity goes well beyond "classic" immune functions and that this system is also required for normal (neuronal) development and activity and general cell and tissue integrity and homeostasis. Furthermore, the recent discovery that complement activation is not confined to the extracellular space but occurs within cells led to the surprising understanding that complement is involved in the regulation of basic processes of the cell, particularly those of metabolic nature-mostly via novel crosstalks between complement and intracellular sensor, and effector, pathways that had been overlooked because of their spatial separation. These paradigm shifts in the field led to a renaissance in complement research and provide new platforms to now better understand the molecular pathways underlying the wide-reaching effects of complement functions in immunity and beyond. In this review, we will cover the current knowledge about complement's emerging relationship with the cellular metabolism machinery with a focus on the functional differences between serum-circulating versus intracellularly active complement during normal cell survival and induction of effector functions. We will also discuss how taking a closer look into the evolution of key complement components not only made the functional connection between complement and metabolism rather "predictable" but how it may also give clues for the discovery of additional roles for complement in basic cellular processes.

MET Receptor Tyrosine Kinase as an Autism Genetic Risk Factor

PubMed Central

Peng, Yun; Huentelman, Matthew; Smith, Christopher; Qiu, Shenfeng

2014-01-01

In this chapter, we will briefly discuss recent literature on the role of MET receptor tyrosine kinase (RTK) in brain development and how perturbation of MET signaling may alter normal neurodevelopmental outcomes. Recent human genetic studies have established MET as a risk factor for autism, and the molecular and cellular underpinnings of this genetic risk are only beginning to emerge from obscurity. Unlike many autism risk genes that encode synaptic proteins, the spatial and temporal expression pattern of MET RTK indicates this signaling system is ideally situated to regulate neuronal growth, functional maturation, and establishment of functional brain circuits, particularly in those brain structures involved in higher levels of cognition, social skills, and executive functions. PMID:24290385

Sox proteins in melanocyte development and melanoma

PubMed Central

Harris, Melissa L.; Baxter, Laura L.; Loftus, Stacie K.; Pavan, William J.

2010-01-01

Over ten years has passed since the first Sox gene was implicated in melanocyte development. Since then, we have discovered that SOX5, SOX9, SOX10 and SOX18 all participate as transcription factors that affect key melanocytic genes in both regulatory and modulatory fashions. Both SOX9 and SOX10 play major roles in the establishment and normal function of the melanocyte; SOX10 has been shown to heavily influence melanocyte development and SOX9 has been implicated in melanogenesis in the adult. Despite these advances, the precise cellular and molecular details of how these SOX proteins are regulated and interact during all stages of the melanocyte life cycle remain unknown. Improper regulation of SOX9 or SOX10 is also associated with cancerous transformation, and thus understanding the normal function of SOX proteins in the melanocyte will be key to revealing how these proteins contribute to melanoma. PMID:20444197

MECHANISMS IN ENDOCRINOLOGY: Nutrition as a mediator of oxidative stress in metabolic and reproductive disorders in women.

PubMed

Diamanti-Kandarakis, Evanthia; Papalou, Olga; Kandaraki, Eleni A; Kassi, Georgia

2017-02-01

Nutrition can generate oxidative stress and trigger a cascade of molecular events that can disrupt oxidative and hormonal balance. Nutrient ingestion promotes a major inflammatory and oxidative response at the cellular level in the postprandial state, altering the metabolic state of tissues. A domino of unfavorable metabolic changes is orchestrated in the main metabolic organs, including adipose tissue, skeletal muscle, liver and pancreas, where subclinical inflammation, endothelial dysfunction, mitochondrial deregulation and impaired insulin response and secretion take place. Simultaneously, in reproductive tissues, nutrition-induced oxidative stress can potentially violate delicate oxidative balance that is mandatory to secure normal reproductive function. Taken all the above into account, nutrition and its accompanying postprandial oxidative stress, in the unique context of female hormonal background, can potentially compromise normal metabolic and reproductive functions in women and may act as an active mediator of various metabolic and reproductive disorders. © 2017 European Society of Endocrinology.

Altered RECQL5 expression in urothelial bladder carcinoma increases cellular proliferation and makes RECQL5 helicase activity a novel target for chemotherapy

PubMed Central

Patterson, Karl; Arya, Lovleen; Bottomley, Sarah; Morgan, Susan; Cox, Angela; Catto, James; Bryant, Helen E.

2016-01-01

RECQ helicases are a family of enzymes with both over lapping and unique functions. Functional autosomal recessive loss of three members of the family BLM, WRN and RECQL4, results in hereditary human syndromes characterized by cancer predisposition and premature aging, but despite the finding that RECQL5 deficient mice are cancer prone, no such link has been made to human RECQL5. Here we demonstrate that human urothelial carcinoma of the bladder (UCC) has increased expression of RECQL5 compared to normal bladder tissue and that increasing RECQL5 expression can drive proliferation of normal bladder cells and is associated with poor prognosis. Further, by expressing a helicase dead RECQL5 and by depleting bladder cancer cells of RECQL5 we show that inhibition of RECQL5 activity has potential as a new target for treatment of UCC. PMID:27764811

Impaired eye-blink conditioning in waggler, a mutant mouse with cerebellar BDNF deficiency.

PubMed

Bao, S; Chen, L; Qiao, X; Knusel, B; Thompson, R F

1998-01-01

In addition to their trophic functions, neurotrophins are also implicated in synaptic modulation and learning and memory. Although gene knockout techniques have been used widely in studying the roles of neurotrophins at molecular and cellular levels, behavioral studies using neurotrophin knockouts are limited by the early-onset lethality and various sensory deficits associated with the gene knockout mice. In the present study, we found that in a spontaneous mutant mouse, waggler, the expression of brain-derived neurotrophic factor (BDNF) was selectively absent in the cerebellar granule cells. The cytoarchitecture of the waggler cerebellum appeared to be normal at the light microscope level. The mutant mice exhibited no sensory deficits to auditory stimuli or heat-induced pain. However, they were massively impaired in classic eye-blink conditioning. These results suggest that BDNF may have a role in normal cerebellar neuronal function, which, in turn, is essential for classic eye-blink conditioning.

Dissecting social cell biology and tumors using Drosophila genetics.

PubMed

Pastor-Pareja, José Carlos; Xu, Tian

2013-01-01

Cancer was seen for a long time as a strictly cell-autonomous process in which oncogenes and tumor-suppressor mutations drive clonal cell expansions. Research in the past decade, however, paints a more integrative picture of communication and interplay between neighboring cells in tissues. It is increasingly clear as well that tumors, far from being homogenous lumps of cells, consist of different cell types that function together as complex tissue-level communities. The repertoire of interactive cell behaviors and the quantity of cellular players involved call for a social cell biology that investigates these interactions. Research into this social cell biology is critical for understanding development of normal and tumoral tissues. Such complex social cell biology interactions can be parsed in Drosophila. Techniques in Drosophila for analysis of gene function and clonal behavior allow us to generate tumors and dissect their complex interactive biology with cellular resolution. Here, we review recent Drosophila research aimed at understanding tissue-level biology and social cell interactions in tumors, highlighting the principles these studies reveal.

The Esophagiome: concept, status, and future perspectives.

PubMed

Gregersen, Hans; Liao, Donghua; Brasseur, James G

2016-09-01

The term "Esophagiome" is meant to imply a holistic, multiscale treatment of esophageal function from cellular and muscle physiology to the mechanical responses that transport and mix fluid contents. The development and application of multiscale mathematical models of esophageal function are central to the Esophagiome concept. These model elements underlie the development of a "virtual esophagus" modeling framework to characterize and analyze function and disease by quantitatively contrasting normal and pathophysiological function. Functional models incorporate anatomical details with sensory-motor properties and functional responses, especially related to biomechanical functions, such as bolus transport and gastrointestinal fluid mixing. This brief review provides insight into Esophagiome research. Future advanced models can provide predictive evaluations of the therapeutic consequences of surgical and endoscopic treatments and will aim to facilitate clinical diagnostics and treatment. © 2016 New York Academy of Sciences.

Changes in leucocyte migration after renal transplantation

PubMed Central

Smith, M. G. M.; Eddleston, A. L. W. F.; Dominguez, J. A.; Evans, D. B.; Bewick, M.; Williams, Roger

1969-01-01

The leucocyte migration test, an in-vitro measure of cellular immunity, has been used to follow the changes in cell-mediated hypersensitivity to kidney and histocompatibility antigens in three patients after renal transplantation. Inhibition of leucocyte migration, indicating strong sensitization to the antigens used, occurred in each patient, starting five to seven days after transplantation. Satisfactory renal function had not been established in any of the patients at this time. In one case inhibition of leucocyte migration persisted almost continuously until the 24th day and was associated with poor renal function proved histologically to be due to rejection. Treatment with increased dosage of prednisone was associated with a rapid reversion to normal of the migration index and improvement in renal function. Later, inhibition of migration occurred again, and shortly afterwards the graft ceased to function. In the other two cases the migration index became normal without alteration in immunosuppressive therapy and a satisfactory diuresis followed. It is suggested that this simple test should prove useful in the specific diagnosis of rejection and in control of immunosuppressive therapy. ImagesFig. 3Fig. 4 PMID:4899455

A mitochondrial-focused genetic interaction map reveals a scaffold-like complex required for inner membrane organization in mitochondria

PubMed Central

Hoppins, Suzanne; Collins, Sean R.; Cassidy-Stone, Ann; Hummel, Eric; DeVay, Rachel M.; Lackner, Laura L.; Westermann, Benedikt; Schuldiner, Maya

2011-01-01

To broadly explore mitochondrial structure and function as well as the communication of mitochondria with other cellular pathways, we constructed a quantitative, high-density genetic interaction map (the MITO-MAP) in Saccharomyces cerevisiae. The MITO-MAP provides a comprehensive view of mitochondrial function including insights into the activity of uncharacterized mitochondrial proteins and the functional connection between mitochondria and the ER. The MITO-MAP also reveals a large inner membrane–associated complex, which we term MitOS for mitochondrial organizing structure, comprised of Fcj1/Mitofilin, a conserved inner membrane protein, and five additional components. MitOS physically and functionally interacts with both outer and inner membrane components and localizes to extended structures that wrap around the inner membrane. We show that MitOS acts in concert with ATP synthase dimers to organize the inner membrane and promote normal mitochondrial morphology. We propose that MitOS acts as a conserved mitochondrial skeletal structure that differentiates regions of the inner membrane to establish the normal internal architecture of mitochondria. PMID:21987634

Continuous exposure to low amplitude extremely low frequency electrical fields characterizing the vascular streaming potential alters elastin accumulation in vascular smooth muscle cells.

PubMed

Bergethon, Peter R; Kindler, Dean D; Hallock, Kevin; Blease, Susan; Toselli, Paul

2013-07-01

In normal development and pathology, the vascular system depends on complex interactions between cellular elements, biochemical molecules, and physical forces. The electrokinetic vascular streaming potential (EVSP) is an endogenous extremely low frequency (ELF) electrical field resulting from blood flowing past the vessel wall. While generally unrecognized, it is a ubiquitous electrical biophysical force to which the vascular tree is exposed. Extracellular matrix elastin plays a central role in normal blood vessel function and in the development of atherosclerosis. It was hypothesized that ELF fields of low amplitude would alter elastin accumulation, supporting a link between the EVSP and the biology of vascular smooth muscle cells. Neonatal rat aortic smooth muscle cell cultures were exposed chronically to electrical fields characteristic of the EVSP. Extracellular protein accumulation, DNA content, and electron microscopic (EM) evaluation were performed after 2 weeks of exposure. Stimulated cultures showed no significant change in cellular proliferation as measured by the DNA concentration. The per-DNA normalized protein in the extracellular matrix was unchanged while extracellular elastin accumulation decreased 38% on average. EM analysis showed that the stimulated cells had a 2.85-fold increase in mitochondrial number. These results support the formulation that ELF fields are a potential factor in both normal vessel biology and in the pathogenesis of atherosclerotic diseases including heart disease, stroke, and peripheral vascular disease. Copyright © 2013 Wiley Periodicals, Inc.

Study of MicroRNAs Related to the Liver Regeneration of the Whitespotted Bamboo Shark, Chiloscyllium plagiosum

PubMed Central

Lu, Conger; Nie, Zuoming; Chen, Jian; Zhang, Wenping; Ren, Xiaoyuan; Yu, Wei; Liu, Lili; Jiang, Caiying; Zhang, Yaozhou; Guo, Jiangfeng; Wu, Wutong; Shu, Jianhong; Lv, Zhengbing

2013-01-01

To understand the mechanisms of liver regeneration better to promote research examining liver diseases and marine biology, normal and regenerative liver tissues of Chiloscyllium plagiosum were harvested 0 h and 24 h after partial hepatectomy (PH) and used to isolate small RNAs for miRNA sequencing. In total, 91 known miRNAs and 166 putative candidate (PC) miRNAs were identified for the first time in Chiloscyllium plagiosum. Through target prediction and GO analysis, 46 of 91 known miRNAs were screened specially for cellular proliferation and growth. Differential expression levels of three miRNAs (xtr-miR-125b, fru-miR-204, and hsa-miR-142-3p_R-1) related to cellular proliferation and apoptosis were measured in normal and regenerating liver tissues at 0 h, 6 h, 12 h, and 24 h using real-time PCR. The expression of these miRNAs showed a rising trend in regenerative liver tissues at 6 h and 12 h but exhibited a downward trend compared to normal levels at 24 h. Differentially expressed genes were screened in normal and regenerating liver tissues at 24 h by DDRT-PCR, and ten sequences were identified. This study provided information regarding the function of genes related to liver regeneration, deepened the understanding of mechanisms of liver regeneration, and resulted in the addition of a significant number of novel miRNAs sequences to GenBank. PMID:24151623

Ribosome Synthesis and MAPK Activity Modulate Ionizing Radiation-Induced Germ Cell Apoptosis in Caenorhabditis elegans

PubMed Central

Eberhard, Ralf; Stergiou, Lilli; Hofmann, E. Randal; Hofmann, Jen; Haenni, Simon; Teo, Youjin; Furger, André; Hengartner, Michael O.

2013-01-01

Synthesis of ribosomal RNA by RNA polymerase I (RNA pol I) is an elemental biological process and is key for cellular homeostasis. In a forward genetic screen in C. elegans designed to identify DNA damage-response factors, we isolated a point mutation of RNA pol I, rpoa-2(op259), that leads to altered rRNA synthesis and a concomitant resistance to ionizing radiation (IR)-induced germ cell apoptosis. This weak apoptotic IR response could be phenocopied when interfering with other factors of ribosome synthesis. Surprisingly, despite their resistance to DNA damage, rpoa-2(op259) mutants present a normal CEP-1/p53 response to IR and increased basal CEP-1 activity under normal growth conditions. In parallel, rpoa-2(op259) leads to reduced Ras/MAPK pathway activity, which is required for germ cell progression and physiological germ cell death. Ras/MAPK gain-of-function conditions could rescue the IR response defect in rpoa-2(op259), pointing to a function for Ras/MAPK in modulating DNA damage-induced apoptosis downstream of CEP-1. Our data demonstrate that a single point mutation in an RNA pol I subunit can interfere with multiple key signalling pathways. Ribosome synthesis and growth-factor signalling are perturbed in many cancer cells; such an interplay between basic cellular processes and signalling might be critical for how tumours evolve or respond to treatment. PMID:24278030

Molecular basis for photoreceptor outer segment architecture

PubMed Central

Goldberg, Andrew F. X.; Moritz, Orson L.; Williams, David S.

2016-01-01

To serve vision, vertebrate rod and cone photoreceptors must detect photons, convert the light stimuli into cellular signals, and then convey the encoded information to downstream neurons. Rods and cones are sensory neurons that each rely on specialized ciliary organelles to detect light. These organelles, called outer segments, possess elaborate architectures that include many hundreds of light-sensitive membranous disks arrayed one atop another in precise register. These stacked disks capture light and initiate the chain of molecular and cellular events that underlie normal vision. Outer segment organization is challenged by an inherently dynamic nature; these organelles are subject to a renewal process that replaces a significant fraction of their disks (up to ~10%) on a daily basis. In addition, a broad range of environmental and genetic insults can disrupt outer segment morphology to impair photoreceptor function and viability. In this chapter, we survey the major progress that has been made for understanding the molecular basis of outer segment architecture. We also discuss key aspects of organelle lipid and protein composition, and highlight distributions, interactions, and potential structural functions of key OS-resident molecules, including: kinesin-2, actin, RP1, prominin-1, protocadherin 21, peripherin-2/rds, rom-1, glutamic acid-rich proteins, and rhodopsin. Finally, we identify key knowledge gaps and challenges that remain for understanding how normal outer segment architecture is established and maintained. PMID:27260426

The relationship between in vitro cellular aging and in vivo human age.

PubMed Central

Schneider, E L; Mitsui, Y

1976-01-01

Differences between early and late passage cell cultures on the organelle and macromolecular levels have been attributed to cellular "aging". However, concern has been expressed over whether changes in diploid cell populations after serial passage in vitro accurately reflect human cellular aging in vivo. Studies were therefore undertaken to determine if significant differences would be observed in the in vitro lifespans of skin fibroblast cultures from old and young normal, non-hospitalized volunteers and to examine if parameters that change with in vitro "aging" are altered as a function of age in vivo. Statistically signigificant (P less than 0.05) decreases were found in the rate of fibroblast migration, onset of cell culture senescence, in vitro lifespan, cell population replication rate, and cell number at confluency of fibroblast cultures derived from the old donor group when compared to parallel cultures from young donors. No significant differences were observed in modal cell volumes and cellular macromolecular contents. The differences observed in cell cultures from old and young donors were quantitatively and qualitatively distinct from those cellular alterations observed in early and late passage WI-38 cells (in vitro "aging"). Therefore, although early and late passage cultures of human diploid cells may provide an important cell system for examining loss of replicative potential, fibroblast cultures derived from old and young human donors may be a more appropriate model system for studying human cellular aging. PMID:1068470

Disruptive environmental chemicals and cellular mechanisms that confer resistance to cell death

PubMed Central

Narayanan, Kannan Badri; Ali, Manaf; Barclay, Barry J.; Cheng, Qiang (Shawn); D’Abronzo, Leandro; Dornetshuber-Fleiss, Rita; Ghosh, Paramita M.; Gonzalez Guzman, Michael J.; Lee, Tae-Jin; Leung, Po Sing; Li, Lin; Luanpitpong, Suidjit; Ratovitski, Edward; Rojanasakul, Yon; Romano, Maria Fiammetta; Romano, Simona; Sinha, Ranjeet K.; Yedjou, Clement; Al-Mulla, Fahd; Al-Temaimi, Rabeah; Amedei, Amedeo; Brown, Dustin G.; Ryan, Elizabeth P.; Colacci, Anna Maria; Hamid, Roslida A.; Mondello, Chiara; Raju, Jayadev; Salem, Hosni K.; Woodrick, Jordan; Scovassi, A.Ivana; Singh, Neetu; Vaccari, Monica; Roy, Rabindra; Forte, Stefano; Memeo, Lorenzo; Kim, Seo Yun; Bisson, William H.; Lowe, Leroy; Park, Hyun Ho

2015-01-01

Cell death is a process of dying within biological cells that are ceasing to function. This process is essential in regulating organism development, tissue homeostasis, and to eliminate cells in the body that are irreparably damaged. In general, dysfunction in normal cellular death is tightly linked to cancer progression. Specifically, the up-regulation of pro-survival factors, including oncogenic factors and antiapoptotic signaling pathways, and the down-regulation of pro-apoptotic factors, including tumor suppressive factors, confers resistance to cell death in tumor cells, which supports the emergence of a fully immortalized cellular phenotype. This review considers the potential relevance of ubiquitous environmental chemical exposures that have been shown to disrupt key pathways and mechanisms associated with this sort of dysfunction. Specifically, bisphenol A, chlorothalonil, dibutyl phthalate, dichlorvos, lindane, linuron, methoxychlor and oxyfluorfen are discussed as prototypical chemical disruptors; as their effects relate to resistance to cell death, as constituents within environmental mixtures and as potential contributors to environmental carcinogenesis. PMID:26106145

NAD+ cellular redox and SIRT1 regulate the diurnal rhythms of tyrosine hydroxylase and conditioned cocaine reward.

PubMed

Logan, Ryan W; Parekh, Puja K; Kaplan, Gabrielle N; Becker-Krail, Darius D; Williams, Wilbur P; Yamaguchi, Shintaro; Yoshino, Jun; Shelton, Micah A; Zhu, Xiyu; Zhang, Hui; Waplinger, Spencer; Fitzgerald, Ethan; Oliver-Smith, Jeffrey; Sundarvelu, Poornima; Enwright, John F; Huang, Yanhua H; McClung, Colleen A

2018-05-04

The diurnal regulation of dopamine is important for normal physiology and diseases such as addiction. Here we find a novel role for the CLOCK protein to antagonize CREB-mediated transcriptional activity at the tyrosine hydroxylase (TH) promoter, which is mediated by the interaction with the metabolic sensing protein, Sirtuin 1 (SIRT1). Additionally, we demonstrate that the transcriptional activity of TH is modulated by the cellular redox state, and daily rhythms of redox balance in the ventral tegmental area (VTA), along with TH transcription, are highly disrupted following chronic cocaine administration. Furthermore, CLOCK and SIRT1 are important for regulating cocaine reward and dopaminergic (DAergic) activity, with interesting differences depending on whether DAergic activity is in a heightened state and if there is a functional CLOCK protein. Taken together, we find that rhythms in cellular metabolism and circadian proteins work together to regulate dopamine synthesis and the reward value for drugs of abuse.

Proteomics-level analysis of myelin formation and regeneration in a mouse model for Vanishing White Matter disease.

PubMed

Gat-Viks, Irit; Geiger, Tamar; Barbi, Mali; Raini, Gali; Elroy-Stein, Orna

2015-08-01

Vanishing white matter (VWM) is a recessive neurodegenerative disease caused by mutations in translation initiation factor eIF2B and leading to progressive brain myelin deterioration, secondary axonal damage, and death in early adolescence. Eif2b5(R132H/R132H) mice exhibit delayed developmental myelination, mild early neurodegeneration and a robust remyelination defect in response to cuprizone-induced demyelination. In the current study we used Eif2b5(R132H/R132H) mice for mass-spectrometry analyses, to follow the changes in brain protein abundance in normal- versus cuprizone-diet fed mice during the remyelination recovery phase. Analysis of proteome profiles suggested that dysregulation of mitochondrial functions, altered proteasomal activity and impaired balance between protein synthesis and degradation play a role in VWM pathology. Consistent with these findings, we detected elevated levels of reactive oxygen species in mutant-derived primary fibroblasts and reduced 20S proteasome activity in mutant brain homogenates. These observations highlight the importance of tight translational control to precise coordination of processes involved in myelin formation and regeneration and point at cellular functions that may contribute to VWM pathology. Eif2b5(R132H/R132H) mouse model for vanishing white matter (VWM) disease was used for mass spectrometry of brain proteins at two time points under normal conditions and along recovery from cuprizone-induced demyelination. Comparisons of proteome profiles revealed the importance of mitochondrial function and tight coordination between protein synthesis and degradation to myelination formation and regeneration, pointing at cellular functions that contribute to VWM pathology. © 2015 International Society for Neurochemistry.

Russell body inducing threshold depends on the variable domain sequences of individual human IgG clones and the cellular protein homeostasis.

PubMed

Stoops, Janelle; Byrd, Samantha; Hasegawa, Haruki

2012-10-01

Russell bodies are intracellular aggregates of immunoglobulins. Although the mechanism of Russell body biogenesis has been extensively studied by using truncated mutant heavy chains, the importance of the variable domain sequences in this process and in immunoglobulin biosynthesis remains largely unknown. Using a panel of structurally and functionally normal human immunoglobulin Gs, we show that individual immunoglobulin G clones possess distinctive Russell body inducing propensities that can surface differently under normal and abnormal cellular conditions. Russell body inducing predisposition unique to each immunoglobulin G clone was corroborated by the intrinsic physicochemical properties encoded in the heavy chain variable domain/light chain variable domain sequence combinations that define each immunoglobulin G clone. While the sequence based intrinsic factors predispose certain immunoglobulin G clones to be more prone to induce Russell bodies, extrinsic factors such as stressful cell culture conditions also play roles in unmasking Russell body propensity from immunoglobulin G clones that are normally refractory to developing Russell bodies. By taking advantage of heterologous expression systems, we dissected the roles of individual subunit chains in Russell body formation and examined the effect of non-cognate subunit chain pair co-expression on Russell body forming propensity. The results suggest that the properties embedded in the variable domain of individual light chain clones and their compatibility with the partnering heavy chain variable domain sequences underscore the efficiency of immunoglobulin G biosynthesis, the threshold for Russell body induction, and the level of immunoglobulin G secretion. We propose that an interplay between the unique properties encoded in variable domain sequences and the state of protein homeostasis determines whether an immunoglobulin G expressing cell will develop the Russell body phenotype in a dynamic cellular setting. Copyright © 2012 Elsevier B.V. All rights reserved.

Xeroderma pigmentosum variants have a slow recovery of DNA synthesis after irradiation with ultraviolet light

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

Cleaver, J.E.; Thomas, G.H.; Park, S.D.

1979-01-01

Human cells (normal and xeroderma pigmentosum variant) irradiated with ultraviolet light and pulse-labelled with (/sup 3/H)thymidine underwent transient decline and recovery of molecular weights of newly synthesized DNA and rates of (/sup 3/H)thymidine incorporation. The ability to synthesize normal-sized DNA recovered more rapidly in both cell types than thymidine incorporation. During recovery cells steadily increased in their ability to replicate normal-sized DNA on damaged templates. The molecular weight versus time curves fitted exponential functions with similar rate constants in normal and heterozygous xeroderma pigmentosum cells, but with a slower rate in two xeroderma pigmentosum variant cell lines. Caffeine added duringmore » the post-irradiation period eliminated the recovery of molecular weights in xeroderma pigmentosum variant but not in normal cells. The recovery of the ability to synthesize normal-sized DNA represents a combination of a number of cellular regulatory processes, some of which are constitutive, and one of which is altered in the xeroderma pigmentosum variant such that recovery becomes slow and caffeine sensitive.« less

Classification of Normal and Apoptotic Cells from Fluorescence Microscopy Images Using Generalized Polynomial Chaos and Level Set Function.

PubMed

Du, Yuncheng; Budman, Hector M; Duever, Thomas A

2016-06-01

Accurate automated quantitative analysis of living cells based on fluorescence microscopy images can be very useful for fast evaluation of experimental outcomes and cell culture protocols. In this work, an algorithm is developed for fast differentiation of normal and apoptotic viable Chinese hamster ovary (CHO) cells. For effective segmentation of cell images, a stochastic segmentation algorithm is developed by combining a generalized polynomial chaos expansion with a level set function-based segmentation algorithm. This approach provides a probabilistic description of the segmented cellular regions along the boundary, from which it is possible to calculate morphological changes related to apoptosis, i.e., the curvature and length of a cell's boundary. These features are then used as inputs to a support vector machine (SVM) classifier that is trained to distinguish between normal and apoptotic viable states of CHO cell images. The use of morphological features obtained from the stochastic level set segmentation of cell images in combination with the trained SVM classifier is more efficient in terms of differentiation accuracy as compared with the original deterministic level set method.

Creation of a virtual cutaneous tissue bank

NASA Astrophysics Data System (ADS)

LaFramboise, William A.; Shah, Sujal; Hoy, R. W.; Letbetter, D.; Petrosko, P.; Vennare, R.; Johnson, Peter C.

2000-04-01

Cellular and non-cellular constituents of skin contain fundamental morphometric features and structural patterns that correlate with tissue function. High resolution digital image acquisitions performed using an automated system and proprietary software to assemble adjacent images and create a contiguous, lossless, digital representation of individual microscope slide specimens. Serial extraction, evaluation and statistical analysis of cutaneous feature is performed utilizing an automated analysis system, to derive normal cutaneous parameters comprising essential structural skin components. Automated digital cutaneous analysis allows for fast extraction of microanatomic dat with accuracy approximating manual measurement. The process provides rapid assessment of feature both within individual specimens and across sample populations. The images, component data, and statistical analysis comprise a bioinformatics database to serve as an architectural blueprint for skin tissue engineering and as a diagnostic standard of comparison for pathologic specimens.

Context Specificity of Stress-activated Mitogen-activated Protein (MAP) Kinase Signaling: The Story as Told by Caenorhabditis elegans*

PubMed Central

Andrusiak, Matthew G.; Jin, Yishi

2016-01-01

Stress-associated p38 and JNK mitogen-activated protein (MAP) kinase signaling cascades trigger specific cellular responses and are involved in multiple disease states. At the root of MAP kinase signaling complexity is the differential use of common components on a context-specific basis. The roundworm Caenorhabditis elegans was developed as a system to study genes required for development and nervous system function. The powerful genetics of C. elegans in combination with molecular and cellular dissections has led to a greater understanding of how p38 and JNK signaling affects many biological processes under normal and stress conditions. This review focuses on the studies revealing context specificity of different stress-activated MAPK components in C. elegans. PMID:26907690

Parasitoid wasp venom SERCA regulates Drosophila calcium levels and inhibits cellular immunity.

PubMed

Mortimer, Nathan T; Goecks, Jeremy; Kacsoh, Balint Z; Mobley, James A; Bowersock, Gregory J; Taylor, James; Schlenke, Todd A

2013-06-04

Because parasite virulence factors target host immune responses, identification and functional characterization of these factors can provide insight into poorly understood host immune mechanisms. The fruit fly Drosophila melanogaster is a model system for understanding humoral innate immunity, but Drosophila cellular innate immune responses remain incompletely characterized. Fruit flies are regularly infected by parasitoid wasps in nature and, following infection, flies mount a cellular immune response culminating in the cellular encapsulation of the wasp egg. The mechanistic basis of this response is largely unknown, but wasps use a mixture of virulence proteins derived from the venom gland to suppress cellular encapsulation. To gain insight into the mechanisms underlying wasp virulence and fly cellular immunity, we used a joint transcriptomic/proteomic approach to identify venom genes from Ganaspis sp.1 (G1), a previously uncharacterized Drosophila parasitoid species, and found that G1 venom contains a highly abundant sarco/endoplasmic reticulum calcium ATPase (SERCA) pump. Accordingly, we found that fly immune cells termed plasmatocytes normally undergo a cytoplasmic calcium burst following infection, and that this calcium burst is required for activation of the cellular immune response. We further found that the plasmatocyte calcium burst is suppressed by G1 venom in a SERCA-dependent manner, leading to the failure of plasmatocytes to become activated and migrate toward G1 eggs. Finally, by genetically manipulating plasmatocyte calcium levels, we were able to alter fly immune success against G1 and other parasitoid species. Our characterization of parasitoid wasp venom proteins led us to identify plasmatocyte cytoplasmic calcium bursts as an important aspect of fly cellular immunity.

Povidone-Iodine Has a Profound Effect on In Vitro Osteoblast Proliferation and Metabolic Function and Inhibits Their Ability to Mineralize and Form Bone.

PubMed

Newton Ede, Matthew P; Philp, Ashleigh M; Philp, Andrew; Richardson, Stephen M; Mohammad, Saeed; Jones, Simon W

2016-05-01

A study examining the clinical protocol of scoliosis wound irrigation, demonstrating povidone-iodine's (PVI) effect on human osteoblast cells. Primary and immortal cell line osteoblasts were treated with 0.35% PVI for 3 minutes, and analyzed for proliferation rate, oxidative capacity, and mineralization. To model spinal wound irrigation with dilute PVI in vitro, in order to investigate the effect of PVI on osteoblast proliferation, metabolism, and bone mineralization. Previously PVI irrigation has been proposed as a safe and effective practice to avoid bacterial growth after spinal surgery. However, recent evidence in multiple cell types suggests that PVI has a deleterious effect on cellular viability and cellular function. Primary and immortal human osteoblast cells were exposed to either phosphate buffered saline control or with 0.35% PVI for 3 minutes. Cellular proliferation was measured over the duration of 7 days by MTS assay. Oxygen consumption rate, extracellular acidification rate, and proton production rate were analyzed using a Seahorse XF24 Bioanalyzer. Protein expression of the electron transport chain subunits CII-SDHB, CIII-UQRCR2, and CV-ATP5A was measured via Western blotting. Mineralized bone nodules were stained with alizarin red. Expressed as a percentage of normal osteoblast proliferation, osteoblasts exposed to 0.35% PVI exhibited a significant 24% decrease in proliferation after 24 hours. This was a sustained response, resulting in a 72% decline in cellular proliferation at 1 week. There was a significant reduction in oxygen consumption rate, extracellular acidification rate, and proton production rate (P < 0.05), in osteoblasts that had been exposed to 0.35% PVI for 3 minutes, coupled with a marked reduction in the protein expression of CII-SDHB. Osteoblasts exposed to 0.35% PVI exhibited reduced bone nodule mineralization compared to control phosphate buffered saline exposed osteoblasts (P < 0.01). PVI has a rapid and detrimental effect on human osteoblast cellular proliferation, metabolic function, and bone nodule mineralization. NA.

G2019S leucine-rich repeat kinase 2 causes uncoupling protein-mediated mitochondrial depolarization

PubMed Central

Papkovskaia, Tatiana D.; Chau, Kai-Yin; Inesta-Vaquera, Francisco; Papkovsky, Dmitri B.; Healy, Daniel G.; Nishio, Koji; Staddon, James; Duchen, Michael R.; Hardy, John; Schapira, Anthony H.V.; Cooper, J. Mark

2012-01-01

The G2019S leucine rich repeat kinase 2 (LRRK2) mutation is the most common genetic cause of Parkinson's disease (PD), clinically and pathologically indistinguishable from idiopathic PD. Mitochondrial abnormalities are a common feature in PD pathogenesis and we have investigated the impact of G2019S mutant LRRK2 expression on mitochondrial bioenergetics. LRRK2 protein expression was detected in fibroblasts and lymphoblasts at levels higher than those observed in the mouse brain. The presence of G2019S LRRK2 mutation did not influence LRRK2 expression in fibroblasts. However, the expression of the G2019S LRRK2 mutation in both fibroblast and neuroblastoma cells was associated with mitochondrial uncoupling. This was characterized by decreased mitochondrial membrane potential and increased oxygen utilization under basal and oligomycin-inhibited conditions. This resulted in a decrease in cellular ATP levels consistent with compromised cellular function. This uncoupling of mitochondrial oxidative phosphorylation was associated with a cell-specific increase in uncoupling protein (UCP) 2 and 4 expression. Restoration of mitochondrial membrane potential by the UCP inhibitor genipin confirmed the role of UCPs in this mechanism. The G2019S LRRK2-induced mitochondrial uncoupling and UCP4 mRNA up-regulation were LRRK2 kinase-dependent, whereas endogenous LRRK2 levels were required for constitutive UCP expression. We propose that normal mitochondrial function was deregulated by the expression of G2019S LRRK2 in a kinase-dependent mechanism that is a modification of the normal LRRK2 function, and this leads to the vulnerability of selected neuronal populations in PD. PMID:22736029

Cytoplasmic RNA Granules in Somatic Maintenance.

PubMed

Moujaber, Ossama; Stochaj, Ursula

2018-05-30

Cytoplasmic RNA granules represent subcellular compartments that are enriched in protein-bound RNA species. RNA granules are produced by evolutionary divergent eukaryotes, including yeast, mammals, and plants. The functions of cytoplasmic RNA granules differ widely. They are dictated by the cell type and physiological state, which in turn is determined by intrinsic cell properties and environmental factors. RNA granules provide diverse cellular functions. However, all of the granules contribute to aspects of RNA metabolism. This is exemplified by transcription, RNA storage, silencing, and degradation, as well as mRNP remodeling and regulated translation. Several forms of cytoplasmic mRNA granules are linked to normal physiological processes. For instance, they may coordinate protein synthesis and thereby serve as posttranscriptional "operons". RNA granules also participate in cytoplasmic mRNA trafficking, a process particularly well understood for neurons. Many forms of RNA granules support the preservation of somatic cell performance under normal and stress conditions. On the other hand, severe insults or disease can cause the formation and persistence of RNA granules that contribute to cellular dysfunction, especially in the nervous system. Neurodegeneration and many other diseases linked to RNA granules are associated with aging. Nevertheless, information related to the impact of aging on the various types of RNA granules is presently very limited. This review concentrates on cytoplasmic RNA granules and their role in somatic cell maintenance. We summarize the current knowledge on different types of RNA granules in the cytoplasm, their assembly and function under normal, stress, or disease conditions. Specifically, we discuss processing bodies, neuronal granules, stress granules, and other less characterized cytoplasmic RNA granules. Our focus is primarily on mammalian and yeast models, because they have been critical to unravel the physiological role of various RNA granules. RNA granules in plants and pathogens are briefly described. We conclude our viewpoint by summarizing the emerging concepts for RNA granule biology and the open questions that need to be addressed in future studies. © 2018 S. Karger AG, Basel.

Rapid Endolysosomal Escape and Controlled Intracellular Trafficking of Cell Surface Mimetic Quantum-Dots-Anchored Peptides and Glycopeptides.

PubMed

Tan, Roger S; Naruchi, Kentaro; Amano, Maho; Hinou, Hiroshi; Nishimura, Shin-Ichiro

2015-09-18

A novel strategy for the development of a high performance nanoparticules platform was established by means of cell surface mimetic quantum-dots (QDs)-anchored peptides/glycopeptides, which was developed as a model system for nanoparticle-based drug delivery (NDD) vehicles with defined functions helping the specific intracellular trafficking after initial endocytosis. In this paper, we proposed a standardized protocol for the preparation of multifunctional QDs that allows for efficient cellular uptake and rapid escaping from the endolysosomal system and subsequent cytoplasmic molecular delivery to the target cellular compartment. Chemoselective ligation of the ketone-functionalized hexahistidine derivative facilitated both efficient endocytic entry and rapid endolysosomal escape of the aminooxy/phosphorylcholine self-assembled monolayer-coated QDs (AO/PCSAM-QDs) to the cytosol in various cell lines such as human normal and cancer cells, while modifications of these QDs with cell-penetrating arginine-rich peptides showed poor cellular uptake and induced self-aggregation of AO/PCSAM-QDs. Combined use of hexahistidylated AO/PCSAM-QDs with serglycine-like glycopeptides, namely synthetic proteoglycan initiators (PGIs), elicited the entry and controlled intracellular trafficking, Golgi localization, and also excretion of these nanoparticles, which suggested that the present approach would provide an ideal platform for the design of high performance NDD systems.

Prenatal Alcohol Exposure and Cellular Differentiation

PubMed Central

Veazey, Kylee J.; Muller, Daria; Golding, Michael C.

2013-01-01

Exposure to alcohol significantly alters the developmental trajectory of progenitor cells and fundamentally compromises tissue formation (i.e., histogenesis). Emerging research suggests that ethanol can impair mammalian development by interfering with the execution of molecular programs governing differentiation. For example, ethanol exposure disrupts cellular migration, changes cell–cell interactions, and alters growth factor signaling pathways. Additionally, ethanol can alter epigenetic mechanisms controlling gene expression. Normally, lineage-specific regulatory factors (i.e., transcription factors) establish the transcriptional networks of each new cell type; the cell’s identity then is maintained through epigenetic alterations in the way in which the DNA encoding each gene becomes packaged within the chromatin. Ethanol exposure can induce epigenetic changes that do not induce genetic mutations but nonetheless alter the course of fetal development and result in a large array of patterning defects. Two crucial enzyme complexes—the Polycomb and Trithorax proteins—are central to the epigenetic programs controlling the intricate balance between self-renewal and the execution of cellular differentiation, with diametrically opposed functions. Prenatal ethanol exposure may disrupt the functions of these two enzyme complexes, altering a crucial aspect of mammalian differentiation. Characterizing the involvement of Polycomb and Trithorax group complexes in the etiology of fetal alcohol spectrum disorders will undoubtedly enhance understanding of the role that epigenetic programming plays in this complex disorder. PMID:24313167

An ultra-sensitive biophysical risk assessment of light effect on skin cells.

PubMed

Bennet, Devasier; Viswanath, Buddolla; Kim, Sanghyo; An, Jeong Ho

2017-07-18

The aim of this study was to analyze photo-dynamic and photo-pathology changes of different color light radiations on human adult skin cells. We used a real-time biophysical and biomechanics monitoring system for light-induced cellular changes in an in vitro model to find mechanisms of the initial and continuous degenerative process. Cells were exposed to intermittent, mild and intense (1-180 min) light with On/Off cycles, using blue, green, red and white light. Cellular ultra-structural changes, damages, and ECM impair function were evaluated by up/down-regulation of biophysical, biomechanical and biochemical properties. All cells exposed to different color light radiation showed significant changes in a time-dependent manner. Particularly, cell growth, stiffness, roughness, cytoskeletal integrity and ECM proteins of the human dermal fibroblasts-adult (HDF-a) cells showed highest alteration, followed by human epidermal keratinocytes-adult (HEK-a) cells and human epidermal melanocytes-adult (HEM-a) cells. Such changes might impede the normal cellular functions. Overall, the obtained results identify a new insight that may contribute to premature aging, and causes it to look aged in younger people. Moreover, these results advance our understanding of the different color light-induced degenerative process and help the development of new therapeutic strategies.

Physiological functions of MTA family of proteins.

PubMed

Sen, Nirmalya; Gui, Bin; Kumar, Rakesh

2014-12-01

Although the functional significance of the metastasic tumor antigen (MTA) family of chromatin remodeling proteins in the pathobiology of cancer is fairly well recognized, the physiological role of MTA proteins continues to be an understudied research area and is just beginning to be recognized. Similar to cancer cells, MTA1 also modulates the expression of target genes in normal cells either by acting as a corepressor or coactivator. In addition, physiological functions of MTA proteins are likely to be influenced by its differential expression, subcellular localization, and regulation by upstream modulators and extracellular signals. This review summarizes our current understanding of the physiological functions of the MTA proteins in model systems. In particular, we highlight recent advances of the role MTA proteins play in the brain, eye, circadian rhythm, mammary gland biology, spermatogenesis, liver, immunomodulation and inflammation, cellular radio-sensitivity, and hematopoiesis and differentiation. Based on the growth of knowledge regarding the exciting new facets of the MTA family of proteins in biology and medicine, we speculate that the next burst of findings in this field may reveal further molecular regulatory insights of non-redundant functions of MTA coregulators in the normal physiology as well as in pathological conditions outside cancer.

RNA sequencing of chorionic villi from recurrent pregnancy loss patients reveals impaired function of basic nuclear and cellular machinery

PubMed Central

Sõber, Siim; Rull, Kristiina; Reiman, Mario; Ilisson, Piret; Mattila, Pirkko; Laan, Maris

2016-01-01

Recurrent pregnancy loss (RPL) concerns ~3% of couples aiming at childbirth. In the current study, transcriptomes and miRNomes of 1st trimester placental chorionic villi were analysed for 2 RPL cases (≥6 miscarriages) and normal, but electively terminated pregnancies (ETP; n = 8). Sequencing was performed on Illumina HiSeq 2000 platform. Differential expression analyses detected 51 (27%) transcripts with increased and 138 (73%) with decreased expression in RPL compared to ETP (DESeq: FDR P < 0.1 and DESeq2: <0.05). RPL samples had substantially decreased transcript levels of histones, regulatory RNAs and genes involved in telomere, spliceosome, ribosomal, mitochondrial and intra-cellular signalling functions. Downregulated expression of HIST1H1B and HIST1H4A (Wilcoxon test, fc≤0.372, P≤9.37 × 10−4) was validated in an extended sample by quantitative PCR (RPL, n = 14; ETP, n = 24). Several upregulated genes are linked to placental function and pregnancy complications: ATF4, C3, PHLDA2, GPX4, ICAM1, SLC16A2. Analysis of the miRNA-Seq dataset identified no large disturbances in RPL samples. Notably, nearly 2/3 of differentially expressed genes have binding sites for E2F transcription factors, coordinating mammalian endocycle and placental development. For a conceptus destined to miscarriage, the E2F TF-family represents a potential key coordinator in reprogramming the placental genome towards gradually stopping the maintenance of basic nuclear and cellular functions. PMID:27929073

Insight into the tumor suppressor function of CBP through the viral oncoprotein tax.

PubMed

Van Orden, K; Nyborg, J K

2000-01-01

CREB binding protein (CBP) is a cellular coactivator protein that regulates essentially all known pathways of gene expression. The transcriptional coactivator properties of CBP are utilized by at least 25 different transcription factors representing nearly all known classes of DNA binding proteins. Once bound to their target genes, these transcription factors are believed to tether CBP to the promoter, leading to activated transcription. CBP functions to stimulate transcription through direct recruitment of the general transcription machinery as well as acetylation of both histone and transcription factor substrates. Recent observations indicate that a critical dosage of CBP is required for normal development and tumor suppression, and that perturbations in CBP concentrations may disrupt cellular homeostasis. Furthermore, there is accumulating evidence that CBP deregulation plays a direct role in hematopoietic malignancies. However, the molecular events linking CBP deregulation and malignant transformation are unclear. Further insight into the function of CBP, and its role as a tumor suppressor, can be gained through recent studies of the human T-cell leukemia virus, type I (HTLV-I) Tax oncoprotein. Tax is known to utilize CBP to stimulate transcription from the viral promoter. However, recent data suggest that as a consequence of the Tax-CBP interaction, many cellular transcription factor pathways may be deregulated. Tax disruption of CBP function may play a key role in transformation of the HTLV-I-infected cell. Thus, Tax derailment of CBP may lend important information about the tumor suppressor properties of CBP and serve as a model for the role of CBP in hematopoietic malignancies.

What are the origins and relevance of spontaneous bladder contractions? ICI-RS 2017.

PubMed

Drake, Marcus J; Fry, Christopher H; Hashitani, Hikaru; Kirschner-Hermanns, Ruth; Rahnama'i, Mohammad S; Speich, John E; Tomoe, Hikaru; Kanai, Anthony J; McCloskey, Karen D

2018-01-23

Storage phase bladder activity is a counter-intuitive observation of spontaneous contractions. They are potentially an intrinsic feature of the smooth muscle, but interstitial cells in the mucosa and the detrusor itself, as well as other muscular elements in the mucosa may substantially influence them. They are identified in several models explaining lower urinary tract dysfunction. A consensus meeting at the International Consultation on Incontinence Research Society (ICI-RS) 2017 congress considered the origins and relevance of spontaneous bladder contractions by debating which cell type(s) modulate bladder spontaneous activity, whether the methodologies are sufficiently robust, and implications for healthy and abnormal lower urinary tract function. The identified research priorities reflect a wide range of unknown aspects. Cellular contributions to spontaneous contractions in detrusor smooth muscle are still uncertain. Accordingly, insight into the cellular physiology of the bladder wall, particularly smooth muscle cells, interstitial cells, and urothelium, remains important. Upstream influences, such as innervation, endocrine, and paracrine factors, are particularly important. The cellular interactions represent the key understanding to derive the integrative physiology of organ function, notably the nature of signalling between mucosa and detrusor layers. Indeed, it is still not clear to what extent spontaneous contractions generated in isolated preparations mirror their normal and pathological counterparts in the intact bladder. Improved models of how spontaneous contractions influence pressure generation and sensory nerve function are also needed. Deriving approaches to robust evaluation of spontaneous contractions and their influences for experimental and clinical use could yield considerable progress in functional urology. © 2018 Wiley Periodicals, Inc.

Geometric phase transition in the cellular network of the pancreatic islets may underlie the onset of type 1diabetes

NASA Astrophysics Data System (ADS)

Wang, Xujing

Living systems are characterized by complexity in structure and emergent dynamic orders. In many aspects the onset of a chronic disease resembles phase transition in a dynamic system: quantitative changes accumulate largely unnoticed until a critical threshold is reached, which causes abrupt qualitative changes of the system. In this study we investigate this idea in a real example, the insulin-producing pancreatic islet β-cells and the onset of type 1 diabetes. Within each islet, the β-cells are electrically coupled to each other, and function as a network with synchronized actions. Using percolation theory we show how normal islet function is intrinsically linked to network connectivity, and the critical point where the islet cellular network loses site percolation, is consistent with laboratory and clinical observations of the threshold β-cell loss that causes islet functional failure. Numerical simulations confirm that the islet cellular network needs to be percolated for β-cells to synchronize. Furthermore, the interplay between site percolation and bond strength predicts the existence of a transient phase of islet functional recovery after disease onset and introduction of treatment, potentially explaining a long time mystery in the clinical study of type 1 diabetes: the honeymoon phenomenon. Based on these results, we hypothesized that the onset of T1D may be the result of a phase transition of the islet β-cell network. We further discuss the potential applications in identifying disease-driving factors, and the critical parameters that are predictive of disease onset.

A Major Binding Protein for Leukemia Inhibitory Factor in Normal Mouse Serum: Identification as a Soluble Form of the Cellular Receptor

NASA Astrophysics Data System (ADS)

Layton, Meredith J.; Cross, Bronwyn A.; Metcalf, Donald; Ward, Larry D.; Simpson, Richard J.; Nicola, Nicos A.

1992-09-01

A protein that specifically binds leukemia inhibitory factor (LIF) has been isolated from normal mouse serum by using four successive fractionation steps: chromatography on a LIF affinity matrix, anion-exchange chromatography, size-exclusion chromatography, and preparative native gel electrophoresis. The purified LIF-binding protein (LBP) is a glycoprotein with an apparent molecular mass of 90 kDa that specifically binds 125I-labeled murine LIF with an affinity comparable to that of the low-affinity cellular LIF receptor (K_d = 600 pM). N-terminal sequencing has identified this protein as a soluble truncated form of the α chain of the cellular LIF receptor. LBP is present in normal mouse serum at high levels (1 μg/ml) and these levels are elevated in pregnant mice and reduced in neonatal mice. Since normal serum concentrations of LBP can block the biological actions of LIF in culture, LBP may serve as an inhibitor of the systemic effects of locally produced LIF.

How the study of Listeria monocytogenes has led to new concepts in biology.

PubMed

Rolhion, Nathalie; Cossart, Pascale

2017-06-01

The opportunistic intracellular bacterial pathogen Listeria monocytogenes has in 30 years emerged as an exceptional bacterial model system in infection biology. Research on this bacterium has provided considerable insight into how pathogenic bacteria adapt to mammalian hosts, invade eukaryotic cells, move intracellularly, interfere with host cell functions and disseminate within tissues. It also contributed to unveil features of normal host cell pathways and unsuspected functions of previously known cellular proteins. This review provides an updated overview of our knowledge on this pathogen. In many examples, findings on L. monocytogenes provided the basis for new concepts in bacterial regulation, cell biology and infection processes.

Multilevel regulation of gene expression by microRNAs.

PubMed

Makeyev, Eugene V; Maniatis, Tom

2008-03-28

MicroRNAs (miRNAs) are approximately 22-nucleotide-long noncoding RNAs that normally function by suppressing translation and destabilizing messenger RNAs bearing complementary target sequences. Some miRNAs are expressed in a cell- or tissue-specific manner and may contribute to the establishment and/or maintenance of cellular identity. Recent studies indicate that tissue-specific miRNAs may function at multiple hierarchical levels of gene regulatory networks, from targeting hundreds of effector genes incompatible with the differentiated state to controlling the levels of global regulators of transcription and alternative pre-mRNA splicing. This multilevel regulation may allow individual miRNAs to profoundly affect the gene expression program of differentiated cells.

Investigation of a calcium-responsive contrast agent in cellular model systems: feasibility for use as a smart molecular probe in functional MRI.

PubMed

Angelovski, Goran; Gottschalk, Sven; Milošević, Milena; Engelmann, Jörn; Hagberg, Gisela E; Kadjane, Pascal; Andjus, Pavle; Logothetis, Nikos K

2014-05-21

Responsive or smart contrast agents (SCAs) represent a promising direction for development of novel functional MRI (fMRI) methods for the eventual noninvasive assessment of brain function. In particular, SCAs that respond to Ca(2+) may allow tracking neuronal activity independent of brain vasculature, thus avoiding the characteristic limitations of current fMRI techniques. Here we report an in vitro proof-of-principle study with a Ca(2+)-sensitive, Gd(3+)-based SCA in an attempt to validate its potential use as a functional in vivo marker. First, we quantified its relaxometric response in a complex 3D cell culture model. Subsequently, we examined potential changes in the functionality of primary glial cells following administration of this SCA. Monitoring intracellular Ca(2+) showed that, despite a reduction in the Ca(2+) level, transport of Ca(2+) through the plasma membrane remained unaffected, while stimulation with ATP induced Ca(2+)-transients suggested normal cellular signaling in the presence of low millimolar SCA concentrations. SCAs merely lowered the intracellular Ca(2+) level. Finally, we estimated the longitudinal relaxation times (T1) for an idealized in vivo fMRI experiment with SCA, for extracellular Ca(2+) concentration level changes expected during intense neuronal activity which takes place upon repetitive stimulation. The values we obtained indicate changes in T1 of around 1-6%, sufficient to be robustly detectable using modern MRI methods in high field scanners. Our results encourage further attempts to develop even more potent SCAs and appropriate fMRI protocols. This would result in novel methods that allow monitoring of essential physiological processes at the cellular and molecular level.

Khat a drug of abuse: roles of free radicals and antioxidants.

PubMed

Aleryani, Samir L; Aleryani, Rowaida A; Al-Akwa, Ahmed A

2011-09-01

Many articles have reviewed the health impact of Khat consumption; however the role of free radicals in the pathogenesis associated with short- and long-term consumption of Khat is absent in the literature. As free radicals and antioxidants converge across various mechanisms in normal physiological function and in disease, this review attempts to uncover the role of endogenous free radicals and the mechanism of cellular injury associated with Khat consumption. Copyright © 2010 John Wiley & Sons, Ltd.

Therapeutic Value of PLK1 Knockdown in Combination with Prostate Cancer Drugs in PIM-1 Overexpressing Prostate Cancer Cells

DTIC Science & Technology

2014-11-13

PIM kinases are not required for essential cellular functions . Furthermore, the presence of a unique hinge region in the ATP-binding site of PIM1...washing and blocking, cells were incubated with the appropriate primary antibodies overnight and incubated with fluorescent secondary antibodies...determined after 72 hrs of reverse transfection by using the CellTiter-Glo Luminescent cell viability assay and the results were normalized to RISC -free siRNA

Role of cholesterol and lipid organization in disease

NASA Astrophysics Data System (ADS)

Maxfield, Frederick R.; Tabas, Ira

2005-12-01

Membrane lipids are essential for biological functions ranging from membrane trafficking to signal transduction. The composition of lipid membranes influences their organization and properties, so it is not surprising that disorders in lipid metabolism and transport have a role in human disease. Significant recent progress has enhanced our understanding of the molecular and cellular basis of lipid-associated disorders such as Tangier disease, Niemann-Pick disease type C and atherosclerosis. These insights have also led to improved understanding of normal physiology.

Quantitative biology of single neurons

PubMed Central

Eberwine, James; Lovatt, Ditte; Buckley, Peter; Dueck, Hannah; Francis, Chantal; Kim, Tae Kyung; Lee, Jaehee; Lee, Miler; Miyashiro, Kevin; Morris, Jacqueline; Peritz, Tiina; Schochet, Terri; Spaethling, Jennifer; Sul, Jai-Yoon; Kim, Junhyong

2012-01-01

The building blocks of complex biological systems are single cells. Fundamental insights gained from single-cell analysis promise to provide the framework for understanding normal biological systems development as well as the limits on systems/cellular ability to respond to disease. The interplay of cells to create functional systems is not well understood. Until recently, the study of single cells has concentrated primarily on morphological and physiological characterization. With the application of new highly sensitive molecular and genomic technologies, the quantitative biochemistry of single cells is now accessible. PMID:22915636

Heart failure: when form fails to follow function.

PubMed

Katz, Arnold M; Rolett, Ellis L

2016-02-01

Cardiac performance is normally determined by architectural, cellular, and molecular structures that determine the heart's form, and by physiological and biochemical mechanisms that regulate the function of these structures. Impaired adaptation of form to function in failing hearts contributes to two syndromes initially called systolic heart failure (SHF) and diastolic heart failure (DHF). In SHF, characterized by high end-diastolic volume (EDV), the left ventricle (LV) cannot eject a normal stroke volume (SV); in DHF, with normal or low EDV, the LV cannot accept a normal venous return. These syndromes are now generally defined in terms of ejection fraction (EF): SHF became 'heart failure with reduced ejection fraction' (HFrEF) while DHF became 'heart failure with normal or preserved ejection fraction' (HFnEF or HFpEF). However, EF is a chimeric index because it is the ratio between SV--which measures function, and EDV--which measures form. In SHF the LV dilates when sarcomere addition in series increases cardiac myocyte length, whereas sarcomere addition in parallel can cause concentric hypertrophy in DHF by increasing myocyte thickness. Although dilatation in SHF allows the LV to accept a greater venous return, it increases the energy cost of ejection and initiates a vicious cycle that contributes to progressive dilatation. In contrast, concentric hypertrophy in DHF facilitates ejection but impairs filling and can cause heart muscle to deteriorate. Differences in the molecular signals that initiate dilatation and concentric hypertrophy can explain why many drugs that improve prognosis in SHF have little if any benefit in DHF. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2015. For permissions please email: journals.permissions@oup.com.

Evaluation of biocompatibility of sodium perborate and 30% hydrogen peroxide using the analysis of the adherence capacity and morphology of macrophages.

PubMed

Asfora, Kattyenne Kabbaz; Santos, Maria do Carmo Moreira da Silva; Montes, Marcos Antonio Japiassú Resende; de Castro, Célia Maria Machado Barbosa

2005-02-01

The purpose of this study was to evaluate the biocompatibility of the most used bleaching materials for pulpless teeth, sodium perborate and 30% hydrogen peroxide, in an experimental model of macrophages, through analysis of the adherence index and the cellular morphology. Inflammatory macrophages were obtained from peritoneal washed of Wistar rats. The evaluation of the adherence capacity of these cells to the plastic surface was conducted in Eppendorf tubes containing RPMI, after treatment with the bleaching agents diluted in 1:10, 1:100 and 1:1000 for 15 and 30 min, and incubation at 37 degrees C and humidified atmosphere of 5% CO(2) in air. The cellular morphology was verified after incubation of the cells treated with the bleaching agents in culture plaques and compared with normal cells in culture medium. Results showed that sodium perborate neither increased the adherence index, nor altered the cellular morphology when compared to the control group. The distribution, cellular morphology, cytoplasmatic and nuclear characteristics, reproduced the aspects observed in normal macrophages. However, the treatment with 30% hydrogen peroxide presented an increase in adherence index when compared to the control group (RPMI), in all dilutions, according to Mann-Whitney test (n=08 and p=0.001 for dilutions 1:10 and 1:100, and n=08 and p=0.004 for dilution 1:1000). The morphology of the cells treated with this product presented structural alterations proportionally greater, depending on the dilution of this bleaching agent, and even in the highest dilution (1:1000) the cells presented very evident alterations. This irreversible cellular damage as well as the elevation of the adherence index, characterizes the aggressive potential of 30% hydrogen peroxide, regardless of its dilution. Sodium perborate, on the other hand, showed biocompatibitity, since, no morphological nor functional alteration was observed in macrophages.

An "elite hacker": breast tumors exploit the normal microenvironment program to instruct their progression and biological diversity.

PubMed

Boudreau, Aaron; van't Veer, Laura J; Bissell, Mina J

2012-01-01

The year 2011 marked the 40 year anniversary of Richard Nixon signing the National Cancer Act, thus declaring the beginning of the "War on Cancer" in the United States. Whereas we have made tremendous progress toward understanding the genetics of tumors in the past four decades, and in developing enabling technology to dissect the molecular underpinnings of cancer at unprecedented resolution, it is only recently that the important role of the stromal microenvironment has been studied in detail. Cancer is a tissue-specific disease, and it is becoming clear that much of what we know about breast cancer progression parallels the biology of the normal breast differentiation, of which there is still much to learn. In particular, the normal breast and breast tumors share molecular, cellular, systemic and microenvironmental influences necessary for their progression. It is therefore enticing to consider a tumor to be a "rogue hacker"--one who exploits the weaknesses of a normal program for personal benefit. Understanding normal mammary gland biology and its "security vulnerabilities" may thus leave us better equipped to target breast cancer. In this review, we will provide a brief overview of the heterotypic cellular and molecular interactions within the microenvironment of the developing mammary gland that are necessary for functional differentiation, provide evidence suggesting that similar biology--albeit imbalanced and exaggerated--is observed in breast cancer progression particularly during the transition from carcinoma in situ to invasive disease. Lastly we will present evidence suggesting that the multigene signatures currently used to model cancer heterogeneity and clinical outcome largely reflect signaling from a heterogeneous microenvironment-a recurring theme that could potentially be exploited therapeutically.

Expression pattern, ethanol-metabolizing activities, and cellular localization of alcohol and aldehyde dehydrogenases in human large bowel: association of the functional polymorphisms of ADH and ALDH genes with hemorrhoids and colorectal cancer.

PubMed

Chiang, Chien-Ping; Jao, Shu-Wen; Lee, Shiao-Pieng; Chen, Pei-Chi; Chung, Chia-Chi; Lee, Shou-Lun; Nieh, Shin; Yin, Shih-Jiun

2012-02-01

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are principal enzymes responsible for metabolism of ethanol. Functional polymorphisms of ADH1B, ADH1C, and ALDH2 genes occur among racial populations. The goal of this study was to systematically determine the functional expressions and cellular localization of ADHs and ALDHs in human rectal mucosa, the lesions of adenocarcinoma and hemorrhoid, and the genetic association of allelic variations of ADH and ALDH with large bowel disorders. Twenty-one surgical specimens of rectal adenocarcinoma and the adjacent normal mucosa, including 16 paired tissues of rectal tumor, normal mucosae of rectum and sigmoid colon from the same individuals, and 18 surgical mixed hemorrhoid specimens and leukocyte DNA samples from 103 colorectal cancer patients, 67 hemorrhoid patients, and 545 control subjects recruited in previous study, were investigated. The isozyme/allozyme expression patterns of ADH and ALDH were identified by isoelectric focusing and the activities were assayed spectrophotometrically. The protein contents of ADH/ALDH isozymes were determined by immunoblotting using the corresponding purified class-specific antibodies; the cellular activity and protein localizations were detected by immunohistochemistry and histochemistry, respectively. Genotypes of ADH1B, ADH1C, and ALDH2 were determined by polymerase chain reaction-restriction fragment length polymorphisms. At 33mM ethanol, pH 7.5, the activity of ADH1C*1/1 phenotypes exhibited 87% higher than that of the ADH1C*1/*2 phenotypes in normal rectal mucosa. The activity of ALDH2-active phenotypes of rectal mucosa was 33% greater than ALDH2-inactive phenotypes at 200μM acetaldehyde. The protein contents in normal rectal mucosa were in the following order: ADH1>ALDH2>ADH3≈ALDH1A1, whereas those of ADH2, ADH4, and ALDH3A1 were fairly low. Both activity and content of ADH1 were significantly decreased in rectal tumors, whereas the ALDH activity remained unchanged. The ADH activity was also significantly reduced in hemorrhoids. ADH4 and ALDH3A1 were uniquely expressed in the squamous epithelium of anus at anorectal junctions. The allele frequencies of ADH1C*1 and ALDH2*2 were significantly higher in colorectal cancer and that of ALDH2*2 also significantly greater in hemorrhoids. In conclusion, ADH and ALDH isozymes are differentially expressed in mucosal cells of rectum and anus. The results suggest that acetaldehyde, an immediate metabolite of ethanol, may play an etiological role in pathogenesis of large bowel diseases. Copyright © 2012 Elsevier Inc. All rights reserved.

Mapping the cellular and molecular heterogeneity of normal and malignant breast tissues and cultured cell lines

PubMed Central

2010-01-01

Introduction Normal and neoplastic breast tissues are comprised of heterogeneous populations of epithelial cells exhibiting various degrees of maturation and differentiation. While cultured cell lines have been derived from both normal and malignant tissues, it remains unclear to what extent they retain similar levels of differentiation and heterogeneity as that found within breast tissues. Methods We used 12 reduction mammoplasty tissues, 15 primary breast cancer tissues, and 20 human breast epithelial cell lines (16 cancer lines, 4 normal lines) to perform flow cytometry for CD44, CD24, epithelial cell adhesion molecule (EpCAM), and CD49f expression, as well as immunohistochemistry, and in vivo tumor xenograft formation studies to extensively analyze the molecular and cellular characteristics of breast epithelial cell lineages. Results Human breast tissues contain four distinguishable epithelial differentiation states (two luminal phenotypes and two basal phenotypes) that differ on the basis of CD24, EpCAM and CD49f expression. Primary human breast cancer tissues also contain these four cellular states, but in altered proportions compared to normal tissues. In contrast, cultured cancer cell lines are enriched for rare basal and mesenchymal epithelial phenotypes, which are normally present in small numbers within human tissues. Similarly, cultured normal human mammary epithelial cell lines are enriched for rare basal and mesenchymal phenotypes that represent a minor fraction of cells within reduction mammoplasty tissues. Furthermore, although normal human mammary epithelial cell lines exhibit features of bi-potent progenitor cells they are unable to differentiate into mature luminal breast epithelial cells under standard culture conditions. Conclusions As a group breast cancer cell lines represent the heterogeneity of human breast tumors, but individually they exhibit increased lineage-restricted profiles that fall short of truly representing the intratumoral heterogeneity of individual breast tumors. Additionally, normal human mammary epithelial cell lines fail to retain much of the cellular diversity found in human breast tissues and are enriched for differentiation states that are a minority in breast tissues, although they do exhibit features of bi-potent basal progenitor cells. These findings suggest that collections of cell lines representing multiple cell types can be used to model the cellular heterogeneity of tissues. PMID:20964822

Phase transitions in pancreatic islet cellular networks and implications for type-1 diabetes

NASA Astrophysics Data System (ADS)

Stamper, I. J.; Jackson, Elais; Wang, Xujing

2014-01-01

In many aspects the onset of a chronic disease resembles a phase transition in a complex dynamic system: Quantitative changes accumulate largely unnoticed until a critical threshold is reached, which causes abrupt qualitative changes of the system. In this study we examine a special case, the onset of type-1 diabetes (T1D), a disease that results from loss of the insulin-producing pancreatic islet β cells. Within each islet, the β cells are electrically coupled to each other via gap-junctional channels. This intercellular coupling enables the β cells to synchronize their insulin release, thereby generating the multiscale temporal rhythms in blood insulin that are critical to maintaining blood glucose homeostasis. Using percolation theory we show how normal islet function is intrinsically linked to network connectivity. In particular, the critical amount of β-cell death at which the islet cellular network loses site percolation is consistent with laboratory and clinical observations of the threshold loss of β cells that causes islet functional failure. In addition, numerical simulations confirm that the islet cellular network needs to be percolated for β cells to synchronize. Furthermore, the interplay between site percolation and bond strength predicts the existence of a transient phase of islet functional recovery after onset of T1D and introduction of treatment, potentially explaining the honeymoon phenomenon. Based on these results, we hypothesize that the onset of T1D may be the result of a phase transition of the islet β-cell network.

Oxidative stress and mitochondrial adaptive shift during pituitary tumoral growth.

PubMed

Sabatino, Maria Eugenia; Grondona, Ezequiel; Sosa, Liliana D V; Mongi Bragato, Bethania; Carreño, Lucia; Juarez, Virginia; da Silva, Rodrigo A; Remor, Aline; de Bortoli, Lucila; de Paula Martins, Roberta; Pérez, Pablo A; Petiti, Juan Pablo; Gutiérrez, Silvina; Torres, Alicia I; Latini, Alexandra; De Paul, Ana L

2018-05-20

The cellular transformation of normal functional cells to neoplastic ones implies alterations in the cellular metabolism and mitochondrial function in order to provide the bioenergetics and growth requirements for tumour growth progression. Currently, the mitochondrial physiology and dynamic shift during pituitary tumour development are not well understood. Pituitary tumours present endocrine neoplastic benign growth which, in previous reports, we had shown that in addition to increased proliferation, these tumours were also characterized by cellular senescence signs with no indication of apoptosis. Here, we show clear evidence of oxidative stress in pituitary cells, accompanied by bigger and round mitochondria during tumour development, associated with augmented biogenesis and an increased fusion process. An activation of the Nrf2 stress response pathway together with the attenuation of the oxidative damage signs occurring during tumour development were also observed which will probably provide survival advantages to the pituitary cells. These neoplasms also presented a progressive increase in lactate production, suggesting a metabolic shift towards glycolysis metabolism. These findings might imply an oxidative stress state that could impact on the pathogenesis of pituitary tumours. These data may also reflect that pituitary cells can modulate their metabolism to adapt to different energy requirements and signalling events in a pathophysiological situation to obtain protection from damage and enhance their survival chances. Thus, we suggest that mitochondria function, oxidative stress or damage might play a critical role in pituitary tumour progression. Copyright © 2018 Elsevier Inc. All rights reserved.

Optical and Nanoparticle Analysis of Normal and Cancer Cells by Light Transmission Spectroscopy

NASA Astrophysics Data System (ADS)

Deatsch, Alison; Sun, Nan; Johnson, Jeffery; Stack, Sharon; Szajko, John; Sander, Christopher; Rebuyon, Roland; Easton, Judah; Tanner, Carol; Ruggiero, Steven

2015-03-01

We have investigated the optical properties of human oral and ovarian cancer and normal cells. Specifically, we have measured the absolute optical extinction for intra-cellular material (lysates) in aqueous suspension. Measurements were conducted over a wavelength range of 250 to 1000 nm with 1 nm resolution using Light Transmission Spectroscopy (LTS). This provides both the absolute extinction of materials under study and, with Mie inversion, the absolute number of particles of a given diameter as a function of diameter in the range of 1 to 3000 nm. Our preliminary studies show significant differences in both the extinction and particle size distributions associated with cancer versus normal cells, which appear to be correlated with differences in the particle size distribution in the range of approximately 50 to 250 nm. Especially significant is a clearly higher density of particles at about 100 nm and smaller for normal cells. Department of Physics, Harper Cancer Research Institute, and the Office of Research at the University of Notre Dame.

MET receptor tyrosine kinase as an autism genetic risk factor.

PubMed

Peng, Yun; Huentelman, Matthew; Smith, Christopher; Qiu, Shenfeng

2013-01-01

In this chapter, we will briefly discuss recent literature on the role of MET receptor tyrosine kinase (RTK) in brain development and how perturbation of MET signaling may alter normal neurodevelopmental outcomes. Recent human genetic studies have established MET as a risk factor for autism, and the molecular and cellular underpinnings of this genetic risk are only beginning to emerge from obscurity. Unlike many autism risk genes that encode synaptic proteins, the spatial and temporal expression pattern of MET RTK indicates this signaling system is ideally situated to regulate neuronal growth, functional maturation, and establishment of functional brain circuits, particularly in those brain structures involved in higher levels of cognition, social skills, and executive functions. © 2013 Elsevier Inc. All rights reserved.

Human Immune Disorder Arising from Mutation of the α Chain of the Interleukin-2 Receptor

NASA Astrophysics Data System (ADS)

Sharfe, Nigel; Dadi, Harjit K.; Shahar, Michal; Roifman, Chaim M.

1997-04-01

Profound cellular immunodeficiency occurs as the result of mutations in proteins involved in both the differentiation and function of mature lymphoid cells. We describe here a novel human immune aberration arising from a truncation mutation of the interleukin-2 receptor α chain (CD25), a subunit of the tripartite high-affinity receptor for interleukin 2. This immunodeficiency is characterized by decreased numbers of peripheral T cells displaying abnormal proliferation but normal B cell development. Extensive lymphocytic infiltration of tissues, including lung, liver, gut, and bone, is observed, accompanied by tissue atrophy and inflammation. Although mature T cells are present, the absence of CD25 does affect the differentiation of thymocytes. While displaying normal development of CD2, CD3, CD4, and CD8 expression, CD25-deficient cortical thymocytes do not express CD1, and furthermore they fail to normally down-regulate levels of the anti-apoptotic protein bcl-2.

Integration of the ubiquitin-proteasome pathway with a cytosolic oligopeptidase activity

PubMed Central

Wang, Evelyn W.; Kessler, Benedikt M.; Borodovsky, Anna; Cravatt, Benjamin F.; Bogyo, Matthew; Ploegh, Hidde L.; Glas, Rickard

2000-01-01

Cytosolic proteolysis is carried out predominantly by the proteasome. We show that a large oligopeptidase, tripeptidylpeptidase II (TPPII), can compensate for compromised proteasome activity. Overexpression of TPPII is sufficient to prevent accumulation of polyubiquitinated proteins and allows survival of EL-4 cells at otherwise lethal concentrations of the covalent proteasome inhibitor NLVS (NIP-leu-leu-leu-vinylsulfone). Elevated TPPII activity also partially restores peptide loading of MHC molecules. Purified proteasomes from adapted cells lack the chymotryptic-like activity, but still degrade longer peptide substrates via residual activity of their Z subunits. However, growth of adapted cells depends on induction of other proteolytic activities. Therefore, cytosolic oligopeptidases such as TPPII normalize rates of intracellular protein breakdown required for normal cellular function and viability. PMID:10954757

Physiological Ranges of Matrix Rigidity Modulate Primary Mouse Hepatocyte Function In Part Through Hepatocyte Nuclear Factor 4 Alpha

PubMed Central

Desai, Seema S.; Tung, Jason C.; Zhou, Vivian X.; Grenert, James P.; Malato, Yann; Rezvani, Milad; Español-Suñer, Regina; Willenbring, Holger; Weaver, Valerie M.; Chang, Tammy T.

2016-01-01

Matrix rigidity has important effects on cell behavior and is increased during liver fibrosis; however, its effect on primary hepatocyte function is unknown. We hypothesized that increased matrix rigidity in fibrotic livers would activate mechanotransduction in hepatocytes and lead to inhibition of hepatic-specific functions. To determine the physiologically relevant ranges of matrix stiffness at the cellular level, we performed detailed atomic force microscopy analysis across liver lobules from normal and fibrotic livers. We determined that normal liver matrix stiffness was around 150Pa and increased to 1–6kPa in areas near fibrillar collagen deposition in fibrotic livers. In vitro culture of primary hepatocytes on collagen matrix of tunable rigidity demonstrated that fibrotic levels of matrix stiffness had profound effects on cytoskeletal tension and significantly inhibited hepatocyte-specific functions. Normal liver stiffness maintained functional gene regulation by hepatocyte nuclear factor 4 alpha (HNF4α) whereas fibrotic matrix stiffness inhibited the HNF4α transcriptional network. Fibrotic levels of matrix stiffness activated mechanotransduction in primary hepatocytes through focal adhesion kinase (FAK). In addition, blockade of the Rho/Rho-associated protein kinase (ROCK) pathway rescued HNF4α expression from hepatocytes cultured on stiff matrix. Conclusion Fibrotic levels of matrix stiffness significantly inhibit hepatocyte-specific functions in part by inhibiting the HNF4α transcriptional network mediated through the Rho/ROCK pathway. Increased appreciation of the role of matrix rigidity in modulating hepatocyte function will advance our understanding of the mechanisms of hepatocyte dysfunction in liver cirrhosis and spur development of novel treatments for chronic liver disease. PMID:26755329

Roles of the Drosophila LRRK2 homolog in Rab7-dependent lysosomal positioning.

PubMed

Dodson, Mark W; Zhang, Ting; Jiang, Changan; Chen, Shengdi; Guo, Ming

2012-03-15

LRRK2 (PARK8) is the most common genetic determinant of Parkinson's disease (PD), with dominant mutations in LRRK2 causing inherited PD and sequence variation at the LRRK2 locus associated with increased risk for sporadic PD. Although LRRK2 has been implicated in diverse cellular processes encompassing almost all cellular compartments, the precise functions of LRRK2 remain unclear. Here, we show that the Drosophila homolog of LRRK2 (Lrrk) localizes to the membranes of late endosomes and lysosomes, physically interacts with the crucial mediator of late endosomal transport Rab7 and negatively regulates rab7-dependent perinuclear localization of lysosomes. We also show that a mutant form of lrrk analogous to the pathogenic LRRK2(G2019S) allele behaves oppositely to wild-type lrrk in that it promotes rather than inhibits rab7-dependent perinuclear lysosome clustering, with these effects of mutant lrrk on lysosome position requiring both microtubules and dynein. These data suggest that LRRK2 normally functions in Rab7-dependent lysosomal positioning, and that this function is disrupted by the most common PD-causing LRRK2 mutation, linking endolysosomal dysfunction to the pathogenesis of LRRK2-mediated PD.

Poliovirus Replication Requires the N-terminus but not the Catalytic Sec7 Domain of ArfGEF GBF1

PubMed Central

Belov, George A.; Kovtunovych, Gennadiy; Jackson, Catherine L.; Ehrenfeld, Ellie

2010-01-01

Viruses are intracellular parasites whose reproduction relies on factors provided by the host. The cellular protein GBF1 is critical for poliovirus replication. Here we show that the contribution of GBF1 to virus replication is different from its known activities in uninfected cells. Normally GBF1 activates the Arf GTPases necessary for formation of COPI transport vesicles. GBF1 function is modulated by p115 and Rab1b. However, in polio-infected cells, p115 is degraded and neither p115 nor Rab1b knock-down affects virus replication. Poliovirus infection is very sensitive to BFA, an inhibitor of Arf activation by GBF1. BFA targets the catalytic Sec7 domain of GBF1. Nevertheless the BFA block of polio replication is rescued by expression of only the N-terminal region of GBF1 lacking the Sec7 domain. Replication of BFA-resistant poliovirus in the presence of BFA is uncoupled from Arf activation but is dependent on GBF1. Thus the function(s) of this protein essential for viral replication can be separated from those required for cellular metabolism. PMID:20497182

Protein and cell micropatterning and its integration with micro/nanoparticles assembly.

PubMed

Yap, F L; Zhang, Y

2007-01-15

Micropatterning of proteins and cells has become very popular over the past decade due to its importance in the development of biosensors, microarrays, tissue engineering and cellular studies. This article reviews the techniques developed for protein and cell micropatterning and its biomedical applications. The prospect of integrating micro and nanoparticles with protein and cell micropatterning is discussed. The micro/nanoparticles are assembled into patterns and form the substrate for proteins and cell attachment. The assembled particles create a micro or nanotopography, depending on the size of the particles employed. The nonplanar structure can increase the surface area for biomolecules attachment and therefore enhance the sensitivity for detection in biosensors. Furthermore, a nanostructured substrate can influence the conformation and functionality of protein attached to it, while cellular response in terms of morphology, adhesion, proliferation, differentiation, etc. can be affected by a surface expressing micro or nanoscale structures. Proteins and cells tend to lose their normal functions upon attachment to substrate. By recognizing the types of topography that are favourable for preserving proteins and cell behaviour, and integrating it with micropattering will lead to the development of functional protein and cell patterns.

α7 nicotinic ACh receptors as a ligand-gated source of Ca(2+) ions: the search for a Ca(2+) optimum.

PubMed

Uteshev, Victor V

2012-01-01

The spatiotemporal distribution of cytosolic Ca(2+) ions is a key determinant of neuronal behavior and survival. Distinct sources of Ca(2+) ions including ligand- and voltage-gated Ca(2+) channels contribute to intracellular Ca(2+) homeostasis. Many normal physiological and therapeutic neuronal functions are Ca(2+)-dependent, however an excess of cytosolic Ca(2+) or a lack of the appropriate balance between Ca(2+) entry and clearance may destroy cellular integrity and cause cellular death. Therefore, the existence of optimal spatiotemporal patterns of cytosolic Ca(2+) elevations and thus, optimal activation of ligand- and voltage-gated Ca(2+) ion channels are postulated to benefit neuronal function and survival. Alpha7 nicotinic -acetylcholine receptors (nAChRs) are highly permeable to Ca(2+) ions and play an important role in modulation of neurotransmitter release, gene expression and neuroprotection in a variety of neuronal and non-neuronal cells. In this review, the focus is placed on α7 nAChR-mediated currents and Ca(2+) influx and how this source of Ca(2+) entry compares to NMDA receptors in supporting cytosolic Ca(2+) homeostasis, neuronal function and survival.

Roles of the Drosophila LRRK2 homolog in Rab7-dependent lysosomal positioning

PubMed Central

Dodson, Mark W.; Zhang, Ting; Jiang, Changan; Chen, Shengdi; Guo, Ming

2012-01-01

LRRK2 (PARK8) is the most common genetic determinant of Parkinson's disease (PD), with dominant mutations in LRRK2 causing inherited PD and sequence variation at the LRRK2 locus associated with increased risk for sporadic PD. Although LRRK2 has been implicated in diverse cellular processes encompassing almost all cellular compartments, the precise functions of LRRK2 remain unclear. Here, we show that the Drosophila homolog of LRRK2 (Lrrk) localizes to the membranes of late endosomes and lysosomes, physically interacts with the crucial mediator of late endosomal transport Rab7 and negatively regulates rab7-dependent perinuclear localization of lysosomes. We also show that a mutant form of lrrk analogous to the pathogenic LRRK2G2019S allele behaves oppositely to wild-type lrrk in that it promotes rather than inhibits rab7-dependent perinuclear lysosome clustering, with these effects of mutant lrrk on lysosome position requiring both microtubules and dynein. These data suggest that LRRK2 normally functions in Rab7-dependent lysosomal positioning, and that this function is disrupted by the most common PD-causing LRRK2 mutation, linking endolysosomal dysfunction to the pathogenesis of LRRK2-mediated PD. PMID:22171073

Cellular function reinstitution of offspring red blood cells cloned from the sickle cell disease patient blood post CRISPR genome editing.

PubMed

Wen, Jianguo; Tao, Wenjing; Hao, Suyang; Zu, Youli

2017-06-13

Sickle cell disease (SCD) is a disorder of red blood cells (RBCs) expressing abnormal hemoglobin-S (HbS) due to genetic inheritance of homologous HbS gene. However, people with the sickle cell trait (SCT) carry a single allele of HbS and do not usually suffer from SCD symptoms, thus providing a rationale to treat SCD. To validate gene therapy potential, hematopoietic stem cells were isolated from the SCD patient blood and treated with CRISPR/Cas9 approach. To precisely dissect genome-editing effects, erythroid progenitor cells were cloned from single colonies of CRISPR-treated cells and then expanded for simultaneous gene, protein, and cellular function studies. Genotyping and sequencing analysis revealed that the genome-edited erythroid progenitor colonies were converted to SCT genotype from SCD genotype. HPLC protein assays confirmed reinstallation of normal hemoglobin at a similar level with HbS in the cloned genome-edited erythroid progenitor cells. For cell function evaluation, in vitro RBC differentiation of the cloned erythroid progenitor cells was induced. As expected, cell sickling assays indicated function reinstitution of the genome-edited offspring SCD RBCs, which became more resistant to sickling under hypoxia condition. This study is an exploration of genome editing of SCD HSPCs.

Oxidative Modulation of Voltage-Gated Potassium Channels

PubMed Central

Sahoo, Nirakar; Hoshi, Toshinori

2014-01-01

Abstract Significance: Voltage-gated K+ channels are a large family of K+-selective ion channel protein complexes that open on membrane depolarization. These K+ channels are expressed in diverse tissues and their function is vital for numerous physiological processes, in particular of neurons and muscle cells. Potentially reversible oxidative regulation of voltage-gated K+ channels by reactive species such as reactive oxygen species (ROS) represents a contributing mechanism of normal cellular plasticity and may play important roles in diverse pathologies including neurodegenerative diseases. Recent Advances: Studies using various protocols of oxidative modification, site-directed mutagenesis, and structural and kinetic modeling provide a broader phenomenology and emerging mechanistic insights. Critical Issues: Physicochemical mechanisms of the functional consequences of oxidative modifications of voltage-gated K+ channels are only beginning to be revealed. In vivo documentation of oxidative modifications of specific amino-acid residues of various voltage-gated K+ channel proteins, including the target specificity issue, is largely absent. Future Directions: High-resolution chemical and proteomic analysis of ion channel proteins with respect to oxidative modification combined with ongoing studies on channel structure and function will provide a better understanding of how the function of voltage-gated K+ channels is tuned by ROS and the corresponding reducing enzymes to meet cellular needs. Antioxid. Redox Signal. 21, 933–952. PMID:24040918

Soluble interleukin 2 receptors are released from activated human lymphoid cells in vitro

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

Rubin, L.A.; Kurman, C.C.; Fritz, M.E.

1985-11-01

With the use of an enzyme-linked immunoabsorbent assay to measure soluble human interleukin 2 receptors (IL 2R), certain human T cell leukemia virus I (HTLV I)-positive T cell lines were found to spontaneously release large quantities of IL 2R into culture supernatants. This was not found with HTLV I-negative and IL 2 independent T cell lines, and only one of seven B cell-derived lines examined produced small amounts of IL 2R. In addition to this constitutive production of soluble IL 2R by certain cell lines, normal human peripheral blood mononuclear cells (PBMC) could be induced to release soluble IL 2Rmore » by plant lectins, the murine monoclonal antibody OKT3, tetanus toxoid, and allogeneic cells. Such activated cells also expressed cellular IL 2R measurable in detergent solubilized cell extracts. The generation of cellular and supernatant IL 2R was: dependent on cellular activation, rapid, radioresistant (3000 rad), and inhibited by cycloheximide treatment. NaDodSO4-polyacrylamide gel electrophoresis analysis of soluble IL 2R demonstrated molecules of apparent Mr = 35,000 to 40,000, and 45,000 to 50,000, respectively, somewhat smaller than the mature surface receptor on these cells. The release of soluble IL 2R appears to be a characteristic marker of T lymphocyte activation and might serve an immunoregulatory function during both normal and abnormal cell growth and differentiation.« less

Targeting pH regulating proteins for cancer therapy-Progress and limitations.

PubMed

Parks, Scott K; Pouysségur, Jacques

2017-04-01

Tumour acidity induced by metabolic alterations and incomplete vascularisation sets cancer cells apart from normal cellular physiology. This distinguishing tumour characteristic has been an area of intense study, as cellular pH (pH i ) disturbances disrupt protein function and therefore multiple cellular processes. Tumour cells effectively utilise pH i regulating machinery present in normal cells with enhancements provided by additional oncogenic or hypoxia induced protein modifications. This overall improvement of pH regulation enables maintenance of an alkaline pH i in the continued presence of external acidification (pH e ). Considerable experimentation has revealed targets that successfully disrupt tumour pH i regulation in efforts to develop novel means to weaken or kill tumour cells. However, redundancy in these pH-regulating proteins, which include Na + /H + exchangers (NHEs), carbonic anhydrases (CAs), Na + /HCO 3 - co-transporters (NBCs) and monocarboxylate transporters (MCTs) has prevented effective disruption of tumour pH i when individual protein targeting is performed. Here we synthesise recent advances in understanding both normoxic and hypoxic pH regulating mechanisms in tumour cells with an ultimate focus on the disruption of tumour growth, survival and metastasis. Interactions between tumour acidity and other cell types are also proving to be important in understanding therapeutic applications such as immune therapy. Promising therapeutic developments regarding pH manipulation along with current limitations are highlighted to provide a framework for future research directives. Copyright © 2017 Elsevier Ltd. All rights reserved.

Kinetic memory based on the enzyme-limited competition.

PubMed

Hatakeyama, Tetsuhiro S; Kaneko, Kunihiko

2014-08-01

Cellular memory, which allows cells to retain information from their environment, is important for a variety of cellular functions, such as adaptation to external stimuli, cell differentiation, and synaptic plasticity. Although posttranslational modifications have received much attention as a source of cellular memory, the mechanisms directing such alterations have not been fully uncovered. It may be possible to embed memory in multiple stable states in dynamical systems governing modifications. However, several experiments on modifications of proteins suggest long-term relaxation depending on experienced external conditions, without explicit switches over multi-stable states. As an alternative to a multistability memory scheme, we propose "kinetic memory" for epigenetic cellular memory, in which memory is stored as a slow-relaxation process far from a stable fixed state. Information from previous environmental exposure is retained as the long-term maintenance of a cellular state, rather than switches over fixed states. To demonstrate this kinetic memory, we study several models in which multimeric proteins undergo catalytic modifications (e.g., phosphorylation and methylation), and find that a slow relaxation process of the modification state, logarithmic in time, appears when the concentration of a catalyst (enzyme) involved in the modification reactions is lower than that of the substrates. Sharp transitions from a normal fast-relaxation phase into this slow-relaxation phase are revealed, and explained by enzyme-limited competition among modification reactions. The slow-relaxation process is confirmed by simulations of several models of catalytic reactions of protein modifications, and it enables the memorization of external stimuli, as its time course depends crucially on the history of the stimuli. This kinetic memory provides novel insight into a broad class of cellular memory and functions. In particular, applications for long-term potentiation are discussed, including dynamic modifications of calcium-calmodulin kinase II and cAMP-response element-binding protein essential for synaptic plasticity.

Luminescent single-walled carbon nanotube-sensitized europium nanoprobes for cellular imaging

PubMed Central

Avti, Pramod K; Sitharaman, Balaji

2012-01-01

Lanthanoid-based optical probes with excitation wavelengths in the ultra-violet (UV) range (300–325 nm) have been widely developed as imaging probes. Efficient cellular imaging requires that lanthanoid optical probes be excited at visible wavelengths, to avoid UV damage to cells. The efficacy of europium-catalyzed single-walled carbon nanotubes (Eu-SWCNTs), as visible nanoprobes for cellular imaging, is reported in this study. Confocal fluorescence microscopy images of breast cancer cells (SK-BR-3 and MCF-7) and normal cells (NIH 3T3), treated with Eu-SWCNT at 0.2 μg/mL concentration, showed bright red luminescence after excitation at 365 nm and 458 nm wavelengths. Cell viability analysis showed no cytotoxic effects after the incubation of cells with Eu-SWCNTs at this concentration. Eu-SWCNT uptake is via the endocytosis mechanism. Labeling efficiency, defined as the percentage of incubated cells that uptake Eu-SWCNT, was 95%–100% for all cell types. The average cellular uptake concentration was 6.68 ng Eu per cell. Intracellular localization was further corroborated by transmission electron microscopy and Raman microscopy. The results indicate that Eu-SWCNT shows potential as a novel cellular imaging probe, wherein SWCNT sensitizes Eu3+ ions to allow excitation at visible wavelengths, and stable time-resolved red emission. The ability to functionalize biomolecules on the exterior surface of Eu-SWCNT makes it an excellent candidate for targeted cellular imaging. PMID:22619533

Hippo signaling is required for Notch-dependent smooth muscle differentiation of neural crest.

PubMed

Manderfield, Lauren J; Aghajanian, Haig; Engleka, Kurt A; Lim, Lillian Y; Liu, Feiyan; Jain, Rajan; Li, Li; Olson, Eric N; Epstein, Jonathan A

2015-09-01

Notch signaling has well-defined roles in the assembly of arterial walls and in the development of the endothelium and smooth muscle of the vasculature. Hippo signaling regulates cellular growth in many tissues, and contributes to regulation of organ size, in addition to other functions. Here, we show that the Notch and Hippo pathways converge to regulate smooth muscle differentiation of the neural crest, which is crucial for normal development of the aortic arch arteries and cranial vasculature during embryonic development. Neural crest-specific deletion of the Hippo effectors Yap and Taz produces neural crest precursors that migrate normally, but fail to produce vascular smooth muscle, and Notch target genes such as Jagged1 fail to activate normally. We show that Yap is normally recruited to a tissue-specific Jagged1 enhancer by directly interacting with the Notch intracellular domain (NICD). The Yap-NICD complex is recruited to chromatin by the DNA-binding protein Rbp-J in a Tead-independent fashion. Thus, Hippo signaling can modulate Notch signaling outputs, and components of the Hippo and Notch pathways physically interact. Convergence of Hippo and Notch pathways by the mechanisms described here might be relevant for the function of these signaling cascades in many tissues and in diseases such as cancer. © 2015. Published by The Company of Biologists Ltd.

Calcitonin gene related family peptides: importance in normal placental and fetal development.

PubMed

Yallampalli, Chandra; Chauhan, Madhu; Endsley, Janice; Sathishkumar, Kunju

2014-01-01

Synchronized molecular and cellular events occur between the uterus and the implanting embryo to facilitate successful pregnancy outcome. Nevertheless, the molecular signaling network that coordinates strategies for successful decidualization, placentation and fetal growth are not well understood. The discovery of calcitonin/calcitonin gene-related peptides (CT/CGRP) highlighted new signaling mediators in various physiological processes, including reproduction. It is known that CGRP family peptides including CGRP, adrenomedulin and intermedin play regulatory functions during implantation, trophoblast proliferation and invasion, and fetal organogenesis. In addition, all the CGRP family peptides and their receptor components are found to be expressed in decidual, placental and fetal tissues. Additionally, plasma levels of peptides of the CGRP family were found to fluctuate during normal gestation and to induce placental cellular differentiation, proliferation, and critical hormone signaling. Moreover, aberrant signaling of these CGRP family peptides during gestation has been associated with pregnancy disorders. It indicates the existence of a possible regulatory role for these molecules during decidualization and placentation processes, which are known to be particularly vulnerable. In this review, the influence of the CGRP family peptides in these critical processes is explored and discussed.

Moringa oleifera-based diet protects against nickel-induced hepatotoxicity in rats.

PubMed

Stephen Adeyemi, Oluyomi; Sokolayemji Aroge, Cincin; Adewumi Akanji, Musbau

2017-07-13

Multiple health-promoting effects have been attributed to the consumption of Moringa oleifera leaves, as part of diet without adequate scientific credence. This study evaluated the effect of M. oleifera-based diets on nickel (Ni) - induced hepatotoxicity in rats. Male rats assigned into six groups were given oral administration of 20 mg/kg body weight nickel sulfate in normal saline and either fed normal diet orM. oleifera-based diets for 21 days. All animals were sacrificed under anesthesia 24 hours after the last treatment. Ni exposure elevated the rat plasma activities of alanine transaminase, aspartate transaminase and alkaline phosphatase significantly. Ni exposure also raised the levels of triglyceride, total cholesterol and low-density lipoprotein cholesterol while depleting the high-density lipoprotein cholesterol concentration. Further, Ni exposure raised rat plasma malondialdehyde but depleted reduced glutathione concentrations. The histopathological presentations revealed inflammation and cellular degeneration caused by Ni exposure. We show evidence thatM. oleifera-based diets protected against Ni-induced hepatotoxicity by improving the rat liver function indices, lipid profile as well as restoring cellular architecture and integrity. Study lends credence to the health-promoting value ofM. oleifera as well as underscores its potential to attenuate hepatic injury.

The multifaceted functions of C/EBPα in normal and malignant haematopoiesis.

PubMed

Ohlsson, E; Schuster, M B; Hasemann, M; Porse, B T

2016-04-01

The process of blood formation, haematopoiesis, depends upon a small number of haematopoietic stem cells (HSCs) that reside in the bone marrow. Differentiation of HSCs is characterised by decreased expression of genes associated with self-renewal accompanied by a stepwise activation of genes promoting differentiation. Lineage branching is further directed by groups of cooperating and counteracting genes forming complex networks of lineage-specific transcription factors. Imbalances in such networks can result in blockage of differentiation, lineage reprogramming and malignant transformation. CCAAT/enhancer-binding protein-α (C/EBPα) was originally identified 30 years ago as a transcription factor that binds both promoter and enhancer regions. Most of the early work focused on the role of C/EBPα in regulating transcriptional processes as well as on its functions in key differentiation processes during liver, adipogenic and haematopoietic development. Specifically, C/EBPα was shown to control differentiation by its ability to coordinate transcriptional output with cell cycle progression. Later, its role as an important tumour suppressor, mainly in acute myeloid leukaemia (AML), was recognised and has been the focus of intense studies by a number of investigators. More recent work has revisited the role of C/EBPα in normal haematopoiesis, especially its function in HSCs, and also started to provide more mechanistic insights into its role in normal and malignant haematopoiesis. In particular, the differential actions of C/EBPα isoforms, as well as its importance in chromatin remodelling and cellular reprogramming, are beginning to be elucidated. Finally, recent work has also shed light on the dichotomous function of C/EBPα in AML by demonstrating its ability to act as both a tumour suppressor and promoter. In the present review, we will summarise the current knowledge on the functions of C/EBPα during normal and malignant haematopoiesis with special emphasis on the recent work.

Noncanonical Gβ Gib2 is a scaffolding protein promoting cAMP signaling through functions of Ras1 and Cac1 proteins in Cryptococcus neoformans.

PubMed

Wang, Yanli; Shen, Gui; Gong, Jinjun; Shen, Danyu; Whittington, Amy; Qing, Jiang; Treloar, Joshua; Boisvert, Scott; Zhang, Zhengguang; Yang, Cai; Wang, Ping

2014-05-02

Gβ-like/RACK1 functions as a key mediator of various pathways and contributes to numerous cellular functions in eukaryotic organisms. In the pathogenic fungus Cryptococcus neoformans, noncanonical Gβ Gib2 promotes cAMP signaling in cells lacking normal Gpa1 function while displaying versatility in interactions with Gα Gpa1, protein kinase Pkc1, and endocytic intersectin Cin1. To elucidate the Gib2 functional mechanism(s), we demonstrate that Gib2 is required for normal growth and virulence. We show that Gib2 directly binds to Gpa1 and Gγ Gpg1/Gpg2 and that it interacts with phosphodiesterase Pde2 and monomeric GTPase Ras1. Pde2 remains functionally dispensable, but Ras1 is found to associate with adenylyl cyclase Cac1 through the conserved Ras association domain. In addition, the ras1 mutant exhibits normal capsule formation, whereas the ras1 gpa1 mutant displays enhanced capsule formation, and the ras1 gpa1 cac1 mutant is acapsular. Collectively, these findings suggest that Gib2 promotes cAMP levels by relieving an inhibitory function of Ras1 on Cac1 in the absence of Gpa1. In addition, using GST affinity purification combined with mass spectrometry, we identified 47 additional proteins that interact with Gib2. These proteins have putative functions ranging from signal transduction, energy generation, metabolism, and stress response to ribosomal function. After establishing and validating a protein-protein interactive network, we believe Gib2 to be a key adaptor/scaffolding protein that drives the formation of various protein complexes required for growth and virulence. Our study reveals Gib2 as an essential component in deciphering the complexity of regulatory networks that control growth and virulence in C. neoformans.

Context Specificity of Stress-activated Mitogen-activated Protein (MAP) Kinase Signaling: The Story as Told by Caenorhabditis elegans.

PubMed

Andrusiak, Matthew G; Jin, Yishi

2016-04-08

Stress-associated p38 and JNK mitogen-activated protein (MAP) kinase signaling cascades trigger specific cellular responses and are involved in multiple disease states. At the root of MAP kinase signaling complexity is the differential use of common components on a context-specific basis. The roundwormCaenorhabditis eleganswas developed as a system to study genes required for development and nervous system function. The powerful genetics ofC. elegansin combination with molecular and cellular dissections has led to a greater understanding of how p38 and JNK signaling affects many biological processes under normal and stress conditions. This review focuses on the studies revealing context specificity of different stress-activated MAPK components inC. elegans. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Effects of the space flight environment on man's immune system. II - Lymphocyte counts and reactivity.

NASA Technical Reports Server (NTRS)

Fischer, G. L.; Daniels, J. C.; Levin, W. C.; Kimzey, S. L.; Cobb, E. K.; Ritzmann, S. E.

1972-01-01

The present studies were undertaken to assess the effects of the environment of space flights on the cellular division of the human immune system. Peripheral blood absolute lymphocyte counts were determined at various preflight and postflight intervals for the 21 crewmen of Apollo Missions 7-13. Mean lymphocyte numbers tended to exhibit a delayed significant but fluctuating increase shortly after recovery, although a variety of responses was seen in individual astronauts. The in vitro reactivity of lymphocytes, reflected by RNA and DNA synthesis rates by unstimulated and PHA-stimulated lymphocytes tissue-cultured preflight and postflight from the same participants, was found to remain within previously established normal ranges. These results indicate that functional integrity of cellular immune potential as reflected by in vitro techniques is maintained during this spaceflight experience.

Enhanced Intracellular Delivery and Tissue Retention of Nanoparticles by Mussel-Inspired Surface Chemistry.

PubMed

Chen, Kai; Xu, Xiaoqiu; Guo, Jiawei; Zhang, Xuelin; Han, Songling; Wang, Ruibing; Li, Xiaohui; Zhang, Jianxiang

2015-11-09

Nanomaterials have been broadly studied for intracellular delivery of diverse compounds for diagnosis or therapy. Currently it remains challenging for discovering new biomolecules that can prominently enhance cellular internalization and tissue retention of nanoparticles (NPs). Herein we report for the first time that a mussel-inspired engineering approach may notably promote cellular uptake and tissue retention of NPs. In this strategy, the catechol moiety is covalently anchored onto biodegradable NPs. Thus, fabricated NPs can be more effectively internalized by sensitive and multidrug resistant tumor cells, as well as some normal cells, resulting in remarkably potentiated in vitro activity when an antitumor drug is packaged. Moreover, the newly engineered NPs afford increased tissue retention post local or oral delivery. This biomimetic approach is promising for creating functional nanomaterials for drug delivery, vaccination, and cell therapy.

Using precursor ion scan of 184 with liquid chromatography-electrospray ionization-tandem mass spectrometry for concentration normalization in cellular lipidomic studies.

PubMed

Chao, Hsi-Chun; Chen, Guan-Yuan; Hsu, Lih-Ching; Liao, Hsiao-Wei; Yang, Sin-Yu; Wang, San-Yuan; Li, Yu-Liang; Tang, Sung-Chun; Tseng, Yufeng Jane; Kuo, Ching-Hua

2017-06-08

Cellular lipidomic studies have been favored approaches in many biomedical research areas. To provide fair comparisons of the studied cells, it is essential to perform normalization of the determined concentration before lipidomic analysis. This study proposed a cellular lipidomic normalization method by measuring the phosphatidylcholine (PC) and sphingomyelin (SM) contents in cell extracts. To provide efficient analysis of PC and SM in cell extracts, flow injection analysis-electrospray ionization-tandem mass spectrometry (FIA-ESI-MS/MS) with a precursor ion scan (PIS) of m/z 184 was used, and the parameters affecting the performance of the method were optimized. Good linearity could be observed between the cell extract dilution factor and the reciprocal of the total ion chromatogram (TIC) area in the PIS of m/z 184 within the dilution range of 1- to 16-fold (R 2  = 0.998). The calibration curve could be used for concentration adjustment of the unknown concentration of a cell extract. The intraday and intermediate precisions were below 10%. The accuracy ranged from 93.0% to 105.6%. The performance of the new normalization method was evaluated using different numbers of HCT-116 cells. Sphingosine, ceramide (d18:1/18:0), SM (d18:1/18:0) and PC (16:1/18:0) were selected as the representative test lipid species, and the results showed that the peak areas of each lipid species obtained from different cell numbers were within a 20% variation after normalization. Finally, the PIS of 184 normalization method was applied to study ischemia-induced neuron injury using oxygen and glucose deprivation (OGD) on primary neuronal cultured cells. Our results showed that the PIS of 184 normalization method is an efficient and effective approach for concentration normalization in cellular lipidomic studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Hitting a Moving Target: Basic Mechanisms of Recovery from Acquired Developmental Brain Injury

PubMed Central

Giza, Christopher C.; Kolb, Bryan; Harris, Neil G.; Asarnow, Robert F.; Prins, Mayumi L.

2009-01-01

Acquired brain injuries represent a major cause of disability in the pediatric population. Understanding responses to developmental acquired brain injuries requires knowledge of the neurobiology of normal development, age-at-injury effects and experience-dependent neuroplasticity. In the developing brain, full recovery cannot be considered as a return to the premorbid baseline, since ongoing maturation means that cerebral functioning in normal individuals will continue to advance. Thus, the recovering immature brain has to ‘hit a moving target’ to achieve full functional recovery, defined as parity with age-matched uninjured peers. This review will discuss the consequences of developmental injuries such as focal lesions, diffuse hypoxia and traumatic brain injury (TBI). Underlying cellular and physiological mechanisms relevant to age-at-injury effects will be described in considerable detail, including but not limited to alterations in neurotransmission, connectivity/network functioning, the extracellular matrix, response to oxidative stress and changes in cerebral metabolism. Finally, mechanisms of experience-dependent plasticity will be reviewed in conjunction with their effects on neural repair and recovery. PMID:19956795

qpure: A Tool to Estimate Tumor Cellularity from Genome-Wide Single-Nucleotide Polymorphism Profiles

PubMed Central

Song, Sarah; Nones, Katia; Miller, David; Harliwong, Ivon; Kassahn, Karin S.; Pinese, Mark; Pajic, Marina; Gill, Anthony J.; Johns, Amber L.; Anderson, Matthew; Holmes, Oliver; Leonard, Conrad; Taylor, Darrin; Wood, Scott; Xu, Qinying; Newell, Felicity; Cowley, Mark J.; Wu, Jianmin; Wilson, Peter; Fink, Lynn; Biankin, Andrew V.; Waddell, Nic; Grimmond, Sean M.; Pearson, John V.

2012-01-01

Tumour cellularity, the relative proportion of tumour and normal cells in a sample, affects the sensitivity of mutation detection, copy number analysis, cancer gene expression and methylation profiling. Tumour cellularity is traditionally estimated by pathological review of sectioned specimens; however this method is both subjective and prone to error due to heterogeneity within lesions and cellularity differences between the sample viewed during pathological review and tissue used for research purposes. In this paper we describe a statistical model to estimate tumour cellularity from SNP array profiles of paired tumour and normal samples using shifts in SNP allele frequency at regions of loss of heterozygosity (LOH) in the tumour. We also provide qpure, a software implementation of the method. Our experiments showed that there is a medium correlation 0.42 (-value = 0.0001) between tumor cellularity estimated by qpure and pathology review. Interestingly there is a high correlation 0.87 (-value 2.2e-16) between cellularity estimates by qpure and deep Ion Torrent sequencing of known somatic KRAS mutations; and a weaker correlation 0.32 (-value = 0.004) between IonTorrent sequencing and pathology review. This suggests that qpure may be a more accurate predictor of tumour cellularity than pathology review. qpure can be downloaded from https://sourceforge.net/projects/qpure/. PMID:23049875

MicroRNA-7: A miRNA with expanding roles in development and disease.

PubMed

Horsham, Jessica L; Ganda, Clarissa; Kalinowski, Felicity C; Brown, Rikki A M; Epis, Michael R; Leedman, Peter J

2015-12-01

MicroRNAs (miRNAs) are a family of short, non-coding RNA molecules (∼22nt) involved in post-transcriptional control of gene expression. They act via base-pairing with mRNA transcripts that harbour target sequences, resulting in accelerated mRNA decay and/or translational attenuation. Given miRNAs mediate the expression of molecules involved in many aspects of normal cell development and functioning, it is not surprising that aberrant miRNA expression is closely associated with many human diseases. Their pivotal role in driving a range of normal cellular physiology as well as pathological processes has established miRNAs as potential therapeutics, as well as potential diagnostic and prognostic tools in human health. MicroRNA-7 (miR-7) is a highly conserved miRNA which displays restricted spatiotemporal expression during development and in maturity. In humans and mice, mature miR-7 is generated from three different genes, illustrating unexpected redundancy and also the importance of this miRNA in regulating key cellular processes. In this review we examine the expanding role of miR-7 in the context of health, with emphasis on organ differentiation and development, as well as in various mammalian diseases, particularly of the brain, heart, endocrine pancreas and skin, as well as in cancer. The more we learn about miR-7, the more we realise the complexity of its regulation and potential functional application both from a biomarker and therapeutic perspective. Copyright © 2015 Elsevier Ltd. All rights reserved.

Loss of lysophosphatidic acid receptor LPA1 alters oligodendrocyte differentiation and myelination in the mouse cerebral cortex.

PubMed

García-Díaz, Beatriz; Riquelme, Raquel; Varela-Nieto, Isabel; Jiménez, Antonio Jesús; de Diego, Isabel; Gómez-Conde, Ana Isabel; Matas-Rico, Elisa; Aguirre, José Ángel; Chun, Jerold; Pedraza, Carmen; Santín, Luis Javier; Fernández, Oscar; Rodríguez de Fonseca, Fernando; Estivill-Torrús, Guillermo

2015-11-01

Lysophosphatidic acid (LPA) is an intercellular signaling lipid that regulates multiple cellular functions, acting through specific G-protein coupled receptors (LPA(1-6)). Our previous studies using viable Malaga variant maLPA1-null mice demonstrated the requirement of the LPA1 receptor for normal proliferation, differentiation, and survival of the neuronal precursors. In the cerebral cortex LPA1 is expressed extensively in differentiating oligodendrocytes, in parallel with myelination. Although exogenous LPA-induced effects have been investigated in myelinating cells, the in vivo contribution of LPA1 to normal myelination remains to be demonstrated. This study identified a relevant in vivo role for LPA1 as a regulator of cortical myelination. Immunochemical analysis in adult maLPA1-null mice demonstrated a reduction in the steady-state levels of the myelin proteins MBP, PLP/DM20, and CNPase in the cerebral cortex. The myelin defects were confirmed using magnetic resonance spectroscopy and electron microscopy. Stereological analysis limited the defects to adult differentiating oligodendrocytes, without variation in the NG2+ precursor cells. Finally, a possible mechanism involving oligodendrocyte survival was demonstrated by the impaired intracellular transport of the PLP/DM20 myelin protein which was accompanied by cellular loss, suggesting stress-induced apoptosis. These findings describe a previously uncharacterized in vivo functional role for LPA1 in the regulation of oligodendrocyte differentiation and myelination in the CNS, underlining the importance of the maLPA1-null mouse as a model for the study of demyelinating diseases.

Generation and Characterization of an Immortalized Human Esophageal Myofibroblast Line.

PubMed

Niu, Chao; Chauhan, Uday; Gargus, Matthew; Shaker, Anisa

2016-01-01

Stromal cells with a myofibroblast phenotype present in the normal human esophagus are increased in individuals with gastro-esophageal reflux disease (GERD). We have previously demonstrated that myofibroblasts stimulated with acid and TLR4 agonists increase IL-6 and IL-8 secretion using primary cultures of myofibroblasts established from normal human esophagus. While primary cultures have the advantage of reflecting the in vivo environment, a short life span and unavoidable heterogeneity limits the usefulness of this model in larger scale in vitro cellular signaling studies. The major aim of this paper therefore was to generate a human esophageal myofibroblast line with an extended lifespan. In the work presented here we have generated and characterized an immortalized human esophageal myofibroblast line by transfection with a commercially available GFP-hTERT lentivirus. Immortalized human esophageal myofibroblasts demonstrate phenotypic, genotypic and functional similarity to primary cultures of esophageal myofibroblasts we have previously described. We found that immortalized esophageal myofibroblasts retain myofibroblast spindle-shaped morphology at low and high confluence beyond passage 80, and express α-SMA, vimentin, and CD90 myofibroblast markers. Immortalized human esophageal myofibroblasts also express the putative acid receptor TRPV1 and TLR4 and retain the functional capacity to respond to stimuli encountered in GERD with secretion of IL-6. Finally, immortalized human esophageal myofibroblasts also support the stratified growth of squamous esophageal epithelial cells in 3D organotypic cultures. This newly characterized immortalized human esophageal myofibroblast cell line can be used in future cellular signaling and co-culture studies.

Regulation of replicative senescence by NADP+ -dependent isocitrate dehydrogenase.

PubMed

Kil, In Sup; Huh, Tae Lin; Lee, Young Sup; Lee, You Mie; Park, Jeen-Woo

2006-01-01

The free radical hypothesis of aging postulates that senescence is due to an accumulation of cellular oxidative damage, caused largely by reactive oxygen species that are produced as by-products of normal metabolic processes. Recently, we demonstrated that the control of cytosolic and mitochondrial redox balance and the cellular defense against oxidative damage is one of the primary functions of cytosolic (IDPc) and mitochondrial NADP+ -dependent isocitrate dehydrogenase (IDPm) by supplying NADPH for antioxidant systems. In this paper, we demonstrate that modulation of IDPc or IDPm activity in IMR-90 cells regulates cellular redox status and replicative senescence. When we examined the regulatory role of IDPc and IDPm against the aging process with IMR-90 cells transfected with cDNA for IDPc or IDPm in sense and antisense orientations, a clear inverse relationship was observed between the amount of IDPc or IDPm expressed in target cells and their susceptibility to senescence, which was reflected by changes in replicative potential, cell cycle, senescence-associated beta-galactosidase activity, expression of p21 and p53, and morphology of cells. Furthermore, lipid peroxidation, oxidative DNA damage, and intracellular peroxide generation were higher and cellular redox status shifted to a prooxidant condition in the cell lines expressing the lower level of IDPc or IDPm. The results suggest that IDPc and IDPm play an important regulatory role in cellular defense against oxidative stress and in the senescence of IMR-90 cells.

Microfluidic device to control interstitial flow-mediated homotypic and heterotypic cellular communication.

PubMed

Alonzo, Luis F; Moya, Monica L; Shirure, Venktesh S; George, Steven C

2015-09-07

Tissue engineering can potentially recreate in vivo cellular microenvironments in vitro for an array of applications such as biological inquiry and drug discovery. However, the majority of current in vitro systems still neglect many biological, chemical, and mechanical cues that are known to impact cellular functions such as proliferation, migration, and differentiation. To address this gap, we have developed a novel microfluidic device that precisely controls the spatial and temporal interactions between adjacent three-dimensional cellular environments. The device consists of four interconnected microtissue compartments (~0.1 mm(3)) arranged in a square. The top and bottom pairs of compartments can be sequentially loaded with discrete cellularized hydrogels creating the opportunity to investigate homotypic (left to right or x-direction) and heterotypic (top to bottom or y-direction) cell-cell communication. A controlled hydrostatic pressure difference across the tissue compartments in both x and y direction induces interstitial flow and modulates communication via soluble factors. To validate the biological significance of this novel platform, we examined the role of stromal cells in the process of vasculogenesis. Our device confirms previous observations that soluble mediators derived from normal human lung fibroblasts (NHLFs) are necessary to form a vascular network derived from endothelial colony forming cell-derived endothelial cells (ECFC-ECs). We conclude that this platform could be used to study important physiological and pathological processes that rely on homotypic and heterotypic cell-cell communication.

Big-conductance Ca2+-activated K+ channels in physiological and pathophysiological urinary bladder smooth muscle cells

PubMed Central

Parajuli, Shankar P.; Zheng, Yun-Min; Levin, Robert; Wang, Yong-Xiao

2016-01-01

ABSTRACT Contraction and relaxation of urinary bladder smooth muscle cells (UBSMCs) represent the important physiological functions of the bladder. Contractile responses in UBSMCs are regulated by a number of ion channels including big-conductance Ca2+- activated K+ (BK) channels. Great progress has been made in studies of BK channels in UBSMCs. The intent of this review is to summarize recent exciting findings with respect to the functional interactions of BK channels with muscarinic receptors, ryanodine receptors (RyRs) and inositol triphosphate receptors (IP3Rs) as well as their functional importance under normal and pathophysiological conditions. BK channels are highly expressed in UBSMCs. Activation of muscarinic M3 receptors inhibits the BK channel activity, facilitates opening of voltage-dependent Ca2+ (CaV) channels, and thereby enhances excitability and contractility of UBSMCs. Signaling molecules and regulatory mechanisms involving RyRs and IP3Rs have a significant effect on functions of BK channels and thereby regulate cellular responses in UBSMCs under normal and pathophysiological conditions including overactive bladders. Moreover, BK channels may represent a novel target for the treatment of bladder dysfunctions. PMID:27101440

Coexpression network based on natural variation in human gene expression reveals gene interactions and functions

PubMed Central

Nayak, Renuka R.; Kearns, Michael; Spielman, Richard S.; Cheung, Vivian G.

2009-01-01

Genes interact in networks to orchestrate cellular processes. Analysis of these networks provides insights into gene interactions and functions. Here, we took advantage of normal variation in human gene expression to infer gene networks, which we constructed using correlations in expression levels of more than 8.5 million gene pairs in immortalized B cells from three independent samples. The resulting networks allowed us to identify biological processes and gene functions. Among the biological pathways, we found processes such as translation and glycolysis that co-occur in the same subnetworks. We predicted the functions of poorly characterized genes, including CHCHD2 and TMEM111, and provided experimental evidence that TMEM111 is part of the endoplasmic reticulum-associated secretory pathway. We also found that IFIH1, a susceptibility gene of type 1 diabetes, interacts with YES1, which plays a role in glucose transport. Furthermore, genes that predispose to the same diseases are clustered nonrandomly in the coexpression network, suggesting that networks can provide candidate genes that influence disease susceptibility. Therefore, our analysis of gene coexpression networks offers information on the role of human genes in normal and disease processes. PMID:19797678

Free radicals, reactive oxygen species, oxidative stress and its classification.

PubMed

Lushchak, Volodymyr I

2014-12-05

Reactive oxygen species (ROS) initially considered as only damaging agents in living organisms further were found to play positive roles also. This paper describes ROS homeostasis, principles of their investigation and technical approaches to investigate ROS-related processes. Especial attention is paid to complications related to experimental documentation of these processes, their diversity, spatiotemporal distribution, relationships with physiological state of the organisms. Imbalance between ROS generation and elimination in favor of the first with certain consequences for cell physiology has been called "oxidative stress". Although almost 30years passed since the first definition of oxidative stress was introduced by Helmut Sies, to date we have no accepted classification of oxidative stress. In order to fill up this gape here classification of oxidative stress based on its intensity is proposed. Due to that oxidative stress may be classified as basal oxidative stress (BOS), low intensity oxidative stress (LOS), intermediate intensity oxidative stress (IOS), and high intensity oxidative stress (HOS). Another classification of potential interest may differentiate three categories such as mild oxidative stress (MOS), temperate oxidative stress (TOS), and finally severe (strong) oxidative stress (SOS). Perspective directions of investigations in the field include development of sophisticated classification of oxidative stresses, accurate identification of cellular ROS targets and their arranged responses to ROS influence, real in situ functions and operation of so-called "antioxidants", intracellular spatiotemporal distribution and effects of ROS, deciphering of molecular mechanisms responsible for cellular response to ROS attacks, and ROS involvement in realization of normal cellular functions in cellular homeostasis. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

In vitro and in vivo assessment of lactic acid-modified chitosan scaffolds for potential treatment of full-thickness burns.

PubMed

Velasquillo, Cristina; Silva-Bermudez, Phaedra; Vázquez, Nadia; Martínez, Alan; Espadín, Andres; García-López, Julieta; Medina-Vega, Antonio; Lecona, Hugo; Pichardo-Baena, Raúl; Ibarra, Clemente; Shirai, Keiko

2017-10-01

Autologous skin transplantation is today's "gold standard" treatment for full-thickness burns. However, when > 30% of total body surface area is damaged, there is an important shortage of autologous donor sites for skin grafting; then, treatment alternatives become crucial. Such alternatives can be based on polymeric scaffolds capable of functioning as protective covers and cells/factors carriers. Chitosan (CTS) is a natural-derived polymer with relevant biological-related properties but poor mechanical performance. Improved mechanical properties can be achieved through lactic acid grafting (LA-g); nevertheless, LA-g affects the biological response towards the CTS-based materials. In this work, CTS-LA scaffolds with different LA-g percentages were synthesized and evaluated to determine appropriate LA-g degrees for full-thickness burns treatment. In vitro results indicated that the higher the LA-g percentage, the lower the capability of the scaffolds to sustain fibroblasts culture. Scaffolds with LA-g around 28% (CTS-LA28) sustained cell culture and allowed normal cell functionality. Further evaluation of CTS-LA28 as acellular and cellular grafts in a full-thickness burn mouse model showed that at 28 days post-burn, macroscopic characteristic of the reparation tissue were closer to healthy skin when cellular grafts were used for treatment; histological evaluation also showed that dermis cellularity and collagenous fibers structure were similar to those in healthy skin when cellular grafts were used for burns treatment. © 2017 Wiley Periodicals Inc. J Biomed Mater Res Part A: 105A: 2875-2891, 2017. © 2017 Wiley Periodicals, Inc.

Distinguishing between biochemical and cellular function: Are there peptide signatures for cellular function of proteins?

PubMed

Jain, Shruti; Bhattacharyya, Kausik; Bakshi, Rachit; Narang, Ankita; Brahmachari, Vani

2017-04-01

The genome annotation and identification of gene function depends on conserved biochemical activity. However, in the cell, proteins with the same biochemical function can participate in different cellular pathways and cannot complement one another. Similarly, two proteins of very different biochemical functions are put in the same class of cellular function; for example, the classification of a gene as an oncogene or a tumour suppressor gene is not related to its biochemical function, but is related to its cellular function. We have taken an approach to identify peptide signatures for cellular function in proteins with known biochemical function. ATPases as a test case, we classified ATPases (2360 proteins) and kinases (517 proteins) from the human genome into different cellular function categories such as transcriptional, replicative, and chromatin remodelling proteins. Using publicly available tool, MEME, we identify peptide signatures shared among the members of a given category but not between cellular functional categories; for example, no motif sharing is seen between chromatin remodelling and transporter ATPases, similarly between receptor Serine/Threonine Kinase and Receptor Tyrosine Kinase. There are motifs shared within each category with significant E value and high occurrence. This concept of signature for cellular function was applied to developmental regulators, the polycomb and trithorax proteins which led to the prediction of the role of INO80, a chromatin remodelling protein, in development. This has been experimentally validated earlier for its role in homeotic gene regulation and its interaction with regulatory complexes like the Polycomb and Trithorax complex. Proteins 2017; 85:682-693. © 2016 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Bioinformatics analysis reveals biophysical and evolutionary insights into the 3-nitrotyrosine post-translational modification in the human proteome

PubMed Central

Ng, John Y.; Boelen, Lies; Wong, Jason W. H.

2013-01-01

Protein 3-nitrotyrosine is a post-translational modification that commonly arises from the nitration of tyrosine residues. This modification has been detected under a wide range of pathological conditions and has been shown to alter protein function. Whether 3-nitrotyrosine is important in normal cellular processes or is likely to affect specific biological pathways remains unclear. Using GPS-YNO2, a recently described 3-nitrotyrosine prediction algorithm, a set of predictions for nitrated residues in the human proteome was generated. In total, 9.27 per cent of the proteome was predicted to be nitratable (27 922/301 091). By matching the predictions against a set of curated and experimentally validated 3-nitrotyrosine sites in human proteins, it was found that GPS-YNO2 is able to predict 73.1 per cent (404/553) of these sites. Furthermore, of these sites, 42 have been shown to be nitrated endogenously, with 85.7 per cent (36/42) of these predicted to be nitrated. This demonstrates the feasibility of using the predicted dataset for a whole proteome analysis. A comprehensive bioinformatics analysis was subsequently performed on predicted and all experimentally validated nitrated tyrosine. This found mild but specific biophysical constraints that affect the susceptibility of tyrosine to nitration, and these may play a role in increasing the likelihood of 3-nitrotyrosine to affect processes, including phosphorylation and DNA binding. Furthermore, examining the evolutionary conservation of predicted 3-nitrotyrosine showed that, relative to non-nitrated tyrosine residues, 3-nitrotyrosine residues are generally less conserved. This suggests that, at least in the majority of cases, 3-nitrotyrosine is likely to have a deleterious effect on protein function and less likely to be important in normal cellular function. PMID:23389939

Comparative RNA-Seq transcriptome analyses reveal distinct metabolic pathways in diabetic nerve and kidney disease.

PubMed

Hinder, Lucy M; Park, Meeyoung; Rumora, Amy E; Hur, Junguk; Eichinger, Felix; Pennathur, Subramaniam; Kretzler, Matthias; Brosius, Frank C; Feldman, Eva L

2017-09-01

Treating insulin resistance with pioglitazone normalizes renal function and improves small nerve fibre function and architecture; however, it does not affect large myelinated nerve fibre function in mouse models of type 2 diabetes (T2DM), indicating that pioglitazone affects the body in a tissue-specific manner. To identify distinct molecular pathways regulating diabetic peripheral neuropathy (DPN) and nephropathy (DN), as well those affected by pioglitazone, we assessed DPN and DN gene transcript expression in control and diabetic mice with or without pioglitazone treatment. Differential expression analysis and self-organizing maps were then used in parallel to analyse transcriptome data. Differential expression analysis showed that gene expression promoting cell death and the inflammatory response was reversed in the kidney glomeruli but unchanged or exacerbated in sciatic nerve by pioglitazone. Self-organizing map analysis revealed that mitochondrial dysfunction was normalized in kidney and nerve by treatment; however, conserved pathways were opposite in their directionality of regulation. Collectively, our data suggest inflammation may drive large fibre dysfunction, while mitochondrial dysfunction may drive small fibre dysfunction in T2DM. Moreover, targeting both of these pathways is likely to improve DN. This study supports growing evidence that systemic metabolic changes in T2DM are associated with distinct tissue-specific metabolic reprogramming in kidney and nerve and that these changes play a critical role in DN and small fibre DPN pathogenesis. These data also highlight the potential dangers of a 'one size fits all' approach to T2DM therapeutics, as the same drug may simultaneously alleviate one complication while exacerbating another. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Regulatory and effector functions of gamma-delta (γδ) T cells and their therapeutic potential in adoptive cellular therapy for cancer.

PubMed

Paul, Sourav; Lal, Girdhari

2016-09-01

γδ T cells are an important innate immune component of the tumor microenvironment and are known to affect the immune response in a wide variety of tumors. Unlike αβ T cells, γδ T cells are capable of spontaneous secretion of IL-17A and IFN-γ without undergoing clonal expansion. Although γδ T cells do not require self-MHC-restricted priming, they can distinguish "foreign" or transformed cells from healthy self-cells by using activating and inhibitory killer Ig-like receptors. γδ T cells were used in several clinical trials to treat cancer patient due to their MHC-unrestricted cytotoxicity, ability to distinguish transformed cells from normal cells, the capacity to secrete inflammatory cytokines and also their ability to enhance the generation of antigen-specific CD8(+) and CD4(+) T cell response. In this review, we discuss the effector and regulatory function of γδ T cells in the tumor microenvironment with special emphasis on the potential for their use in adoptive cellular immunotherapy. © 2016 UICC.

Laser ablation of Dbx1 neurons in the pre-Bötzinger complex stops inspiratory rhythm and impairs output in neonatal mice

PubMed Central

Wang, Xueying; Hayes, John A; Revill, Ann L; Song, Hanbing; Kottick, Andrew; Vann, Nikolas C; LaMar, M Drew; Picardo, Maria Cristina D; Akins, Victoria T; Funk, Gregory D; Del Negro, Christopher A

2014-01-01

To understand the neural origins of rhythmic behavior one must characterize the central pattern generator circuit and quantify the population size needed to sustain functionality. Breathing-related interneurons of the brainstem pre-Bötzinger complex (preBötC) that putatively comprise the core respiratory rhythm generator in mammals are derived from Dbx1-expressing precursors. Here, we show that selective photonic destruction of Dbx1 preBötC neurons in neonatal mouse slices impairs respiratory rhythm but surprisingly also the magnitude of motor output; respiratory hypoglossal nerve discharge decreased and its frequency steadily diminished until rhythm stopped irreversibly after 85±20 (mean ± SEM) cellular ablations, which corresponds to ∼15% of the estimated population. These results demonstrate that a single canonical interneuron class generates respiratory rhythm and contributes in a premotor capacity, whereas these functions are normally attributed to discrete populations. We also establish quantitative cellular parameters that govern network viability, which may have ramifications for respiratory pathology in disease states. DOI: http://dx.doi.org/10.7554/eLife.03427.001 PMID:25027440

Gold nanoparticles cellular toxicity and recovery: adipose Derived Stromal cells.

PubMed

Mironava, Tatsiana; Hadjiargyrou, Michael; Simon, Marcia; Rafailovich, Miriam H

2014-03-01

Gold nanoparticles (AuNPs) are currently used in numerous medical applications. Herein, we describe their in vitro impact on human adipose-derived stromal cells (ADSCs) using 13 nm and 45 nm citrate-coated AuNPs. In their non-differentiated state, ADSCs were penetrated by the AuNPs and stored in vacuoles. The presence of the AuNPs in ADSCs resulted in increased population doubling times, decreased cell motility and cell-mediated collagen contraction. The degree to which the cells were impacted was a function of particle concentration, where the smaller particles required a sevenfold higher concentration to have the same effect as the larger ones. Furthermore, AuNPs reduced adipogenesis as measured by lipid droplet accumulation and adiponectin secretion. These effects correlated with transient increases in DLK1 and with relative reductions in fibronectin. Upon removal of exogenous AuNPs, cellular NP levels decreased and normal ADSC functions were restored. As adiponectin helps regulate energy metabolism, local fluctuations triggered by AuNPs can lead to systemic changes. Hence, careful choice of size, concentration and clinical application duration of AuNPs is warranted.

Disruptive environmental chemicals and cellular mechanisms that confer resistance to cell death.

PubMed

Narayanan, Kannan Badri; Ali, Manaf; Barclay, Barry J; Cheng, Qiang Shawn; D'Abronzo, Leandro; Dornetshuber-Fleiss, Rita; Ghosh, Paramita M; Gonzalez Guzman, Michael J; Lee, Tae-Jin; Leung, Po Sing; Li, Lin; Luanpitpong, Suidjit; Ratovitski, Edward; Rojanasakul, Yon; Romano, Maria Fiammetta; Romano, Simona; Sinha, Ranjeet K; Yedjou, Clement; Al-Mulla, Fahd; Al-Temaimi, Rabeah; Amedei, Amedeo; Brown, Dustin G; Ryan, Elizabeth P; Colacci, Annamaria; Hamid, Roslida A; Mondello, Chiara; Raju, Jayadev; Salem, Hosni K; Woodrick, Jordan; Scovassi, A Ivana; Singh, Neetu; Vaccari, Monica; Roy, Rabindra; Forte, Stefano; Memeo, Lorenzo; Kim, Seo Yun; Bisson, William H; Lowe, Leroy; Park, Hyun Ho

2015-06-01

Cell death is a process of dying within biological cells that are ceasing to function. This process is essential in regulating organism development, tissue homeostasis, and to eliminate cells in the body that are irreparably damaged. In general, dysfunction in normal cellular death is tightly linked to cancer progression. Specifically, the up-regulation of pro-survival factors, including oncogenic factors and antiapoptotic signaling pathways, and the down-regulation of pro-apoptotic factors, including tumor suppressive factors, confers resistance to cell death in tumor cells, which supports the emergence of a fully immortalized cellular phenotype. This review considers the potential relevance of ubiquitous environmental chemical exposures that have been shown to disrupt key pathways and mechanisms associated with this sort of dysfunction. Specifically, bisphenol A, chlorothalonil, dibutyl phthalate, dichlorvos, lindane, linuron, methoxychlor and oxyfluorfen are discussed as prototypical chemical disruptors; as their effects relate to resistance to cell death, as constituents within environmental mixtures and as potential contributors to environmental carcinogenesis. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Expression of different functional isoforms in haematopoiesis.

PubMed

Grech, Godfrey; Pollacco, Joel; Portelli, Mark; Sacco, Keith; Baldacchino, Shawn; Grixti, Justine; Saliba, Christian

2014-01-01

Haematopoiesis is a complex process regulated at various levels facilitating rapid responses to external factors including stress, modulation of lineage commitment and terminal differentiation of progenitors. Although the transcription program determines the RNA pool of a cell, various mRNA strands can be obtained from the same template, giving rise to multiple protein isoforms. The majority of variants and isoforms co-occur in normal haematopoietic cells or are differentially expressed at various maturity stages of progenitor maturation and cellular differentiation within the same lineage or across lineages. Genetic aberrations or specific cellular states result in the predominant expression of abnormal isoforms leading to deregulation and disease. The presence of upstream open reading frames (uORF) in 5' untranslated regions (UTRs) of a transcript, couples the utilization of start codons with the cellular status and availability of translation initiation factors (eIFs). In addition, tissue-specific and cell lineage-specific alternative promoter use, regulates several transcription factors producing transcript variants with variable 5' exons. In this review, we propose to give a detailed account of the differential isoform formation, causing haematological malignancies.

Surface-anchored poly(acryloyl-L(D)-valine) with enhanced chirality-selective effect on cellular uptake of gold nanoparticles

PubMed Central

Deng, Jun; Wu, Sai; Yao, Mengyun; Gao, Changyou

2016-01-01

Chirality is one of the ubiquitous phenomena in biological systems. The left handed (L-) amino acids and right handed (D-) sugars are normally found in proteins, and in RNAs and DNAs, respectively. The effect of chiral surfaces at the nanoscale on cellular uptake has, however, not been explored. This study reveals for the first time the molecular chirality on gold nanoparticles (AuNPs) functions as a direct regulator for cellular uptake. Monolayers of 2-mercaptoacetyl-L(D)-valine (L(D)-MAV) and poly(acryloyl-L(D)-valine (L(D)-PAV) chiral molecules were formed on AuNPs surface, respectively. The internalized amount of PAV-AuNPs was several times larger than that of MAV-AuNPs by A549 and HepG2 cells, regardless of the chirality difference. However, the D-PAV-AuNPs were internalized with significantly larger amount than the L-PAV-AuNPs. This chirality-dependent uptake effect is likely attributed to the preferable interaction between the L-phospholipid-based cell membrane and the D-enantiomers. PMID:27531648

Surface-anchored poly(acryloyl-L(D)-valine) with enhanced chirality-selective effect on cellular uptake of gold nanoparticles

NASA Astrophysics Data System (ADS)

Deng, Jun; Wu, Sai; Yao, Mengyun; Gao, Changyou

2016-08-01

Chirality is one of the ubiquitous phenomena in biological systems. The left handed (L-) amino acids and right handed (D-) sugars are normally found in proteins, and in RNAs and DNAs, respectively. The effect of chiral surfaces at the nanoscale on cellular uptake has, however, not been explored. This study reveals for the first time the molecular chirality on gold nanoparticles (AuNPs) functions as a direct regulator for cellular uptake. Monolayers of 2-mercaptoacetyl-L(D)-valine (L(D)-MAV) and poly(acryloyl-L(D)-valine (L(D)-PAV) chiral molecules were formed on AuNPs surface, respectively. The internalized amount of PAV-AuNPs was several times larger than that of MAV-AuNPs by A549 and HepG2 cells, regardless of the chirality difference. However, the D-PAV-AuNPs were internalized with significantly larger amount than the L-PAV-AuNPs. This chirality-dependent uptake effect is likely attributed to the preferable interaction between the L-phospholipid-based cell membrane and the D-enantiomers.

Surface-anchored poly(acryloyl-L(D)-valine) with enhanced chirality-selective effect on cellular uptake of gold nanoparticles.

PubMed

Deng, Jun; Wu, Sai; Yao, Mengyun; Gao, Changyou

2016-08-17

Chirality is one of the ubiquitous phenomena in biological systems. The left handed (L-) amino acids and right handed (D-) sugars are normally found in proteins, and in RNAs and DNAs, respectively. The effect of chiral surfaces at the nanoscale on cellular uptake has, however, not been explored. This study reveals for the first time the molecular chirality on gold nanoparticles (AuNPs) functions as a direct regulator for cellular uptake. Monolayers of 2-mercaptoacetyl-L(D)-valine (L(D)-MAV) and poly(acryloyl-L(D)-valine (L(D)-PAV) chiral molecules were formed on AuNPs surface, respectively. The internalized amount of PAV-AuNPs was several times larger than that of MAV-AuNPs by A549 and HepG2 cells, regardless of the chirality difference. However, the D-PAV-AuNPs were internalized with significantly larger amount than the L-PAV-AuNPs. This chirality-dependent uptake effect is likely attributed to the preferable interaction between the L-phospholipid-based cell membrane and the D-enantiomers.

Microtubule-Actin Cross-Linking Factor 1: Domains, Interaction Partners, and Tissue-Specific Functions.

PubMed

Goryunov, Dmitry; Liem, Ronald K H

2016-01-01

The cytoskeleton of most eukaryotic cells is composed of three principal filamentous components: actin filaments, microtubules (MTs), and intermediate filaments. It is a highly dynamic system that plays crucial roles in a wide range of cellular processes, including migration, adhesion, cytokinesis, morphogenesis, intracellular traffic and signaling, and structural flexibility. Among the large number of cytoskeleton-associated proteins characterized to date, microtubule-actin cross-linking factor 1 (MACF1) is arguably the most versatile integrator and modulator of cytoskeleton-related processes. MACF1 belongs to the plakin family of proteins, and within it, to the spectraplakin subfamily. These proteins are characterized by the ability to bridge MT and actin cytoskeletal networks in a dynamic fashion, which underlies their involvement in the regulation of cell migration, axonal extension, and vesicular traffic. Studying MACF1 functions has provided insights not only into the regulation of the cytoskeleton but also into molecular mechanisms of both normal cellular physiology and cellular pathology. Multiple MACF1 isoforms exist, composed of a large variety of alternatively spliced domains. Each of these domains mediates a specific set of interactions and functions. These functions are manifested in tissue and cell-specific phenotypes observed in conditional MACF1 knockout mice. The conditional models described to date reveal critical roles of MACF1 in mammalian skin, nervous system, heart muscle, and intestinal epithelia. Complete elimination of MACF1 is early embryonic lethal, indicating an essential role for MACF1 in early development. Further studies of MACF1 domains and their interactions will likely reveal multiple new roles of this protein in various tissues. © 2016 Elsevier Inc. All rights reserved.

Pyridostigmine bromide (PYR) alters immune function in B6C3F1 mice.

PubMed

Peden-Adams, M M; Dudley, A C; EuDaly, J G; Allen, C T; Gilkeson, G S; Keil, D E

2004-02-01

Pyridostigmine bromide (PYR) is an anticholinesterase drug indicated for the treatment of myasthenia gravis and neuromuscular blockade reversal. It acts as a reversible cholinesterase inhibitor and was used as a pretreatment for soldiers during Operation Desert Storm to protect against possible nerve gas attacks. Since that time, PYR has been implicated as a possible causative agent contributing to Gulf War Illness. PYR's mechanism of action has been well-delineated with regards to its effects on the nervous system, yet little is known regarding potential effects on immunological function. To evaluate the effects of PYR on immunological function, adult female B6C3F1 mice were gavaged daily for 14 days with PYR (0, 1, 5, 10, or 20 mg/kg/day). Immune parameters assessed were lymphoproliferation, natural killer cell activity, the SRBC-specific antibody plaque-forming cell (PFC) response, thymus and spleen weight and cellularity, and thymic and splenic CD4/CD8 lymphocyte subpopulations. Exposure to PYR did not alter splenic and thymus weight or splenic cellularity. However, 20 mg PYR/kg/day decreased thymic cellularity with decreases in both CD4+/CD8+ (20 mg/kg/day) and CD4-/CD8- (10 and 20 mg/kg/day) cell types. Functional immune assays indicated that lymphocyte proliferative responses and natural killer cell activity were normal; whereas exposure to PYR significantly decreased primary IgM antibody responses to a T-cell dependent antigen at the 1, 5, 10 and 20 mg/kg treatment levels for 14 days. This is the first study to examine the immunotoxicological effects of PYR and demonstrate that this compound selectively suppresses humoral antibody responses.

Ablation of ceramide synthase 2 exacerbates dextran sodium sulphate-induced colitis in mice due to increased intestinal permeability.

PubMed

Kim, Ye-Ryung; Volpert, Giora; Shin, Kyong-Oh; Kim, So-Yeon; Shin, Sun-Hye; Lee, Younghay; Sung, Sun Hee; Lee, Yong-Moon; Ahn, Jung-Hyuck; Pewzner-Jung, Yael; Park, Woo-Jae; Futerman, Anthony H; Park, Joo-Won

2017-12-01

Ceramides mediate crucial cellular processes including cell death and inflammation and have recently been implicated in inflammatory bowel disease. Ceramides consist of a sphingoid long-chain base to which fatty acids of various length can be attached. We now investigate the effect of alerting the ceramide acyl chain length on a mouse model of colitis. Ceramide synthase (CerS) 2 null mice, which lack very-long acyl chain ceramides with concomitant increase of long chain bases and C16-ceramides, were more susceptible to dextran sodium sulphate-induced colitis, and their survival rate was markedly decreased compared with that of wild-type littermates. Using mixed bone-marrow chimeric mice, we showed that the host environment is primarily responsible for intestinal barrier dysfunction and increased intestinal permeability. In the colon of CerS2 null mice, the expression of junctional adhesion molecule-A was markedly decreased and the phosphorylation of myosin light chain 2 was increased. In vitro experiments using Caco-2 cells also confirmed an important role of CerS2 in maintaining epithelial barrier function; CerS2-knockdown via CRISPR-Cas9 technology impaired barrier function. In vivo myriocin administration, which normalized long-chain bases and C16-ceramides of the colon of CerS2 null mice, increased intestinal permeability as measured by serum FITC-dextran levels, indicating that altered SLs including deficiency of very-long-chain ceramides are critical for epithelial barrier function. In conclusion, deficiency of CerS2 influences intestinal barrier function and the severity of experimental colitis and may represent a potential mechanism for inflammatory bowel disease pathogenesis. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Effect of compound glycyrrhizin injection on liver function and cellular immunity of children with infectious mononucleosis complicated liver impairment.

PubMed

Cao, Zong-xin; Zhao, Zhong-fang; Zhao, Xiu-fen

2006-12-01

To investigate the effects of Compound Glycyrrhizin Injection (CGI) on liver function and cellular immunity of children with infectious mononucleosis complicated liver impairment (IM-LI) and to explore its clinical therapeutic effect. Forty-two patients with IM-LI were randomly assigned, according to the randomizing number table, to two groups, 20 in the control group and 22 in the treated group. All the patients were treated with conventional treatment, but to those in the treated group, CGI was given additionally once a day, at the dosage of 10 ml for children aged below 2 years, 20 ml for 2-4 years old, 30 ml for 5-7 years old and 40 ml for 8- 12 years old, in 100-200 ml of 5% glucose solution by intravenous dripping. The treatment lasted for 2 weeks. T lymphocyte subsets and serum levels of alanine transaminase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBil) were detected before and after treatment. Besides, a normal control group consisting of 20 healthy children was also set up. Baseline of the percentage of CD3 + , CD8 + lymphocyte and serum levels of ALT, AST, TBiL in the children with IM-LI were markedly higher, while the percentage of CD4 + lymphocyte and the CD4 + /CD8 + ratio was markedly lower in IM-LI children as compared with the corresponding indices in the healthy children ( P<0.01). These indices were improved after treatment in both groups of patients, but the improvement in the treated group was better than that in the control group (P<0.01). Cellular immunity dysfunction often occurs in patients with IM-LI, and CGI treatment can not only obviously promote the recovery of liver function, but also regulate the immune function in organism.

Tau and spectraplakins promote synapse formation and maintenance through Jun kinase and neuronal trafficking

PubMed Central

Voelzmann, Andre; Okenve-Ramos, Pilar; Qu, Yue; Chojnowska-Monga, Monika; del Caño-Espinel, Manuela; Prokop, Andreas; Sanchez-Soriano, Natalia

2016-01-01

The mechanisms regulating synapse numbers during development and ageing are essential for normal brain function and closely linked to brain disorders including dementias. Using Drosophila, we demonstrate roles of the microtubule-associated protein Tau in regulating synapse numbers, thus unravelling an important cellular requirement of normal Tau. In this context, we find that Tau displays a strong functional overlap with microtubule-binding spectraplakins, establishing new links between two different neurodegenerative factors. Tau and the spectraplakin Short Stop act upstream of a three-step regulatory cascade ensuring adequate delivery of synaptic proteins. This cascade involves microtubule stability as the initial trigger, JNK signalling as the central mediator, and kinesin-3 mediated axonal transport as the key effector. This cascade acts during development (synapse formation) and ageing (synapse maintenance) alike. Therefore, our findings suggest novel explanations for intellectual disability in Tau deficient individuals, as well as early synapse loss in dementias including Alzheimer’s disease. DOI: http://dx.doi.org/10.7554/eLife.14694.001 PMID:27501441

The Neurotensin Receptor-1 Pathway Contributes to Human Ductal Breast Cancer Progression

PubMed Central

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian

2009-01-01

Background The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. Methods and Results we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. Conclusion these data support the activation of neurotensinergic deleterious pathways in breast cancer progression. PMID:19156213

The neurotensin receptor-1 pathway contributes to human ductal breast cancer progression.

PubMed

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian; Gompel, Anne; Forgez, Patricia

2009-01-01

The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. these data support the activation of neurotensinergic deleterious pathways in breast cancer progression.

The Genetics and Epigenetics of Kidney Development

PubMed Central

Patel, Sanjeevkumar R.; Dressler, Gregory R.

2013-01-01

The development of the mammalian kidney has been studied at the genetic, biochemical, and cell biological level for more than 40 years. As such, detailed mechanisms governing early patterning, cell lineages, and inductive interactions are well described. How genes interact to specify the renal epithelial cells of the nephrons and how this specification is relevant to maintaining normal renal function is discussed. Implicit in the development of the kidney are epigenetic mechanisms that mark renal cell types and connect certain developmental regulatory factors to chromatin modifications that control gene expression patterns and cellular physiology. In adults, such regulatory factors and their epigenetic pathways may function in regeneration and may be disturbed in disease processes. PMID:24011574

Expression of Ras-related C3 botulinum toxin substrate 1 (RAC1) in human cholesteatoma.

PubMed

Lee, No Hee; Chang, Ji-Won; Choi, June; Jung, Hak Hyun; Im, Gi Jung

2013-02-01

Ras-related C3 botulinum toxin substrate 1 (RAC1) is a 21-kDa signaling G protein that functions as a pleiotropic regulator of many cellular processes including epithelial differentiation. RAC1 activates the nicotinamide adenine dinucleotide phosphate oxidase complex which promotes formation of reactive oxygen species and degradation enzymes. RAC1 has been associated with rapid epithelial differentiation and invasive properties in human cholesteatoma. This study aimed to identify the presence of RAC1 in human cholesteatoma and analyze its functional role as a regulator of proteolysis and overgrowth. Tissue samples from human cholesteatoma and normal postaural skin were obtained from patients during otologic surgery for cholesteatoma. The expression of RAC1 mRNA was quantified by real-time RT-PCR, and localization of RAC1 expression was confirmed using immunohistochemical staining. Expression of RAC1 mRNA in the epithelium of cholesteatoma was significantly elevated 2.94 fold on average, compared with normal control skin. RAC1 expression in the suprabasal and basal layer of cholesteatoma epithelium was stronger than normal control skin. Our results suggest that RAC1 can be associated with rapid epithelial differentiation and invasive properties of human cholesteatoma.

A crucial role of ROCK for alleviation of senescence-associated phenotype.

PubMed

Park, Joon Tae; Kang, Hyun Tae; Park, Chi Hyun; Lee, Young-Sam; Cho, Kyung A; Park, Sang Chul

2018-06-01

In our previous study, we uncovered a novel mechanism in which amelioration of Hutchinson-Gilford progeria syndrome (HGPS) phenotype is mediated by mitochondrial functional recovery upon rho-associated protein kinase (ROCK) inhibition. However, it remains elusive whether this mechanism is also applied to the amelioration of normal aging cells. In this study, we used Y-27632 and fasudil as effective ROCK inhibitors, and examined their role in senescence. We found that ROCK inhibition induced the functional recovery of the mitochondria as well as the metabolic reprogramming, which are two salient features that are altered in normal aging cells. Moreover, microarray analysis revealed that the up-regulated pathway upon ROCK inhibition is enriched for chromatin remodeling genes, which may play an important role in the alleviation of senescence-associated cell cycle arrest. Indeed, ROCK inhibition induced cellular proliferation, concomitant with the amelioration of senescent phenotype. Furthermore, the restorative effect by ROCK inhibition was observed in vivo as evidenced by the facilitated cutaneous wound healing. Taken together, our data indicate that ROCK inhibition might be utilized to ameliorate normal aging process and to treat age-related disease. Copyright © 2018 Elsevier Inc. All rights reserved.

Evidence that the tri-cellular metabolism of N-acetylaspartate functions as the brain's "operating system": how NAA metabolism supports meaningful intercellular frequency-encoded communications.

PubMed

Baslow, Morris H

2010-11-01

N-acetylaspartate (NAA), an acetylated derivative of L-aspartate (Asp), and N-acetylaspartylglutamate (NAAG), a derivative of NAA and L-glutamate (Glu), are synthesized by neurons in brain. However, neurons cannot catabolize either of these substances, and so their metabolism requires the participation of two other cell types. Neurons release both NAA and NAAG to extra-cellular fluid (ECF) upon stimulation, where astrocytes, the target cells for NAAG, hydrolyze it releasing NAA back into ECF, and oligodendrocytes, the target cells for NAA, hydrolyze it releasing Asp to ECF for recycling to neurons. This sequence is unique as it is the only known amino acid metabolic cycle in brain that requires three cell types for its completion. The results of this cycling are two-fold. First, neuronal metabolic water is transported to ECF for its removal from brain. Second, the rate of neuronal activity is coupled with focal hyperemia, providing stimulated neurons with the energy required for transmission of meaningful frequency-encoded messages. In this paper, it is proposed that the tri-cellular metabolism of NAA functions as the "operating system" of the brain, and is essential for normal cognitive and motor activities. Evidence in support of this hypothesis is provided by the outcomes of two human inborn errors in NAA metabolism.

Cell-autonomous correction of ring chromosomes in human induced pluripotent stem cells

NASA Astrophysics Data System (ADS)

Bershteyn, Marina; Hayashi, Yohei; Desachy, Guillaume; Hsiao, Edward C.; Sami, Salma; Tsang, Kathryn M.; Weiss, Lauren A.; Kriegstein, Arnold R.; Yamanaka, Shinya; Wynshaw-Boris, Anthony

2014-03-01

Ring chromosomes are structural aberrations commonly associated with birth defects, mental disabilities and growth retardation. Rings form after fusion of the long and short arms of a chromosome, and are sometimes associated with large terminal deletions. Owing to the severity of these large aberrations that can affect multiple contiguous genes, no possible therapeutic strategies for ring chromosome disorders have been proposed. During cell division, ring chromosomes can exhibit unstable behaviour leading to continuous production of aneuploid progeny with low viability and high cellular death rate. The overall consequences of this chromosomal instability have been largely unexplored in experimental model systems. Here we generated human induced pluripotent stem cells (iPSCs) from patient fibroblasts containing ring chromosomes with large deletions and found that reprogrammed cells lost the abnormal chromosome and duplicated the wild-type homologue through the compensatory uniparental disomy (UPD) mechanism. The karyotypically normal iPSCs with isodisomy for the corrected chromosome outgrew co-existing aneuploid populations, enabling rapid and efficient isolation of patient-derived iPSCs devoid of the original chromosomal aberration. Our results suggest a fundamentally different function for cellular reprogramming as a means of `chromosome therapy' to reverse combined loss-of-function across many genes in cells with large-scale aberrations involving ring structures. In addition, our work provides an experimentally tractable human cellular system for studying mechanisms of chromosomal number control, which is of critical relevance to human development and disease.

Progressing neurobiological strategies against proteostasis failure: Challenges in neurodegeneration.

PubMed

Amanullah, Ayeman; Upadhyay, Arun; Joshi, Vibhuti; Mishra, Ribhav; Jana, Nihar Ranjan; Mishra, Amit

2017-12-01

Proteins are ordered useful cellular entities, required for normal health and organism's survival. The proteome is the absolute set of cellular expressed proteins, which regulates a wide range of physiological functions linked with all domains of life. In aging cells or under unfavorable cellular conditions, misfolding of proteins generates common pathological events linked with neurodegenerative diseases and aging. Current advances of proteome studies systematically generates some progress in our knowledge that how misfolding of proteins or their accumulation can contribute to the impairment or depletion of proteome functions. Still, the underlying causes of this unrecoverable loss are not clear that how such unsolved transitions give rise to multifactorial challengeable degenerative pathological conditions in neurodegeneration. In this review, we specifically focus and systematically summarize various molecular mechanisms of proteostasis maintenance, as well as discuss progressing neurobiological strategies, promising natural and pharmacological candidates, which can be useful to counteract the problem of proteopathies. Our article emphasizes an urgent need that now it is important for us to recognize the fundamentals of proteostasis to design a new molecular framework and fruitful strategies to uncover how the proteome defects are associated with aging and neurodegenerative diseases. A enhance understanding of progress link with proteome and neurobiological challenges may provide new basic concepts in the near future, based on pharmacological agents, linked with impaired proteostasis and neurodegenerative diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

Prenatal alcohol exposure and cellular differentiation: a role for Polycomb and Trithorax group proteins in FAS phenotypes?

PubMed

Veazey, Kylee J; Muller, Daria; Golding, Michael C

2013-01-01

Exposure to alcohol significantly alters the developmental trajectory of progenitor cells and fundamentally compromises tissue formation (i.e., histogenesis). Emerging research suggests that ethanol can impair mammalian development by interfering with the execution of molecular programs governing differentiation. For example, ethanol exposure disrupts cellular migration, changes cell-cell interactions, and alters growth factor signaling pathways. Additionally, ethanol can alter epigenetic mechanisms controlling gene expression. Normally, lineage-specific regulatory factors (i.e., transcription factors) establish the transcriptional networks of each new cell type; the cell's identity then is maintained through epigenetic alterations in the way in which the DNA encoding each gene becomes packaged within the chromatin. Ethanol exposure can induce epigenetic changes that do not induce genetic mutations but nonetheless alter the course of fetal development and result in a large array of patterning defects. Two crucial enzyme complexes--the Polycomb and Trithorax proteins--are central to the epigenetic programs controlling the intricate balance between self-renewal and the execution of cellular differentiation, with diametrically opposed functions. Prenatal ethanol exposure may disrupt the functions of these two enzyme complexes, altering a crucial aspect of mammalian differentiation. Characterizing the involvement of Polycomb and Trithorax group complexes in the etiology of fetal alcohol spectrum disorders will undoubtedly enhance understanding of the role that epigenetic programming plays in this complex disorder.

Establishment of a Mouse Model with Misregulated Chromosome Condensation due to Defective Mcph1 Function

PubMed Central

Walther, Diego J.; Dopatka, Monika; Dutrannoy, Véronique; Busche, Andreas; Meyer, Franziska; Nowak, Stefanie; Nowak, Jean; Zabel, Claus; Klose, Joachim; Esquitino, Veronica; Garshasbi, Masoud; Kuss, Andreas W.; Ropers, Hans-Hilger; Mueller, Susanne; Poehlmann, Charlotte; Gavvovidis, Ioannis; Schindler, Detlev; Sperling, Karl; Neitzel, Heidemarie

2010-01-01

Mutations in the human gene MCPH1 cause primary microcephaly associated with a unique cellular phenotype with premature chromosome condensation (PCC) in early G2 phase and delayed decondensation post-mitosis (PCC syndrome). The gene encodes the BRCT-domain containing protein microcephalin/BRIT1. Apart from its role in the regulation of chromosome condensation, the protein is involved in the cellular response to DNA damage. We report here on the first mouse model of impaired Mcph1-function. The model was established based on an embryonic stem cell line from BayGenomics (RR0608) containing a gene trap in intron 12 of the Mcph1 gene deleting the C-terminal BRCT-domain of the protein. Although residual wild type allele can be detected by quantitative real-time PCR cell cultures generated from mouse tissues bearing the homozygous gene trap mutation display the cellular phenotype of misregulated chromosome condensation that is characteristic for the human disorder, confirming defective Mcph1 function due to the gene trap mutation. While surprisingly the DNA damage response (formation of repair foci, chromosomal breakage, and G2/M checkpoint function after irradiation) appears to be largely normal in cell cultures derived from Mcph1gt/gt mice, the overall survival rates of the Mcph1gt/gt animals are significantly reduced compared to wild type and heterozygous mice. However, we could not detect clear signs of premature malignant disease development due to the perturbed Mcph1 function. Moreover, the animals show no obvious physical phenotype and no reduced fertility. Body and brain size are within the range of wild type controls. Gene expression on RNA and protein level did not reveal any specific pattern of differentially regulated genes. To the best of our knowledge this represents the first mammalian transgenic model displaying a defect in mitotic chromosome condensation and is also the first mouse model for impaired Mcph1-function. PMID:20169082

Establishment of a mouse model with misregulated chromosome condensation due to defective Mcph1 function.

PubMed

Trimborn, Marc; Ghani, Mahdi; Walther, Diego J; Dopatka, Monika; Dutrannoy, Véronique; Busche, Andreas; Meyer, Franziska; Nowak, Stefanie; Nowak, Jean; Zabel, Claus; Klose, Joachim; Esquitino, Veronica; Garshasbi, Masoud; Kuss, Andreas W; Ropers, Hans-Hilger; Mueller, Susanne; Poehlmann, Charlotte; Gavvovidis, Ioannis; Schindler, Detlev; Sperling, Karl; Neitzel, Heidemarie

2010-02-16

Mutations in the human gene MCPH1 cause primary microcephaly associated with a unique cellular phenotype with premature chromosome condensation (PCC) in early G2 phase and delayed decondensation post-mitosis (PCC syndrome). The gene encodes the BRCT-domain containing protein microcephalin/BRIT1. Apart from its role in the regulation of chromosome condensation, the protein is involved in the cellular response to DNA damage. We report here on the first mouse model of impaired Mcph1-function. The model was established based on an embryonic stem cell line from BayGenomics (RR0608) containing a gene trap in intron 12 of the Mcph1 gene deleting the C-terminal BRCT-domain of the protein. Although residual wild type allele can be detected by quantitative real-time PCR cell cultures generated from mouse tissues bearing the homozygous gene trap mutation display the cellular phenotype of misregulated chromosome condensation that is characteristic for the human disorder, confirming defective Mcph1 function due to the gene trap mutation. While surprisingly the DNA damage response (formation of repair foci, chromosomal breakage, and G2/M checkpoint function after irradiation) appears to be largely normal in cell cultures derived from Mcph1(gt/gt) mice, the overall survival rates of the Mcph1(gt/gt) animals are significantly reduced compared to wild type and heterozygous mice. However, we could not detect clear signs of premature malignant disease development due to the perturbed Mcph1 function. Moreover, the animals show no obvious physical phenotype and no reduced fertility. Body and brain size are within the range of wild type controls. Gene expression on RNA and protein level did not reveal any specific pattern of differentially regulated genes. To the best of our knowledge this represents the first mammalian transgenic model displaying a defect in mitotic chromosome condensation and is also the first mouse model for impaired Mcph1-function.

TWO INNEXINS OF Spodoptera litura INFLUENCES HEMICHANNEL AND GAP JUNCTION FUNCTIONS IN CELLULAR IMMUNE RESPONSES.

PubMed

Pang, Zunyu; Li, Ming; Yu, Dongshuai; Yan, Zhang; Liu, Xinyi; Ji, Xinglai; Yang, Yang; Hu, Jiansheng; Luo, Kaijun

2015-09-01

Insect cellular immune responses include encapsulation, nodule formation, and phagocytosis. Hemichannels and gap junctions are involved in these cellular actions. Innexins (Inxs: analogous to the vertebrate connexins) form hemichannels and gap junctions, but the molecular mechanisms underlying their biology is still unclear. In this article, we reported a steady-state level of Inxs (SpliInxs) in hemocytes of Spodoptera litura, which formed nonfunctional hemichannels on the cell surface to maintain normal metabolism. We also reported that two innnexins (SpliInx2 and SpliInx3) were expressed significantly higher in hemocytes compared to other tissues, suggesting that they play important roles in hemocytes. Amino acid analysis found that two cysteine residues in two extracellular loops provided the capability for SpliInx2 and SpliInx3 hemichannels to dock into gap junctions. Western blotting demonstrated that both extracellular and intracellular loops of SpliInx3 and the extracellular loops of SpliInx2 might undergo posttranslational modification during the formation of a steady-state hemichannel. During hemichannel formation, SpliInx2 presented as one isoform, while SpliInx3 presented as three isoforms. These results provide fundamental knowledge for further study of how steady-state levels of SpliInxs are dynamically adjusted to perform cellular immune responses under immune challenge. © 2015 Wiley Periodicals, Inc.

Investigation of phosphoserine and cytidine 5'-phosphate by heteronuclear two-dimensional spectroscopy: samples with strong proton coupling

NASA Astrophysics Data System (ADS)

Bolton, Philip H.

Heteronuclear two-dimensional magnetic resonance is a novel method for investigating the conformations of cellular phosphates. The two-dimensional proton spectra are detected indirectly via the phosphorus-31 nucleus and thus allow determination of proton chemical shifts and coupling constants in situations in which the normal proton spectrum is obscured. Previous investigations of cellular phosphates with relatively simple spin systems have shown that the two-dimensional proton spectrum can be readily related to the normal proton spectrum by subspectral analysis. The normal proton spectrum can be decomposed into two subspectra, one for each polarization of the phosphorus-31 nucleus. The two-dimensional spectrum arises from the difference between the subspectra, and the normal proton spectrum is the sum. This allows simulation of the two-dimensional spectra and hence determination of the proton chemical shifts and coupling constants. Many cellular phosphates of interest, such as 5'-nucleotides and phosphoserine, contain three protons coupled to the phosphorus which are strongly coupled to one another. These samples are amenable to the two-dimensional method and the straightforward subspectral analysis is preserved when a 90° pulse is applied to the protons in the magnetization transfer step. The two-dimensional proton spectra of the samples investigated here have higher resolution than the normal proton spectra, revealing spectral features not readily apparent in the normal proton spectra.

Tribbles in normal and malignant haematopoiesis.

PubMed

Stein, Sarah J; Mack, Ethan A; Rome, Kelly S; Pear, Warren S

2015-10-01

The tribbles protein family, an evolutionarily conserved group of pseudokinases, have been shown to regulate multiple cellular events including those involved in normal and malignant haematopoiesis. The three mammalian Tribbles homologues, Trib1, Trib2 and Trib3 are characterized by conserved motifs, including a pseudokinase domain and a C-terminal E3 ligase-binding domain. In this review, we focus on the role of Trib (mammalian Tribbles homologues) proteins in mammalian haematopoiesis and leukaemia. The Trib proteins show divergent expression in haematopoietic cells, probably indicating cell-specific functions. The roles of the Trib proteins in oncogenesis are also varied and appear to be tissue-specific. Finally, we discuss the potential mechanisms by which the Trib proteins preferentially regulate these processes in multiple cell types. © 2015 Authors; published by Portland Press Limited.

Semi-quantitative estimation of cellular SiO2 nanoparticles using flow cytometry combined with X-ray fluorescence measurements.

PubMed

Choi, Seo Yeon; Yang, Nuri; Jeon, Soo Kyung; Yoon, Tae Hyun

2014-09-01

In this study, we have demonstrated feasibility of a semi-quantitative approach for the estimation of cellular SiO2 nanoparticles (NPs), which is based on the flow cytometry measurements of their normalized side scattering intensity. In order to improve our understanding on the quantitative aspects of cell-nanoparticle interactions, flow cytometry, transmission electron microscopy, and X-ray fluorescence experiments were carefully performed for the HeLa cells exposed to SiO2 NPs with different core diameters, hydrodynamic sizes, and surface charges. Based on the observed relationships among the experimental data, a semi-quantitative cellular SiO2 NPs estimation method from their normalized side scattering and core diameters was proposed, which can be applied for the determination of cellular SiO2 NPs within their size-dependent linear ranges. © 2014 International Society for Advancement of Cytometry.

Simulated hypogravity and synaptogenesis in culture

NASA Technical Reports Server (NTRS)

Gruener, R.

1985-01-01

A study on the effects of simulated microgravity on spinal neurons and myocytes cultured from X. laevis, is performed. Horizontal clinorotation at 1-10 rpm lasted from 16-36 hours, a sufficient time for cells to proceed through ontogenetic maturation. Late appearance of striations, retarded consumpton of yolk platelets and fewer and thinner neurites indicate subnormal expression of cell functions. Furthermore, these cells do not respond normally to environmental cues like trophic substances or surface contact. The observed delay in cell maturation is consistent with a hypothesis that cellular graviperception may effect the centriole and cytoskeleton.

Selenium in bone health: roles in antioxidant protection and cell proliferation.

PubMed

Zeng, Huawei; Cao, Jay J; Combs, Gerald F

2013-01-10

Selenium (Se) is an essential trace element for humans and animals, and several findings suggest that dietary Se intake may be necessary for bone health. Such findings may relate to roles of Se in antioxidant protection, enhanced immune surveillance and modulation of cell proliferation. Elucidation of the mechanisms by which Se supports these cellular processes can lead to a better understanding of the role of this nutrient in normal bone metabolism. This article reviews the current knowledge concerning the molecular functions of Se relevant to bone health.

Selenium in Bone Health: Roles in Antioxidant Protection and Cell Proliferation

PubMed Central

Zeng, Huawei; Cao, Jay J.; Combs, Gerald F.

2013-01-01

Selenium (Se) is an essential trace element for humans and animals, and several findings suggest that dietary Se intake may be necessary for bone health. Such findings may relate to roles of Se in antioxidant protection, enhanced immune surveillance and modulation of cell proliferation. Elucidation of the mechanisms by which Se supports these cellular processes can lead to a better understanding of the role of this nutrient in normal bone metabolism. This article reviews the current knowledge concerning the molecular functions of Se relevant to bone health. PMID:23306191

High vascular delivery of EGF, but low receptor binding rate is observed in AsPC-1 tumors as compared to normal pancreas.

PubMed

Samkoe, Kimberley S; Sexton, Kristian; Tichauer, Kenneth M; Hextrum, Shannon K; Pardesi, Omar; Davis, Scott C; O'Hara, Julia A; Hoopes, P Jack; Hasan, Tayyaba; Pogue, Brian W

2012-08-01

Cellular receptor targeted imaging agents present the potential to target extracellular molecular expression in cancerous lesions; however, the image contrast in vivo does not reflect the magnitude of overexpression expected from in vitro data. Here, the in vivo delivery and binding kinetics of epidermal growth factor receptor (EGFR) was determined for normal pancreas and AsPC-1 orthotopic pancreatic tumors known to overexpress EGFR. EGFR in orthotopic xenograft AsPC-1 tumors was targeted with epidermal growth factor (EGF) conjugated with IRDye800CW. The transfer rate constants (k(e), K₁₂, k₂₁, k₂₃, and k₃₂) associated with a three-compartment model describing the vascular delivery, leakage rate and binding of targeted agents were determined experimentally. The plasma excretion rate, k (e), was determined from extracted blood plasma samples. K₁₂, k₂₁, and k₃₂ were determined from ex vivo tissue washing studies at time points ≥ 24 h. The measured in vivo uptake of IRDye800CW-EGF and a non-targeted tracer dye, IRDye700DX-carboxylate, injected simultaneously was used to determined k₂₃. The vascular exchange of IRDye800CW-EGF in the orthotopic tumor (K₁₂ and k₂₁) was higher than in the AsPC-1 tumor as compared to normal pancreas, suggesting that more targeted agent can be taken up in tumor tissue. However, the cellular associated (binding) rate constant (k₂₃) was slightly lower for AsPC-1 pancreatic tumor (4.1 × 10(-4) s(-1)) than the normal pancreas (5.5 × 10(-4) s(-1)), implying that less binding is occurring. Higher vascular delivery but low cellular association in the AsPC-1 tumor compared to the normal pancreas may be indicative of low receptor density due to low cellular content. This attribute of the AsPC-1 tumor may indicate one contributing cause of the difficulty in treating pancreatic tumors with cellular targeted agents.

Physiological ranges of matrix rigidity modulate primary mouse hepatocyte function in part through hepatocyte nuclear factor 4 alpha.

PubMed

Desai, Seema S; Tung, Jason C; Zhou, Vivian X; Grenert, James P; Malato, Yann; Rezvani, Milad; Español-Suñer, Regina; Willenbring, Holger; Weaver, Valerie M; Chang, Tammy T

2016-07-01

Matrix rigidity has important effects on cell behavior and is increased during liver fibrosis; however, its effect on primary hepatocyte function is unknown. We hypothesized that increased matrix rigidity in fibrotic livers would activate mechanotransduction in hepatocytes and lead to inhibition of liver-specific functions. To determine the physiologically relevant ranges of matrix stiffness at the cellular level, we performed detailed atomic force microscopy analysis across liver lobules from normal and fibrotic livers. We determined that normal liver matrix stiffness was around 150 Pa and increased to 1-6 kPa in areas near fibrillar collagen deposition in fibrotic livers. In vitro culture of primary hepatocytes on collagen matrix of tunable rigidity demonstrated that fibrotic levels of matrix stiffness had profound effects on cytoskeletal tension and significantly inhibited hepatocyte-specific functions. Normal liver stiffness maintained functional gene regulation by hepatocyte nuclear factor 4 alpha (HNF4α), whereas fibrotic matrix stiffness inhibited the HNF4α transcriptional network. Fibrotic levels of matrix stiffness activated mechanotransduction in primary hepatocytes through focal adhesion kinase. In addition, blockade of the Rho/Rho-associated protein kinase pathway rescued HNF4α expression from hepatocytes cultured on stiff matrix. Fibrotic levels of matrix stiffness significantly inhibit hepatocyte-specific functions in part by inhibiting the HNF4α transcriptional network mediated through the Rho/Rho-associated protein kinase pathway. Increased appreciation of the role of matrix rigidity in modulating hepatocyte function will advance our understanding of the mechanisms of hepatocyte dysfunction in liver cirrhosis and spur development of novel treatments for chronic liver disease. (Hepatology 2016;64:261-275). © 2016 by the American Association for the Study of Liver Diseases.

Signaling pathway activation drift during aging: Hutchinson-Gilford Progeria Syndrome fibroblasts are comparable to normal middle-age and old-age cells.

PubMed

Aliper, Alexander M; Csoka, Antonei Benjamin; Buzdin, Anton; Jetka, Tomasz; Roumiantsev, Sergey; Moskalev, Alexy; Zhavoronkov, Alex

2015-01-01

For the past several decades, research in understanding the molecular basis of human aging has progressed significantly with the analysis of premature aging syndromes. Progerin, an altered form of lamin A, has been identified as the cause of premature aging in Hutchinson-Gilford Progeria Syndrome (HGPS), and may be a contributing causative factor in normal aging. However, the question of whether HGPS actually recapitulates the normal aging process at the cellular and organismal level, or simply mimics the aging phenotype is widely debated. In the present study we analyzed publicly available microarray datasets for fibroblasts undergoing cellular aging in culture, as well as fibroblasts derived from young, middle-age, and old-age individuals, and patients with HGPS. Using GeroScope pathway analysis and drug discovery platform we analyzed the activation states of 65 major cellular signaling pathways. Our analysis reveals that signaling pathway activation states in cells derived from chronologically young patients with HGPS strongly resemble cells taken from normal middle-aged and old individuals. This clearly indicates that HGPS may truly represent accelerated aging, rather than being just a simulacrum. Our data also points to potential pathways that could be targeted to develop drugs and drug combinations for both HGPS and normal aging.

Improved intra-array and interarray normalization of peptide microarray phosphorylation for phosphorylome and kinome profiling by rational selection of relevant spots

PubMed Central

Scholma, Jetse; Fuhler, Gwenny M.; Joore, Jos; Hulsman, Marc; Schivo, Stefano; List, Alan F.; Reinders, Marcel J. T.; Peppelenbosch, Maikel P.; Post, Janine N.

2016-01-01

Massive parallel analysis using array technology has become the mainstay for analysis of genomes and transcriptomes. Analogously, the predominance of phosphorylation as a regulator of cellular metabolism has fostered the development of peptide arrays of kinase consensus substrates that allow the charting of cellular phosphorylation events (often called kinome profiling). However, whereas the bioinformatical framework for expression array analysis is well-developed, no advanced analysis tools are yet available for kinome profiling. Especially intra-array and interarray normalization of peptide array phosphorylation remain problematic, due to the absence of “housekeeping” kinases and the obvious fallacy of the assumption that different experimental conditions should exhibit equal amounts of kinase activity. Here we describe the development of analysis tools that reliably quantify phosphorylation of peptide arrays and that allow normalization of the signals obtained. We provide a method for intraslide gradient correction and spot quality control. We describe a novel interarray normalization procedure, named repetitive signal enhancement, RSE, which provides a mathematical approach to limit the false negative results occuring with the use of other normalization procedures. Using in silico and biological experiments we show that employing such protocols yields superior insight into cellular physiology as compared to classical analysis tools for kinome profiling. PMID:27225531

Murine colon proteome and characterization of the protein pathways

PubMed Central

2012-01-01

Background Most of the current proteomic researches focus on proteome alteration due to pathological disorders (i.e.: colorectal cancer) rather than normal healthy state when mentioning colon. As a result, there are lacks of information regarding normal whole tissue- colon proteome. Results We report here a detailed murine (mouse) whole tissue- colon protein reference dataset composed of 1237 confident protein (FDR < 2) with comprehensive insight on its peptide properties, cellular and subcellular localization, functional network GO annotation analysis, and its relative abundances. The presented dataset includes wide spectra of pI and Mw ranged from 3–12 and 4–600 KDa, respectively. Gravy index scoring predicted 19.5% membranous and 80.5% globularly located proteins. GO hierarchies and functional network analysis illustrated proteins function together with their relevance and implication of several candidates in malignancy such as Mitogen- activated protein kinase (Mapk8, 9) in colorectal cancer, Fibroblast growth factor receptor (Fgfr 2), Glutathione S-transferase (Gstp1) in prostate cancer, and Cell division control protein (Cdc42), Ras-related protein (Rac1,2) in pancreatic cancer. Protein abundances calculated with 3 different algorithms (NSAF, PAF and emPAI) provide a relative quantification under normal condition as guidance. Conclusions This highly confidence colon proteome catalogue will not only serve as a useful reference for further experiments characterizing differentially expressed proteins induced from diseased conditions, but also will aid in better understanding the ontology and functional absorptive mechanism of the colon as well. PMID:22929016

Cellular immunity and lymphokine production during spaceflights

NASA Technical Reports Server (NTRS)

Konstantinova, I. V.; Sonnenfeld, G.; Lesniak, A. T.; Shaffar, L.; Mandel, A.; Rykova, M. P.; Antropova, E. N.; Ferrua, B.

1991-01-01

Results are presented on changes in cellular immunity and in the production of lymphokine in spacecrews during spaceflights. Measurements were carried out on blood samples collected from 50 cosmonauts before and after spaceflights of different duration, on board Salyut-6, Salyut-7, or Mir. Additional data were obtained from rats flown on board the Cosmos-1667 and Cosmos-1887 biosatellites. The parameters measured included the PHA responsiveness of T lymphocytes, the activity of T-helper cells and of nonspecific T suppressors, the activity of the so-called natural killer lymphocytes, the production of gamma-interferon, and the cell-surface markers. Results showed that the frequency and the extent of changes in the immunologic resistance of subjects depended on the duration of the flight. However, even after the most prolonged (365 days) spaceflight, the changes observed were mostly of a functional character with subsequent rapid return to normal.

Roles of the Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR pathways in controlling growth and sensitivity to therapy-implications for cancer and aging

PubMed Central

Steelman, Linda S.; Chappell, William H.; Abrams, Stephen L.; Kempf, C. Ruth; Long, Jacquelyn; Laidler, Piotr; Mijatovic, Sanja; Maksimovic-Ivanic, Danijela; Stivala, Franca; Mazzarino, Maria C.; Donia, Marco; Fagone, Paolo; Malaponte, Graziella; Nicoletti, Ferdinando; Libra, Massimo; Milella, Michele; Tafuri, Agostino; Bonati, Antonio; Bäsecke, Jörg; Cocco, Lucio; Evangelisti, Camilla; Martelli, Alberto M.; Montalto, Giuseppe; Cervello, Melchiorre; McCubrey, James A.

2011-01-01

Dysregulated signaling through the Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR pathways is often the result of genetic alterations in critical components in these pathways or upstream activators. Unrestricted cellular proliferation and decreased sensitivity to apoptotic-inducing agents are typically associated with activation of these pro-survival pathways. This review discusses the functions these pathways have in normal and neoplastic tissue growth and how they contribute to resistance to apoptotic stimuli. Crosstalk and commonly identified mutations that occur within these pathways that contribute to abnormal activation and cancer growth will also be addressed. Finally the recently described roles of these pathways in cancer stem cells, cellular senescence and aging will be evaluated. Controlling the expression of these pathways could ameliorate human health. PMID:21422497

New Insights into Thyroid Hormone Action

PubMed Central

Mendoza, Arturo; Hollenberg, Anthony N.

2017-01-01

Thyroid hormones (TH) are endocrine messengers essential for normal development and function of virtually every vertebrate. The hypothalamic-pituitary-thyroid axis is exquisitely modulated to maintain nearly constant TH (T4 and T3) concentrations in circulation. However peripheral tissues and the CNS control the intracellular availability of TH, suggesting that circulating concentrations of TH are not fully representative of what each cell type sees. Indeed, recent work in the field has identified that TH transporters, deiodinases and thyroid hormone receptor coregulators can strongly control tissue-specific sensitivity to a set amount of TH. Furthermore, the mechanism by which the thyroid hormone receptors regulate target gene expression can vary by gene, tissue and cellular context. This review will highlight novel insights into the machinery that controls the cellular response to TH, which include unique signaling cascades. These findings shed new light into the pathophysiology of human diseases caused by abnormal TH signaling. PMID:28174093

Control of proliferation and cancer growth by the Hippo signaling pathway

PubMed Central

Ehmer, Ursula; Sage, Julien

2015-01-01

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

Growth compensatory role of sulindac sulfide-induced thrombospondin-1 linked with ERK1/2 and RhoA GTPase signaling pathways

PubMed Central

Moon, Yuseok; Kim, Jeung Il; Yang, Hyun; Eling, Thomas E.

2009-01-01

Previously, we reported that non-steroidal anti-inflammatory drugs (NSAIDs) suppress cellular invasion which was mediated by thrombospondin-1 (TSP-1). As the extending study of the previous observation, we investigated the effect of NSAID-induced TSP-1 on the cellular growth and its related signaling transduction of the TSP-1 production. Among diverse NSAIDs, sulindac sulfide was most potent of inducing the human TSP-1 protein expression. Functionally, induced TSP-1 expression was associated with the growth-compensatory action of NSAID. TSP-1 expression was also elevated by mitogenic signals of ERK1/2 and RhoA GTPase pathway which had also growth-promotive capability after sulindac sulfide treatment. These findings suggest the possible mechanism through which tumor cells can survive the chemopreventive action of NSAIDs or the normal epithelium can reconstitute after NSAID-mediated ulceration in a compensatory way. PMID:18261746

Rational Design of Cancer-Targeted Benzoselenadiazole by RGD Peptide Functionalization for Cancer Theranostics.

PubMed

Yang, Liye; Li, Wenying; Huang, Yanyu; Zhou, Yangliang; Chen, Tianfeng

2015-09-01

A cancer-targeted conjugate of the selenadiazole derivative BSeC (benzo[1,2,5] selenadiazole-5-carboxylic acid) with RGD peptide as targeting molecule and PEI (polyethylenimine) as a linker is rationally designed and synthesized in the present study. The results show that RGD-PEI-BSeC forms nanoparticles in aqueous solution with a core-shell nanostructure and high stability under physiological conditions. This rational design effectively enhances the selective cellular uptake and cellular retention of BSeC in human glioma cells, and increases its selectivity between cancer and normal cells. The nanoparticles enter the cells through receptor-mediated endocytosis via clathrin-mediated and nystatin-dependent lipid raft-mediated pathways. Internalized nanoparticles trigger glioma cell apoptosis by activation of ROS-mediated p53 phosphorylation. Therefore, this study provides a strategy for the rational design of selenium-containing cancer-targeted theranostics. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A cellular, molecular, and pharmacological basis for appendage regeneration in mice

PubMed Central

Leung, Thomas H.; Snyder, Emily R.; Liu, Yinghua; Wang, Jing; Kim, Seung K.

2015-01-01

Regenerative medicine aims to restore normal tissue architecture and function. However, the basis of tissue regeneration in mammalian solid organs remains undefined. Remarkably, mice lacking p21 fully regenerate injured ears without discernable scarring. Here we show that, in wild-type mice following tissue injury, stromal-derived factor-1 (Sdf1) is up-regulated in the wound epidermis and recruits Cxcr4-expressing leukocytes to the injury site. In p21-deficient mice, Sdf1 up-regulation and the subsequent recruitment of Cxcr4-expressing leukocytes are significantly diminished, thereby permitting scarless appendage regeneration. Lineage tracing demonstrates that this regeneration derives from fate-restricted progenitor cells. Pharmacological or genetic disruption of Sdf1–Cxcr4 signaling enhances tissue repair, including full reconstitution of tissue architecture and all cell types. Our findings identify signaling and cellular mechanisms underlying appendage regeneration in mice and suggest new therapeutic approaches for regenerative medicine. PMID:26494786

The polarity protein partitioning-defective 1 (PAR-1) regulates dendritic spine morphogenesis through phosphorylating postsynaptic density protein 95 (PSD-95).

PubMed

Wu, Qian; DiBona, Victoria L; Bernard, Laura P; Zhang, Huaye

2012-08-31

The polarity protein PAR-1 plays an essential role in many cellular contexts, including embryogenesis, asymmetric cell division, directional migration, and epithelial morphogenesis. Despite its known importance in different cellular processes, the role of PAR-1 in neuronal morphogenesis is less well understood. In particular, its role in the morphogenesis of dendritic spines, which are sites of excitatory synaptic inputs, has been unclear. Here, we show that PAR-1 is required for normal spine morphogenesis in hippocampal neurons. We further show that PAR-1 functions through phosphorylating the synaptic scaffolding protein PSD-95 in this process. Phosphorylation at a conserved serine residue in the KXGS motif in PSD-95 regulates spine morphogenesis, and a phosphomimetic mutant of this site can rescue the defects of kinase-dead PAR-1. Together, our findings uncover a role of PAR-1 in spine morphogenesis in hippocampal neurons through phosphorylating PSD-95.

Fracture healing: mechanisms and interventions

PubMed Central

Einhorn, Thomas A.; Gerstenfeld, Louis C.

2015-01-01

Fractures are the most common large-organ, traumatic injuries to humans. The repair of bone fractures is a postnatal regenerative process that recapitulates many of the ontological events of embryonic skeletal development. Although fracture repair usually restores the damaged skeletal organ to its pre-injury cellular composition, structure and biomechanical function, about 10% of fractures will not heal normally. This article reviews the developmental progression of fracture healing at the tissue, cellular and molecular levels. Innate and adaptive immune processes are discussed as a component of the injury response, as are environmental factors, such as the extent of injury to the bone and surrounding tissue, fixation and the contribution of vascular tissues. We also present strategies for fracture treatment that have been tested in animal models and in clinical trials or case series. The biophysical and biological basis of the molecular actions of various therapeutic approaches, including recombinant human bone morphogenetic proteins and parathyroid hormone therapy, are also discussed. PMID:25266456

Alterations in plasma L-arginine and methylarginines in heart failure and after heart transplantation.

PubMed

Lundgren, Jakob; Sandqvist, Anna; Hedeland, Mikael; Bondesson, Ulf; Wikström, Gerhard; Rådegran, Göran

2018-04-12

Endothelial function, including the nitric oxide (NO)-pathway, has previously been extensively investigated in heart failure (HF). In contrast, studies are lacking on the NO pathway after heart transplantation (HT). We therefore investigated substances in the NO pathway prior to and after HT in relation to hemodynamic parameters. 12 patients (median age 50.0 yrs, 2 females), heart transplanted between June 2012 and February 2014, evaluated at our hemodynamic lab, at rest, prior to HT, as well as four weeks and six months after HT were included. All patients had normal left ventricular function post-operatively and none had post-operative pulmonary hypertension or acute cellular rejection requiring therapy at the evaluations. Plasma concentrations of ADMA, SDMA, L-Arginine, L-Ornithine and L-Citrulline were analyzed at each evaluation. In comparison to controls, the plasma L-Arginine concentration was low and ADMA high in HF patients, resulting in low L-Arginine/ADMA-ratio pre-HT. Already four weeks after HT L-Arginine was normalized whereas ADMA remained high. Consequently the L-Arginine/ADMA-ratio improved, but did not normalize. The biomarkers remained unchanged at the six-month evaluation and the L-Arginine/ADMA-ratio correlated inversely to pulmonary vascular resistance (PVR) six months post-HT. Plasma L-Arginine concentrations normalize after HT. However, as ADMA is unchanged, the L-Arginine/ADMA-ratio remained low and correlated inversely to PVR. Together these findings suggest that (i) the L-Arginine/ADMA-ratio may be an indicator of pulmonary vascular tone after HT, and that (ii) NO-dependent endothelial function is partly restored after HT. Considering the good postoperative outcome, the biomarker levels may be considered "normal" after HT.

Ground Testing of the EMCS Seed Cassette for Biocompatibility with the Cellular Slime Mold, Dictyostelium Discoideum

NASA Technical Reports Server (NTRS)

Hanely, Julia C.; Reinsch, Sigrid; Myers, Zachary A.; Freeman, John; Steele, Marianne K.; Sun, Gwo-Shing; Heathcote, David G.

2014-01-01

The European Modular Cultivation System, EMCS, was developed by ESA for plant experiments. To expand the use of flight verified hardware for various model organisms, we performed ground experiments to determine whether ARC EMCS Seed Cassettes could be adapted for use with cellular slime mold for future space flight experiments. Dictyostelium is a cellular slime mold that can exist both as a single-celled independent organism and as a part of a multicellular colony which functions as a unit (pseudoplasmodium). Under certain stress conditions, individual amoebae will aggregate to form multicellular structures. Developmental pathways are very similar to those found in Eukaryotic organisms, making this a uniquely interesting organism for use in genetic studies. Dictyostelium has been used as a genetic model organism for prior space flight experiments. Due to the formation of spores that are resistant to unfavorable conditions such as desiccation, Dictyostelium is also a good candidate for use in the EMCS Seed Cassettes. The growth substratum in the cassettes is a gridded polyether sulfone (PES) membrane. A blotter beneath the PES membranes contains dried growth medium. The goals of this study were to (1) verify that Dictyostelium are capable of normal growth and development on PES membranes, (2) develop a method for dehydration of Dictyostelium spores with successful recovery and development after rehydration, and (3) successful mock rehydration experiments in cassettes. Our results show normal developmental progression in two strains of Dictyostelium discoideum on PES membranes with a bacterial food source. We have successfully performed a mock rehydration of spores with developmental progression from aggregation to slug formation, and production of morphologically normal spores within 9 days of rehydration. Our results indicate that experiments on the ISS using the slime mold, Dictyostelium discoideum could potentially be performed in the flight verified hardware of the EMCS ARC Seed Cassettes.

Functional tissue engineering of tendon: Establishing biological success criteria for improving tendon repair.

PubMed

Breidenbach, Andrew P; Gilday, Steven D; Lalley, Andrea L; Dyment, Nathaniel A; Gooch, Cynthia; Shearn, Jason T; Butler, David L

2014-06-27

Improving tendon repair using Functional Tissue Engineering (FTE) principles has been the focus of our laboratory over the last decade. Although our primary goals were initially focused only on mechanical outcomes, we are now carefully assessing the biological properties of our tissue-engineered tendon repairs so as to link biological influences with mechanics. However, given the complexities of tendon development and healing, it remains challenging to determine which aspects of tendon biology are the most important to focus on in the context of tissue engineering. To address this problem, we have formalized a strategy to identify, prioritize, and evaluate potential biological success criteria for tendon repair. We have defined numerous biological properties of normal tendon relative to cellular phenotype, extracellular matrix and tissue ultra-structure that we would like to reproduce in our tissue-engineered repairs and prioritized these biological criteria by examining their relative importance during both normal development and natural tendon healing. Here, we propose three specific biological criteria which we believe are essential for normal tendon function: (1) scleraxis-expressing cells; (2) well-organized and axially-aligned collagen fibrils having bimodal diameter distribution; and (3) a specialized tendon-to-bone insertion site. Moving forward, these biological success criteria will be used in conjunction with our already established mechanical success criteria to evaluate the effectiveness of our tissue-engineered tendon repairs. © 2013 Published by Elsevier Ltd.

Toward an understanding of mechanism of aging-induced oxidative stress in human mesenchymal stem cells.

PubMed

Benameur, Laila; Charif, Naceur; Li, Yueying; Stoltz, Jean-François; de Isla, Natalia

2015-01-01

Under physiological conditions, there is a production of limited range of free radicals. However, when the cellular antioxidant defence systems, overwhelm and fail to reverse back the free radicals to their normal basal levels, there is a creation of a condition of redox disequilibrium termed "oxidative stress", which is implicated in a very wide spectrum of genetic, metabolic, and cellular responses. The excess of free radicals can, cause unfavourable molecular alterations to biomolecules through oxidation of lipids, proteins, RNA and DNA, that can in turn lead to mutagenesis, carcinogenesis, and aging. Mesenchymal stem cells (MSCs) have been proven to be a promising source of cells for regenerative medicine, and to be useful in the treatment of pathologies in which tissue damage is linked to oxidative stress. Moreover, MSCs appeared to efficiently manage oxidative stress and to be more resistant to oxidative insult than normal somatic cells, making them an interesting and testable model for the role of oxidative stress in the aging process. In addition, aging is accompanied by a progressive decline in stem cell function, resulting in less effective tissue homeostasis and repair. Also, there is an obvious link between intracellular reactive oxygen species levels and cellular senescence. To date, few studies have investigated the promotion of aging by oxidative stress on human MSCs, and the mechanism by which oxidative stress induce stem cell aging is poorly understood. In this context, the aim of this review is to gain insight the current knowledge about the molecular mechanisms of aging-induced oxidative stress in human MSCs.

Physiology and pathophysiology of potassium homeostasis.

PubMed

Palmer, Biff F; Clegg, Deborah J

2016-12-01

Total body potassium content and proper distribution of potassium across the cell membrane is of critical importance for normal cellular function. Potassium homeostasis is maintained by several different methods. In the kidney, total body potassium content is achieved by alterations in renal excretion of potassium in response to variations in intake. Insulin and beta-adrenergic tone play critical roles in maintaining the internal distribution of potassium under normal conditions. Despite homeostatic pathways designed to maintain potassium levels within the normal range, disorders of altered potassium homeostasis are common. The clinical approach to designing effective treatments relies on understanding the pathophysiology and regulatory influences which govern the internal distribution and external balance of potassium. Here we provide an overview of the key regulatory aspects of normal potassium physiology. This review is designed to provide an overview of potassium homeostasis as well as provide references of seminal papers to guide the reader into a more in depth discussion of the importance of potassium balance. This review is designed to be a resource for educators and well-informed clinicians who are teaching trainees about the importance of potassium balance. Copyright © 2016 the American Physiological Society.

Oncogenes in retroviruses and cells

NASA Astrophysics Data System (ADS)

Kurth, Reinhard

1983-09-01

Oncogenes are genes that cause cancer. Retroviruses contain oncogenes and cause cancer in animals and, perhaps, in man. The viruses have appropriated their oncogenes from normal cellular DNA by genetic recombination. Correspondingly, uninfected vertebrate cells contain a family of evolutionary conserved cellular oncogenes. Retrovirus infection, introducing additional viral oncogenes into the cells, as well as carcinogen-mediated activation of cellular oncogenes may both lead to increased synthesis of oncogene encoded transforming proteins which convert normal cells to tumor cells. Unique retroviruses of human origin have recently been identified. They may, on occasion, directly cause tumors in man. However, the general significance of retroviruses may better be illustrated by their remarkable genetic composition which allows them to promote tumor growth by a variety of genetic mechanisms.

A modular framework for biomedical concept recognition

PubMed Central

2013-01-01

Background Concept recognition is an essential task in biomedical information extraction, presenting several complex and unsolved challenges. The development of such solutions is typically performed in an ad-hoc manner or using general information extraction frameworks, which are not optimized for the biomedical domain and normally require the integration of complex external libraries and/or the development of custom tools. Results This article presents Neji, an open source framework optimized for biomedical concept recognition built around four key characteristics: modularity, scalability, speed, and usability. It integrates modules for biomedical natural language processing, such as sentence splitting, tokenization, lemmatization, part-of-speech tagging, chunking and dependency parsing. Concept recognition is provided through dictionary matching and machine learning with normalization methods. Neji also integrates an innovative concept tree implementation, supporting overlapped concept names and respective disambiguation techniques. The most popular input and output formats, namely Pubmed XML, IeXML, CoNLL and A1, are also supported. On top of the built-in functionalities, developers and researchers can implement new processing modules or pipelines, or use the provided command-line interface tool to build their own solutions, applying the most appropriate techniques to identify heterogeneous biomedical concepts. Neji was evaluated against three gold standard corpora with heterogeneous biomedical concepts (CRAFT, AnEM and NCBI disease corpus), achieving high performance results on named entity recognition (F1-measure for overlap matching: species 95%, cell 92%, cellular components 83%, gene and proteins 76%, chemicals 65%, biological processes and molecular functions 63%, disorders 85%, and anatomical entities 82%) and on entity normalization (F1-measure for overlap name matching and correct identifier included in the returned list of identifiers: species 88%, cell 71%, cellular components 72%, gene and proteins 64%, chemicals 53%, and biological processes and molecular functions 40%). Neji provides fast and multi-threaded data processing, annotating up to 1200 sentences/second when using dictionary-based concept identification. Conclusions Considering the provided features and underlying characteristics, we believe that Neji is an important contribution to the biomedical community, streamlining the development of complex concept recognition solutions. Neji is freely available at http://bioinformatics.ua.pt/neji. PMID:24063607

Functional classification of skeletal muscle networks. I. Normal physiology

PubMed Central

Wang, Yu; Winters, Jack

2012-01-01

Extensive measurements of the parts list of human skeletal muscle through transcriptomics and other phenotypic assays offer the opportunity to reconstruct detailed functional models. Through integration of vast amounts of data present in databases and extant knowledge of muscle function combined with robust analyses that include a clustering approach, we present both a protein parts list and network models for skeletal muscle function. The model comprises the four key functional family networks that coexist within a functional space; namely, excitation-activation family (forward pathways that transmit a motoneuronal command signal into the spatial volume of the cell and then use Ca2+ fluxes to bind Ca2+ to troponin C sites on F-actin filaments, plus transmembrane pumps that maintain transmission capacity); mechanical transmission family (a sophisticated three-dimensional mechanical apparatus that bidirectionally couples the millions of actin-myosin nanomotors with external axial tensile forces at insertion sites); metabolic and bioenergetics family (pathways that supply energy for the skeletal muscle function under widely varying demands and provide for other cellular processes); and signaling-production family (which represents various sensing, signal transduction, and nuclear infrastructure that controls the turn over and structural integrity and regulates the maintenance, regeneration, and remodeling of the muscle). Within each family, we identify subfamilies that function as a unit through analysis of large-scale transcription profiles of muscle and other tissues. This comprehensive network model provides a framework for exploring functional mechanisms of the skeletal muscle in normal and pathophysiology, as well as for quantitative modeling. PMID:23085959

Developing and applying the adverse outcome pathway ...

EPA Pesticide Factsheets

To support a paradigm shift in regulatory toxicology testing and risk assessment, the Adverse Outcome Pathway (AOP) concept has recently been proposed. This concept is similar to that for Mode of Action (MOA), describing a sequence of measurable key events triggered by a molecular initiating event in which a stressor interacts with a biological target. The resulting cascade of key events includes molecular, cellular, structural and functional changes in biological systems, resulting in a measurable adverse outcome. Thereby, an AOP ideally provides information relevant to chemical structure-activity relationships as a basis to predict effects for structurally similar compounds. AOPs could potentially also form the basis for qualitative and quantitative predictive modeling of the human adverse outcome resulting from molecular initiating or other key events for which higher-throughput testing methods are available or can be developed.A variety of cellular and molecular processes are known to be critical to normal function of the central (CNS) and peripheral nervous systems (PNS). Because of the biological and functional complexity of the CNS and PNS, it has been challenging to establish causative links and quantitative relationships between key events that comprise the pathways leading from chemical exposure to an adverse outcome in the nervous system. Following introduction of principles of the description and assessment of MOA and AOPs, examples of adverse out

Estrogen Biology: New Insights into GPER Function and Clinical Opportunities

PubMed Central

Prossnitz, Eric R.; Barton, Matthias

2014-01-01

Estrogens play an important role in the regulation of normal physiology, aging and many disease states. Although the nuclear estrogen receptors have classically been described to function as ligand-activated transcription factors mediating genomic effects in hormonally regulated tissues, more recent studies reveal that estrogens also mediate rapid signaling events traditionally associated with G protein-coupled receptors. The G protein-coupled estrogen receptor GPER (formerly GPR30) has now become recognized as a major mediator of estrogen’s rapid cellular effects throughout the body. With the discovery of selective synthetic ligands for GPER, both agonists and antagonists, as well as the use of GPER knockout mice, significant advances have been made in our understanding of GPER function at the cellular, tissue and organismal levels. In many instances, the protective/beneficial effects of estrogen are mimicked by selective GPER agonism and are absent or reduced in GPER knockout mice, suggesting an essential or at least parallel role for GPER in the actions of estrogen. In this review, we will discuss recent advances and our current understanding of the role of GPER and certain drugs such as SERMs and SERDs in physiology and disease. We will also highlight novel opportunities for clinical development towards GPER-targeted therapeutics, for molecular imaging, as well as for theranostic approaches and personalized medicine. PMID:24530924

Poliovirus replication requires the N-terminus but not the catalytic Sec7 domain of ArfGEF GBF1.

PubMed

Belov, George A; Kovtunovych, Gennadiy; Jackson, Catherine L; Ehrenfeld, Ellie

2010-10-01

Viruses are intracellular parasites whose reproduction relies on factors provided by the host. The cellular protein GBF1 is critical for poliovirus replication. Here we show that the contribution of GBF1 to virus replication is different from its known activities in uninfected cells. Normally GBF1 activates the ADP-ribosylation factor (Arf) GTPases necessary for formation of COPI transport vesicles. GBF1 function is modulated by p115 and Rab1b. However, in polio-infected cells, p115 is degraded and neither p115 nor Rab1b knock-down affects virus replication. Poliovirus infection is very sensitive to brefeldin A (BFA), an inhibitor of Arf activation by GBF1. BFA targets the catalytic Sec7 domain of GBF1. Nevertheless the BFA block of polio replication is rescued by expression of only the N-terminal region of GBF1 lacking the Sec7 domain. Replication of BFA-resistant poliovirus in the presence of BFA is uncoupled from Arf activation but is dependent on GBF1. Thus the function(s) of this protein essential for viral replication can be separated from those required for cellular metabolism. © Published 2010. This article is a US Government work and is in the public domain in the USA.

The cell envelope proteome of Aggregatibacter actinomycetemcomitans

PubMed Central

Smith, Kenneth P.; Fields, Julia G.; Voogt, Richard D.; Deng, Bin; Lam, Ying-Wai; Mintz, Keith P.

2014-01-01

Summary The cell envelope of Gram-negative bacteria serves a critical role in maintenance of cellular homeostasis, resistance to external stress, and host-pathogen interactions. Envelope protein composition is influenced by the physiological and environmental demands placed on the bacterium. In this study, we report a comprehensive compilation of cell envelope proteins from the periodontal and systemic pathogen Aggregatibacter actinomycetemcomitans VT1169, an afimbriated serotype b strain. The urea-extracted membrane proteins were identified by mass spectrometry-based shotgun proteomics. The membrane proteome, isolated from actively growing bacteria under normal laboratory conditions, included 648 proteins representing 28% of the predicted ORFs in the genome. Bioinformatic analyses were used to annotate and predict the cellular location and function of the proteins. Surface adhesins, porins, lipoproteins, numerous influx and efflux pumps, multiple sugar, amino acid and iron transporters, and components of the type I, II and V secretion systems were identified. Periplasmic space and cytoplasmic proteins with chaperone function were also identified. 107 proteins with unknown function were associated with the cell envelope. Orthologs of a subset of these uncharacterized proteins are present in other bacterial genomes, while others are found exclusively in A. actinomycetemcomitans. This knowledge will contribute to elucidating the role of cell envelope proteins in bacterial growth and survival in the oral cavity. PMID:25055881

Macrophage Polarization in Chronic Inflammatory Diseases: Killers or Builders?

PubMed Central

Baci, Denisa; Tremolati, Marco; Fanuli, Matteo; Farronato, Giampietro; Mortara, Lorenzo

2018-01-01

Macrophages are key cellular components of the innate immunity, acting as the main player in the first-line defence against the pathogens and modulating homeostatic and inflammatory responses. Plasticity is a major feature of macrophages resulting in extreme heterogeneity both in normal and in pathological conditions. Macrophages are not homogenous, and they are generally categorized into two broad but distinct subsets as either classically activated (M1) or alternatively activated (M2). However, macrophages represent a continuum of highly plastic effector cells, resembling a spectrum of diverse phenotype states. Induction of specific macrophage functions is closely related to the surrounding environment that acts as a relevant orchestrator of macrophage functions. This phenomenon, termed polarization, results from cell/cell, cell/molecule interaction, governing macrophage functionality within the hosting tissues. Here, we summarized relevant cellular and molecular mechanisms driving macrophage polarization in “distant” pathological conditions, such as cancer, type 2 diabetes, atherosclerosis, and periodontitis that share macrophage-driven inflammation as a key feature, playing their dual role as killers (M1-like) and/or builders (M2-like). We also dissect the physio/pathological consequences related to macrophage polarization within selected chronic inflammatory diseases, placing polarized macrophages as a relevant hallmark, putative biomarkers, and possible target for prevention/therapy. PMID:29507865

RHIM-based protein:protein interactions in anti-microbial defence against programmed cell death by necroptosis.

PubMed

Baker, Max O D G; Shanmugam, Nirukshan; Pham, Chi L L; Strange, Merryn; Steain, Megan; Sunde, Margaret

2018-05-05

The Receptor-interacting protein kinase Homotypic Interaction Motif (RHIM) is an amino acid sequence that mediates multiple protein:protein interactions in the mammalian programmed cell death pathway known as necroptosis. At least one key RHIM-based complex has been shown to have a functional amyloid fibril structure, which provides a stable hetero-oligomeric platform for downstream signaling. RHIMs and related motifs are present in immunity-related proteins across nature, from viruses to fungi to metazoans. Necroptosis is a hallmark feature of cellular clearance of infection. For this reason, numerous pathogens, including viruses and bacteria, have developed varied methods to modulate necroptosis, focusing on inhibiting RHIM:RHIM interactions, and thus their downstream cell death effects. This review will discuss current understanding of RHIM:RHIM interactions in normal cellular activation of necroptosis, from a structural and cell biology perspective. It will compare the mechanisms by which pathogens subvert these interactions in order to maintain their replicative and infective cycles and consider the similarities between RHIMs and other functional amyloid-forming proteins associated with cell death and innate immunity. It will discuss the implications of the heteromeric nature and structure of RHIM-based amyloid complexes in the context of other functional amyloids. Copyright © 2018. Published by Elsevier Ltd.

Spectroscopic analysis of normal and neoplastic (WI-FTC) thyroid tissue.

PubMed

Depciuch, Joanna; Stanek-Widera, Agata; Lange, Dariusz; Biskup-Frużyńska, Magdalena; Stanek-Tarkowska, Jadwiga; Czarny, Wojciech; Cebulski, Jozef

2018-06-07

Thyroid cancer holds the first place of the malignant tumors of the endocrine system. One of the less common thyroid cancers is follicular thyroid carcinoma (FTC), which is very difficult to diagnose because it gives the same image as adenoma, which is benign. Certainty of the diagnosis is gained only when FTC gives metastases. Therefore, it was decided to compare normal and neoplastic (FTC) thyroid tissues with Fourier Transform Infrared (FTIR) spectroscopy. The obtained FTIR spectra and Principal Component Analysis (PCA) allowed us to conclude that there are differences in the FTIR spectrum between normal tissues and those affected by cancer. In addition, the results indicate that there is a decrease in the number of functional groups that build cellular and tissue structures in tumoral tissues. The shifts of wave numbers corresponding to the protein and lipid function group vibrations, as well as the calculated second derivative of the FTIR spectra showed the structural changes in neoplastic tissues. Moreover, the deconvolution of the amide I massif indicates that in cancerous tissues the prevailing secondary structure is β-sheet structure, while in normal tissues it is α-helix. The obtained results allow us to conclude that infrared spectroscopy, in addition to providing information on the composition of tested samples, can be an excellent diagnostic tool contributing to understanding the FTC substrate. Copyright © 2018. Published by Elsevier B.V.

Gravitational Effects on Cellular Flame Structure

NASA Technical Reports Server (NTRS)

Dunsky, C. M.; Fernandez-Pello, A. C.

1991-01-01

An experimental investigation has been conducted of the effect of gravity on the structure of downwardly propagating, cellular premixed propane-oxygen-nitrogen flames anchored on a water-cooled porous-plug burner. The flame is subjected to microgravity conditions in the NASA Lewis 2.2-second drop tower, and flame characteristics are recorded on high-speed film. These are compared to flames at normal gravity conditions with the same equivalence ratio, dilution index, mixture flow rate, and ambient pressure. The results show that the cellular instability band, which is located in the rich mixture region, changes little under the absence of gravity. Lifted normal-gravity flames near the cellular/lifted limits, however, are observed to become cellular when gravity is reduced. Observations of a transient cell growth period following ignition point to heat loss as being an important mechanism in the overall flame stability, dominating the stabilizing effect of buoyancy for these downwardly-propagating burner-anchored flames. The pulsations that are observed in the plume and diffusion flame generated downstream of the premixed flame in the fuel rich cases disappear in microgravity, verifying that these fluctuations are gravity related.

Spatial differences of cellular origins and in vivo hypoxia modify contractile properties of pulmonary artery smooth muscle cells: lessons for arterial tissue engineering.

PubMed

Hall, S M; Soueid, A; Smith, T; Brown, R A; Haworth, S G; Mudera, V

2007-01-01

Tissue engineering of functional arteries is challenging. Within the pulmonary artery wall, smooth muscle cells (PASMCs) have site-specific developmental and functional phenotypes, reflecting differing contractile roles. The force generated by PASMCs isolated from the inner 25% and outer 50% of the media of intrapulmonary elastic arteries from five normal and eight chronically hypoxic (hypertensive) 14 day-old piglets was quantified in a three-dimensional (3D) collagen construct, using a culture force monitor. Outer medial PASMCs from normal piglets exerted more force (528 +/- 50 dynes) than those of hypoxic piglets (177 +/- 42 dynes; p < 0.01). Force generation by inner medial PASMCs from normal and hypoxic piglets was similar (349 +/- 35 and 239 +/- 60 dynes). In response to agonist (thromboxane) stimulation, all PASMCs from normal and hypoxic piglets contracted, but the increase in force generated by outer and inner hypoxic PASMCs (ranges 13-72 and 14-56 dynes) was less than by normal PASMCs (ranges 27-154 and 34-159 dynes, respectively; p < 0.05 for both). All hypoxic PASMCs were unresponsive to antagonist (sodium nitroprusside) stimulation, all normal PASMCs relaxed (range - 87 to - 494 dynes). Myosin heavy chain expression by both hypoxic PASMC phenotypes was less than normal (p < 0.05 for both), as was the activity of focal adhesion kinase, regulating contraction, in hypoxic inner PASMCs (p < 0.01). Chronic hypoxia resulted in the development of abnormal PASMC phenotypes, which in collagen constructs exhibited a reduction in contractile force and reactivity to agonists. Characterization of the mechanical response of spatially distinct cells and modification of their behaviour by hypoxia is critical for successful tissue engineering of major blood vessels.

Nonlinear and Stochastic Dynamics in the Heart

PubMed Central

Qu, Zhilin; Hu, Gang; Garfinkel, Alan; Weiss, James N.

2014-01-01

In a normal human life span, the heart beats about 2 to 3 billion times. Under diseased conditions, a heart may lose its normal rhythm and degenerate suddenly into much faster and irregular rhythms, called arrhythmias, which may lead to sudden death. The transition from a normal rhythm to an arrhythmia is a transition from regular electrical wave conduction to irregular or turbulent wave conduction in the heart, and thus this medical problem is also a problem of physics and mathematics. In the last century, clinical, experimental, and theoretical studies have shown that dynamical theories play fundamental roles in understanding the mechanisms of the genesis of the normal heart rhythm as well as lethal arrhythmias. In this article, we summarize in detail the nonlinear and stochastic dynamics occurring in the heart and their links to normal cardiac functions and arrhythmias, providing a holistic view through integrating dynamics from the molecular (microscopic) scale, to the organelle (mesoscopic) scale, to the cellular, tissue, and organ (macroscopic) scales. We discuss what existing problems and challenges are waiting to be solved and how multi-scale mathematical modeling and nonlinear dynamics may be helpful for solving these problems. PMID:25267872

Normalization of environmental metagenomic DNA enhances the discovery of under-represented microbial community members.

PubMed

Ramond, J-B; Makhalanyane, T P; Tuffin, M I; Cowan, D A

2015-04-01

Normalization is a procedure classically employed to detect rare sequences in cellular expression profiles (i.e. cDNA libraries). Here, we present a normalization protocol involving the direct treatment of extracted environmental metagenomic DNA with S1 nuclease, referred to as normalization of metagenomic DNA: NmDNA. We demonstrate that NmDNA, prior to post hoc PCR-based experiments (16S rRNA gene T-RFLP fingerprinting and clone library), increased the diversity of sequences retrieved from environmental microbial communities by detection of rarer sequences. This approach could be used to enhance the resolution of detection of ecologically relevant rare members in environmental microbial assemblages and therefore is promising in enabling a better understanding of ecosystem functioning. This study is the first testing 'normalization' on environmental metagenomic DNA (mDNA). The aim of this procedure was to improve the identification of rare phylotypes in environmental communities. Using hypoliths as model systems, we present evidence that this post-mDNA extraction molecular procedure substantially enhances the detection of less common phylotypes and could even lead to the discovery of novel microbial genotypes within a given environment. © 2014 The Society for Applied Microbiology.

Discovery of small molecules binding to the normal conformation of prion by combining virtual screening and multiple biological activity evaluation methods

NASA Astrophysics Data System (ADS)

Li, Lanlan; Wei, Wei; Jia, Wen-Juan; Zhu, Yongchang; Zhang, Yan; Chen, Jiang-Huai; Tian, Jiaqi; Liu, Huanxiang; He, Yong-Xing; Yao, Xiaojun

2017-12-01

Conformational conversion of the normal cellular prion protein, PrPC, into the misfolded isoform, PrPSc, is considered to be a central event in the development of fatal neurodegenerative diseases. Stabilization of prion protein at the normal cellular form (PrPC) with small molecules is a rational and efficient strategy for treatment of prion related diseases. However, few compounds have been identified as potent prion inhibitors by binding to the normal conformation of prion. In this work, to rational screening of inhibitors capable of stabilizing cellular form of prion protein, multiple approaches combining docking-based virtual screening, steady-state fluorescence quenching, surface plasmon resonance and thioflavin T fluorescence assay were used to discover new compounds interrupting PrPC to PrPSc conversion. Compound 3253-0207 that can bind to PrPC with micromolar affinity and inhibit prion fibrillation was identified from small molecule databases. Molecular dynamics simulation indicated that compound 3253-0207 can bind to the hotspot residues in the binding pocket composed by β1, β2 and α2, which are significant structure moieties in conversion from PrPC to PrPSc.

Potential usage of proteasome inhibitor bortezomib (Velcade, PS-341) in the treatment of metastatic melanoma: basic and clinical aspects

PubMed Central

Shahshahan, Mohammad A; Beckley, Maureen N; Jazirehi, Ali R

2011-01-01

Protein degradation by proteasome is essential to the regulation of important cellular functions including cell cycle progression, proliferation, differentiation and apoptosis. Abnormal proteasomal degradation of key regulatory proteins perturbs the normal dynamics of these cellular processes culminating in uncontrolled cell cycle progression and decreased apoptosis leading to the characteristic cancer cell phenotype. Proteasome inhibitors are a novel group of therapeutic agents designed to oppose the increased proteasomal degradation observed in various cancers while restoring key cellular functions such as apoptosis, cell cycle progression, and the inhibition of angiogenesis. Several proteasome inhibitors have been evaluated in pre- and clinical studies for their potential usage in clinical oncology. Bortezomib (Velcade, PS-341) is the first Food and Drug Administration-approved proteasome inhibitor for the treatment of multiple myeloma and mantle cell lymphoma. Bortezomib's ability to preferentially induce toxicity and cell death in tumor cells while rendering healthy cells unaffected makes it a powerful therapeutic agent and has extended its use in other types of malignancies. The ability of bortezomib and other proteasome inhibitors to synergize with conventional therapies in killing tumors in various in vitro and in vivo models makes this class of drugs a powerful tool in overcoming acquired and inherent resistance observed in many cancers. This is achieved through modulation of aberrant cellular survival signal transduction pathways and their downstream anti-apoptotic gene products. This review will discuss the anti-neoplastic effects of various proteasome inhibitors in a variety of cancers with a special emphasis on bortezomib, its mechanism of action and role in cancer therapy. We further discuss the potential use of bortezomib in the treatment of metastatic melanoma. PMID:22016836

From Embryonic Development to Human Diseases: The Functional Role of Caveolae/Caveolin

PubMed Central

Sohn, Jihee; Brick, Rachel M.; Tuan, Rocky S.

2017-01-01

Caveolae, an almost ubiquitous, structural component of the plasma membrane, play a critical role in many functions essential for proper cell function, including membrane trafficking, signal transduction, extracellular matrix remodeling, and tissue regeneration. Three main types of caveolin proteins have been identified from caveolae since the discovery of caveolin-1 in the early 1990s. All three (Cav-1, Cav-2, and Cav-3) play crucial roles in mammalian physiology, and can effect pathogenesis in a wide range of human diseases. While many biological activities of caveolins have been uncovered since its discovery, their role and regulation in embryonic develop remain largely poorly understood, although there is increasing evidence that caveolins may be linked to lung and brain birth defects. Further investigations are clearly needed to decipher how caveolae/caveolins mediate cellular functions and activities of normal embryogenesis and how their perturbations contribute to developmental disorders. PMID:26991990

Biologic effects of interferons.

PubMed

Gresser, I

1990-12-01

If one were to review articles on IFN published between 1957 and 1967 it would become apparent that virtually none of the tenets held then are still valid today. Whereas IFN was long considered to be a specific antiviral substance without any effect on normal cellular metabolism, we accept today that it affects normal cell division and many specialized cellular functions. In this respect it is not unique; IFN is a prototype of a family of similar substances now called cytokines that all appear to function as regulatory molecules. It was held that the production of IFN constituted a specific response to a viral infection. Today we believe that IFN is an integral part of a cytokine network and that they and other cytokines may be produced constitutively at low levels. These substances exert multiple effects on virtually all cells. They play an important role in host defense against viral and parasitic infections, and in the resistance to experimental tumors. IFN can be shown to exert effects on the immune system and on lymphocyte circulation. Lastly, because of the multiplicity of their biologic effects, they may even contribute to the pathogenesis of certain diseases. Thus, when large amounts of IFN are administered or induced in newborn mice they can cause liver, kidney, and pulmonary disease. The field of IFN and cytokine research continues to expand and there is an increasing number of therapeutic applications. Twenty years from now, scientists and clinicians may be surprised that we understood so little of how IFN act and how inadequately we used them to treat disease.

Dendritic Cytoskeletal Architecture Is Modulated by Combinatorial Transcriptional Regulation in Drosophila melanogaster.

PubMed

Das, Ravi; Bhattacharjee, Shatabdi; Patel, Atit A; Harris, Jenna M; Bhattacharya, Surajit; Letcher, Jamin M; Clark, Sarah G; Nanda, Sumit; Iyer, Eswar Prasad R; Ascoli, Giorgio A; Cox, Daniel N

2017-12-01

Transcription factors (TFs) have emerged as essential cell autonomous mediators of subtype specific dendritogenesis; however, the downstream effectors of these TFs remain largely unknown, as are the cellular events that TFs control to direct morphological change. As dendritic morphology is largely dictated by the organization of the actin and microtubule (MT) cytoskeletons, elucidating TF-mediated cytoskeletal regulatory programs is key to understanding molecular control of diverse dendritic morphologies. Previous studies in Drosophila melanogaster have demonstrated that the conserved TFs Cut and Knot exert combinatorial control over aspects of dendritic cytoskeleton development, promoting actin and MT-based arbor morphology, respectively. To investigate transcriptional targets of Cut and/or Knot regulation, we conducted systematic neurogenomic studies, coupled with in vivo genetic screens utilizing multi-fluor cytoskeletal and membrane marker reporters. These analyses identified a host of putative Cut and/or Knot effector molecules, and a subset of these putative TF targets converge on modulating dendritic cytoskeletal architecture, which are grouped into three major phenotypic categories, based upon neuromorphometric analyses: complexity enhancer, complexity shifter, and complexity suppressor. Complexity enhancer genes normally function to promote higher order dendritic growth and branching with variable effects on MT stabilization and F-actin organization, whereas complexity shifter and complexity suppressor genes normally function in regulating proximal-distal branching distribution or in restricting higher order branching complexity, respectively, with spatially restricted impacts on the dendritic cytoskeleton. Collectively, we implicate novel genes and cellular programs by which TFs distinctly and combinatorially govern dendritogenesis via cytoskeletal modulation. Copyright © 2017 by the Genetics Society of America.

[The blood glucose value not necessarily indicates correctly the cellular metabolic state].

PubMed

Simon, Kornél; Wittmann, István

2017-03-01

In clinical recommendations the normalized blood glucose level is declared as the main target in therapy of diabetes mellitus, i.e. the achievement of euglycemia is the main therapeutic goal. This approach suggests, that the normal blood glucose value is the marker of the normal carbohydrate metabolism (eumetabolism), and vice versa: hyperglycemia is associated with abnormal metabolism (dysmetabolism). However the question arises, whether identical blood glucose values do reflect the same intracellular biochemical mechanisms? On the basis of data published in the literature authors try to answer these questions by studying the relations between the short/longterm blood glucose level and the cellular metabolism in different clinical settings characterized by divergent pathophysiological parameters. The correlations between blood glucose level and cellular metabolism in development of micro-, and macroangiopathy, in the breakthrough phenomenon, as well as during administration of metabolic promoters, the discrepancies of relation between blood glucose values and cellular metabolism in type 1, and type 2 diabetes mellitus, furthermore association between blood glucose value and myocardial metabolism in acute and chronic stress were analyzed. Authors conclude, that the actual blood glucose values reveal the actual cellular metabolism in a very variable manner: neither euglycemia does mandatorily indicate eumetabolism (balance of cellular energy production), nor hyperglycemia is necessarily a marker of abnormal metabolic state (dept of cellular energy production). Moreover, at the same actual blood glucose level both the metabolic efficacy of the same organ may sharply vary, and the intracellular biochemical machinery could also be very different. In case of the very same longterm blood glucose level the metabolic state of the different organs could be very variable: some organs show an energetically balanced metabolism, while others produce a significant deficit. These inconsistencies between blood glucose level and cellular metabolism can be explained by the fact, that blood glucose value is a transport parameter, reflecting the actual steady state of glucose transport from the carbohydrate pools into the blood, and that from the blood into the tissues. Without knowing the speed of these transports of opposite direction, the blood glucose value per se can not reveal the quantitative and qualitative characteristics of cellular metabolism. Orv. Hetil., 2017, 158(11), 409-417.

Single Cell Force Spectroscopy for Quantification of Cellular Adhesion on Surfaces

NASA Astrophysics Data System (ADS)

Christenson, Wayne B.

Cell adhesion is an important aspect of many biological processes. The atomic force microscope (AFM) has made it possible to quantify the forces involved in cellular adhesion using a technique called single cell force spectroscopy (SCFS). AFM based SCFS offers versatile control over experimental conditions for probing directly the interaction between specific cell types and specific proteins, surfaces, or other cells. Transmembrane integrins are the primary proteins involved in cellular adhesion to the extra cellular matix (ECM). One of the chief integrins involved in the adhesion of leukocyte cells is alpha Mbeta2 (Mac-1). The experiments in this dissertation quantify the adhesion of Mac-1 expressing human embryonic kidney (HEK Mac-1), platelets, and neutrophils cells on substrates with different concentrations of fibrinogen and on fibrin gels and multi-layered fibrinogen coated fibrin gels. It was shown that multi-layered fibrinogen reduces the adhesion force of these cells considerably. A novel method was developed as part of this research combining total internal reflection microscopy (TIRFM) with SCFS allowing for optical microscopy of HEK Mac-1 cells interacting with bovine serum albumin (BSA) coated glass after interacting with multi-layered fibrinogen. HEK Mac-1 cells are able to remove fibrinogen molecules from the multi-layered fibrinogen matrix. An analysis methodology for quantifying the kinetic parameters of integrin-ligand interactions from SCFS experiments is proposed, and the kinetic parameters of the Mac-1 fibrinogen bond are quantified. Additional SCFS experiments quantify the adhesion of macrophages and HEK Mac-1 cells on functionalized glass surfaces and normal glass surfaces. Both cell types show highest adhesion on a novel functionalized glass surface that was prepared to induce macrophage fusion. These experiments demonstrate the versatility of AFM based SCFS, and how it can be applied to address many questions in cellular biology offering quantitative insights.

Harnessing Solute Carrier Transporters for Precision Oncology.

PubMed

Nyquist, Michael D; Prasad, Bhagwat; Mostaghel, Elahe A

2017-03-28

Solute Carrier (SLC) transporters are a large superfamily of transmembrane carriers involved in the regulated transport of metabolites, nutrients, ions and drugs across cellular membranes. A subset of these solute carriers play a significant role in the cellular uptake of many cancer therapeutics, ranging from chemotherapeutics such as antimetabolites, topoisomerase inhibitors, platinum-based drugs and taxanes to targeted therapies such as tyrosine kinase inhibitors. SLC transporters are co-expressed in groups and patterns across normal tissues, suggesting they may comprise a coordinated regulatory circuit serving to mediate normal tissue functions. In cancer however, there are dramatic changes in expression patterns of SLC transporters. This frequently serves to feed the increased metabolic demands of the tumor cell for amino acids, nucleotides and other metabolites, but also presents a therapeutic opportunity, as increased transporter expression may serve to increase intracellular concentrations of substrate drugs. In this review, we examine the regulation of drug transporters in cancer and how this impacts therapy response, and discuss novel approaches to targeting therapies to specific cancers via tumor-specific aberrations in transporter expression. We propose that among the oncogenic changes in SLC transporter expression there exist emergent vulnerabilities that can be exploited therapeutically, extending the application of precision medicine from tumor-specific drug targets to tumor-specific determinants of drug uptake.

Aromatic Side Chain Water-to-Lipid Transfer Free Energies Show a Depth Dependence across the Membrane Normal.

PubMed

McDonald, Sarah K; Fleming, Karen G

2016-06-29

Quantitating and understanding the physical forces responsible for the interactions of biomolecules are fundamental to the biological sciences. This is especially challenging for membrane proteins because they are embedded within cellular bilayers that provide a unique medium in which hydrophobic sequences must fold. Knowledge of the energetics of protein-lipid interactions is thus vital to understand cellular processes involving membrane proteins. Here we used a host-guest mutational strategy to calculate the Gibbs free energy changes of water-to-lipid transfer for the aromatic side chains Trp, Tyr, and Phe as a function of depth in the membrane. This work reveals an energetic gradient in the transfer free energies for Trp and Tyr, where transfer was most favorable to the membrane interfacial region and comparatively less favorable into the bilayer center. The transfer energetics follows the concentration gradient of polar atoms across the bilayer normal that naturally occurs in biological membranes. Additional measurements revealed nearest-neighbor coupling in the data set are influenced by a network of aromatic side chains in the host protein. Taken together, these results show that aromatic side chains contribute significantly to membrane protein stability through either aromatic-aromatic interactions or placement at the membrane interface.

Regulation and disregulation of mammalian nucleotide excision repair: A pathway to nongermline breast carcinogenesis

DOE PAGES

Latimer, Jean J.; Majekwana, Vongai J.; Pabon-Padin, Yashira R.; ...

2014-12-19

Nucleotide excision repair (NER) is important as a modulator of disease, especially in constitutive deficiencies, such as the cancer predisposition syndrome Xeroderma pigmentosum. We have found profound variation of NER capacity among normal individuals, between cell-types and during carcinogenesis. NER is a repair system for many types of DNA damage, and therefore many types of genotoxic carcinogenic exposures, including ultraviolet light, products of organic combustion, metals, oxidative stress, etc. Since NER is intimately related to cellular metabolism, requiring components of both the DNA replicative and transcription machinery, it has a narrow range of functional viability. Thus, genes in the NERmore » pathway are expressed at the low levels manifested by, for example, nuclear transcription factors. Since NER activity and gene expression vary by cell-type, it is inherently epigenetically regulated. Furthermore, this epigenetic regulation is disregulated during sporadic breast carcinogenesis. Loss of NER is one basis of genomic instability, a required element in cellular transformation, and one that potentially modulates response to therapy. In this article, we demonstrate differences in NER capacity in eight adult mouse tissues, and place this result into the context of our previous work on mouse extraembryonic tissues, normal human tissues and sporadic early stage human breast cancer.« less

Autophagy in tumorigenesis and energy metabolism: friend by day, foe by night.

PubMed

Mathew, Robin; White, Eileen

2011-02-01

Autophagy is the mechanism by which cells consume parts of themselves to survive starvation and stress. This self-cannibalization limits cell death and tissue inflammation, recycles energy and biosynthetic substrates and removes damaged proteins and organelles, accumulation of which is toxic. In normal tissues, autophagy-mediated damage mitigation may suppress tumorigenesis, while in advanced tumors macromolecular recycling may support survival by buffering metabolic demand under stress. As a result, autophagy-activation in normal cells may suppress tumorigenesis, while autophagy inhibition may be beneficial for the therapy of established tumors. The mechanisms by which autophagy supports cancer cell metabolism are slowly emerging. As cancer is being increasingly recognized as a metabolic disease, how autophagy-mediated catabolism impacts cellular and mammalian metabolism and tumor growth is of great interest. Most cancer therapeutics induce autophagy, either directly by modulating signaling pathways that control autophagy in the case of many targeted therapies, or indirectly in the case of cytotoxic therapy. However, the functional consequence of autophagy induction in the context of cancer therapy is not yet clear. A better understanding of how autophagy modulates cell metabolism under various cellular stresses and the consequences of this on tumorigenesis will help develop better therapeutic strategies against cancer prevention and treatment. Copyright © 2011 Elsevier Ltd. All rights reserved.

The zebrafish maternal-effect gene cellular atoll encodes the centriolar component sas-6 and defects in its paternal function promote whole genome duplication.

PubMed

Yabe, Taijiro; Ge, Xiaoyan; Pelegri, Francisco

2007-12-01

A female-sterile zebrafish maternal-effect mutation in cellular atoll (cea) results in defects in the initiation of cell division starting at the second cell division cycle. This phenomenon is caused by defects in centrosome duplication, which in turn affect the formation of a bipolar spindle. We show that cea encodes the centriolar coiled-coil protein Sas-6, and that zebrafish Cea/Sas-6 protein localizes to centrosomes. cea also has a genetic paternal contribution, which when mutated results in an arrested first cell division followed by normal cleavage. Our data supports the idea that, in zebrafish, paternally inherited centrosomes are required for the first cell division while maternally derived factors are required for centrosomal duplication and cell divisions in subsequent cell cycles. DNA synthesis ensues in the absence of centrosome duplication, and the one-cycle delay in the first cell division caused by cea mutant sperm leads to whole genome duplication. We discuss the potential implications of these findings with regards to the origin of polyploidization in animal species. In addition, the uncoupling of developmental time and cell division count caused by the cea mutation suggests the presence of a time window, normally corresponding to the first two cell cycles, which is permissive for germ plasm recruitment.

Disorders of erythrocyte hydration.

PubMed

Gallagher, Patrick G

2017-12-21

The erythrocyte contains a network of pathways that regulate salt and water content in the face of extracellular and intracellular osmotic perturbations. This allows the erythrocyte to maintain a narrow range of cell hemoglobin concentration, a process critical for normal red blood cell function and survival. Primary disorders that perturb volume homeostasis jeopardize the erythrocyte and may lead to its premature destruction. These disorders are marked by clinical, laboratory, and physiologic heterogeneity. Recent studies have revealed that these disorders are also marked by genetic heterogeneity. They have implicated roles for several proteins, PIEZO1, a mammalian mechanosensory protein; GLUT1, the glucose transporter; SLC4A1, the anion transporter; RhAG, the Rh-associated glycoprotein; KCNN4, the Gardos channel; and ABCB6, an adenosine triphosphate-binding cassette family member, in the maintenance of erythrocyte volume homeostasis. Secondary disorders of erythrocyte hydration include sickle cell disease, thalassemia, hemoglobin CC, and hereditary spherocytosis, where cellular dehydration may be a significant contributor to disease pathology and clinical complications. Understanding the pathways regulating erythrocyte water and solute content may reveal innovative strategies to maintain normal volume in disorders associated with primary or secondary cellular dehydration. These mechanisms will serve as a paradigm for other cells and may reveal new therapeutic targets for disease prevention and treatment beyond the erythrocyte. © 2017 by The American Society of Hematology.

PTEN expression and function in adult cancer stem cells and prospects for therapeutic targeting.

PubMed

Ciuffreda, Ludovica; Falcone, Italia; Incani, Ursula Cesta; Del Curatolo, Anais; Conciatori, Fabiana; Matteoni, Silvia; Vari, Sabrina; Vaccaro, Vanja; Cognetti, Francesco; Milella, Michele

2014-09-01

Phosphatase and tensin homolog deleted on chromosome ten (PTEN) is a non-redundant lipid phosphatase that restrains and fine tunes the phosphatidylinositol-3-kinase (PI3K) signaling pathway. PTEN is involved in inherited syndromes, which predispose to different types of cancers and is among the most frequently inactivated tumor suppressor genes in sporadic cancers. Indeed, loss of PTEN function occurs in a wide spectrum of human cancers through a variety of mechanisms, including mutations, deletions, transcriptional silencing, or protein instability. PTEN prevents tumorigenesis through multiple mechanisms and regulates a plethora of cellular processes, including survival, proliferation, energy metabolism and cellular architecture. Moreover, recent studies have demonstrated that PTEN is able to exit, exist, and function outside the cell, allowing for inhibition of the PI3K pathway in neighboring cells in a paracrine fashion. Most recently, studies have shown that PTEN is also critical for stem cell maintenance and that PTEN loss can lead to the emergence and proliferation of cancer stem cell (CSC) clones. Depending on the cellular and tissue context of origin, PTEN deletion may result in increased self-renewal capacity or normal stem cell exhaustion and PTEN-defìcient stem and progenitor cells have been reported in prostate, lung, intestinal, and pancreatic tissues before tumor formation; moreover, reversible or irreversible PTEN loss is frequently observed in CSC from a variety of solid and hematologic malignancies, where it may contribute to the functional phenotype of CSC. In this review, we will focus on the role of PTEN expression and function and downstream pathway activation in cancer stem cell biology and regulation of the tumorigenic potential; the emerging role of PTEN in mediating the crosstalk between the PI3K and MAPK pathways will also be discussed, together with prospects for the therapeutic targeting of tumors lacking PTEN expression. Copyright © 2014 Elsevier Ltd. All rights reserved.

Functional Mitochondria in Health and Disease.

PubMed

Herst, Patries M; Rowe, Matthew R; Carson, Georgia M; Berridge, Michael V

2017-01-01

The ability to rapidly adapt cellular bioenergetic capabilities to meet rapidly changing environmental conditions is mandatory for normal cellular function and for cancer progression. Any loss of this adaptive response has the potential to compromise cellular function and render the cell more susceptible to external stressors such as oxidative stress, radiation, chemotherapeutic drugs, and hypoxia. Mitochondria play a vital role in bioenergetic and biosynthetic pathways and can rapidly adjust to meet the metabolic needs of the cell. Increased demand is met by mitochondrial biogenesis and fusion of individual mitochondria into dynamic networks, whereas a decrease in demand results in the removal of superfluous mitochondria through fission and mitophagy. Effective communication between nucleus and mitochondria (mito-nuclear cross talk), involving the generation of different mitochondrial stress signals as well as the nuclear stress response pathways to deal with these stressors, maintains bioenergetic homeostasis under most conditions. However, when mitochondrial DNA (mtDNA) mutations accumulate and mito-nuclear cross talk falters, mitochondria fail to deliver critical functional outputs. Mutations in mtDNA have been implicated in neuromuscular and neurodegenerative mitochondriopathies and complex diseases such as diabetes, cardiovascular diseases, gastrointestinal disorders, skin disorders, aging, and cancer. In some cases, drastic measures such as acquisition of new mitochondria from donor cells occurs to ensure cell survival. This review starts with a brief discussion of the evolutionary origin of mitochondria and summarizes how mutations in mtDNA lead to mitochondriopathies and other degenerative diseases. Mito-nuclear cross talk, including various stress signals generated by mitochondria and corresponding stress response pathways activated by the nucleus are summarized. We also introduce and discuss a small family of recently discovered hormone-like mitopeptides that modulate body metabolism. Under conditions of severe mitochondrial stress, mitochondria have been shown to traffic between cells, replacing mitochondria in cells with damaged and malfunctional mtDNA. Understanding the processes involved in cellular bioenergetics and metabolic adaptation has the potential to generate new knowledge that will lead to improved treatment of many of the metabolic, degenerative, and age-related inflammatory diseases that characterize modern societies.

Microglia During Development and Aging

PubMed Central

Harry, G. Jean

2013-01-01

Microglia are critical nervous system-specific cells influencing brain development, maintenance of the neural environment, response to injury, and repair. They contribute to neuronal proliferation and differentiation, pruning of dying neurons, synaptic remodeling and clearance of debris and aberrant proteins. Colonization of the brain occurs during gestation with an expansion following birth with localization stimulated by programmed neuronal death, synaptic pruning, andaxonal degeneration. Changes inmicroglia phenotype relate to cellular processes including specific neurotransmitter, pattern recognition, or immune-related receptor activation. Upon activation, microglia cells have the capacity to release a number of substances, e.g., cytokines, chemokines, nitric oxide, and reactive oxygen species, which could be detrimental or beneficial to the surrounding cells. With aging, microglia shift their morphology and may display diminished capacity for normal functions related to migration, clearance, and the ability to shift from a pro-inflammatory to an anti-inflammatory state to regulate injury and repair. This shift in microgliapotentially contributes to increased susceptibility and neurodegeneration as a function of age. In the current review, information is provided on the colonization of the brain by microglia, the expression of various pattern recognition receptors to regulate migration and phagocytosis, and the shift in related functions that occur in normal aging. PMID:23644076

Cellular Mechanism of Inner Ear Genetic Disease, roles of Kv7.1 (KCNQ1) Channel

NASA Astrophysics Data System (ADS)

Mousavi Nik, Atefeh

Potassium channels are the most diverse and widely distributed membrane protein in all living organisms. They have various roles in the body such as controlling membrane potential, cell volume, and cell migration. Many studies have shown that mutation in these channels is associated with different diseases for example: Hearing Defect, Cardiac Arrhythmia, Episodic Ataxia, Seizure and Neuromyotonia. One of the most important diseases associated with K+ channel mutations is called Jervell and Lange-Nielsen syndrome (JLNS). This disease causes bilateral congenital deafness and the patients also suffer from Long QT and they usually experience syncopal episodes in their life and eventually die as a result of cardiac arrest. The gene KCNQ1 encodes the Kv7.1 voltage gated potassium channel. This channel expresses in apical membrane of marginal cell in stria vasularis of cochlea and secret K+ ion to endolymp to keep the endocochlear potential stable, which is necessary for the inner ear to function properly. Kv7.1 channel also expresses in cardiac myocytes and mutation in this gene is associated with another syndrome called Romano-Ward syndrome (RWS). Although Romano-Ward patients have mutation in KCNQ1, similar to Jervell and Lange-Nielsen patients, they only suffer from cardiac defect, and their hearing is completely normal. Several studies identified that mutations in Kv7.1 gene is associated with JLNS and RWS, but the biophysical and cellular mechanisms of these mutations are still unknown. To determine the cellular mechanisms of JLNS and RWS, and to provide mechanistic insight on the functional outputs of JLNS versus RWS mutations, we generated several mutant forms of the human Kv7.1 ( KCNQ1) clone, using site-directed mutagenesis to define their sub-cellular localization and examined their electrophysiological properties. We identified JLNS and RWS mutations at the S4-S5-linker, the pore loop (P-loop) and the C-terminus of hKv7.1 which have been found to control channel gating, permeation and modulation, respectively. The result showed that for JLNS, all P-loop and C-terminal mutations (seven mutations) yielded non-functional channels when expressed alone. Moreover, the W248F at the end of the S4 domain yielded a functional current, but it became inactivated at positive step potentials, therefore the channel essentially is non-functional. All the JLNS mutant channels are non-functional, and have impaired membrane trafficking. In contrast, the RWS mutants showed wide-ranging functional phenotypes consisting of channels with large current, channel with no measurable current when expressed alone, but significant current upon addition of the WT subunit and finally channel with no measurable current even in presence of WT subunit. The RWS mutants, however, produced dominant negative effect. These findings provide integrated cellular and molecular mechanisms of hKv7.1 functions and the consequent diseased phenotype in JLNS and RWS may come from the tissue-specific function of the channel.

Targeting polyamine metabolism for cancer therapy and prevention

PubMed Central

Murray-Stewart, Tracy R.; Woster, Patrick M.; Casero, Robert A.

2017-01-01

The chemically simple, biologically complex eukaryotic polyamines, spermidine and spermine, are positively charged alkylamines involved in many crucial cellular processes. Along with their diamine precursor putrescine, their normally high intracellular concentrations require fine attenuation by multiple regulatory mechanisms to keep these essential molecules within strict physiologic ranges. Since the metabolism of and requirement for polyamines are frequently dysregulated in neoplastic disease, the metabolic pathway and functions of polyamines provide rational drug targets; however, these targets have been difficult to exploit for chemotherapy. It is the goal of this article to review the latest findings in the field that demonstrate the potential utility of targeting the metabolism and function of polyamines as strategies for both chemotherapy and, possibly more importantly, chemoprevention. PMID:27679855

Plant uncoupling mitochondrial proteins.

PubMed

Vercesi, Aníbal Eugênio; Borecký, Jiri; Maia, Ivan de Godoy; Arruda, Paulo; Cuccovia, Iolanda Midea; Chaimovich, Hernan

2006-01-01

Uncoupling proteins (UCPs) are membrane proteins that mediate purine nucleotide-sensitive free fatty acid-activated H(+) flux through the inner mitochondrial membrane. After the discovery of UCP in higher plants in 1995, it was acknowledged that these proteins are widely distributed in eukaryotic organisms. The widespread presence of UCPs in eukaryotes implies that these proteins may have functions other than thermogenesis. In this review, we describe the current knowledge of plant UCPs, including their discovery, biochemical properties, distribution, gene family, gene expression profiles, regulation of gene expression, and evolutionary aspects. Expression analyses and functional studies on the plant UCPs under normal and stressful conditions suggest that UCPs regulate energy metabolism in the cellular responses to stress through regulation of the electrochemical proton potential (Deltamu(H)+) and production of reactive oxygen species.

Pharmacology of Antisense Drugs.

PubMed

Bennett, C Frank; Baker, Brenda F; Pham, Nguyen; Swayze, Eric; Geary, Richard S

2017-01-06

Recent studies have led to a greater appreciation of the diverse roles RNAs play in maintaining normal cellular function and how they contribute to disease pathology, broadening the number of potential therapeutic targets. Antisense oligonucleotides are the most direct means to target RNA in a selective manner and have become an established platform technology for drug discovery. There are multiple molecular mechanisms by which antisense oligonucleotides can be used to modulate RNAs in cells, including promoting the degradation of the targeted RNA or modulating RNA function without degradation. Antisense drugs utilizing various antisense mechanisms are demonstrating therapeutic potential for the treatment of a broad variety of diseases. This review focuses on some of the advances that have taken place in translating antisense technology from the bench to the clinic.

Molecular and functional characterization of CD133+ stem/progenitor cells infused in patients with end-stage liver disease reveals their interplay with stromal liver cells.

PubMed

Catani, Lucia; Sollazzo, Daria; Bianchi, Elisa; Ciciarello, Marilena; Antoniani, Chiara; Foscoli, Licia; Caraceni, Paolo; Giannone, Ferdinando Antonino; Baldassarre, Maurizio; Giordano, Rosaria; Montemurro, Tiziana; Montelatici, Elisa; D'Errico, Antonia; Andreone, Pietro; Giudice, Valeria; Curti, Antonio; Manfredini, Rossella; Lemoli, Roberto Massimo

2017-12-01

Growing evidence supports the therapeutic potential of bone marrow (BM)-derived stem/progenitor cells for end-stage liver disease (ESLD). We recently demonstrated that CD133 + stem/progenitor cell (SPC) reinfusion in patients with ESLD is feasible and safe and improve, albeit transiently, liver function. However, the mechanism(s) through which BM-derived SPCs may improve liver function are not fully elucidated. Here, we characterized the circulating SPCs compartment of patients with ESLD undergoing CD133 + cell therapy. Next, we set up an in vitro model mimicking SPCs/liver microenvironment interaction by culturing granulocyte colony-stimulating factor (G-CSF)-mobilized CD133 + and LX-2 hepatic stellate cells. We found that patients with ESLD show normal basal levels of circulating hematopoietic and endothelial progenitors with impaired clonogenic ability. After G-CSF treatment, patients with ESLD were capable to mobilize significant numbers of functional multipotent SPCs, and interestingly, this was associated with increased levels of selected cytokines potentially facilitating SPC function. Co-culture experiments showed, at the molecular and functional levels, the bi-directional cross-talk between CD133 + SPCs and human hepatic stellate cells LX-2. Human hepatic stellate cells LX-2 showed reduced activation and fibrotic potential. In turn, hepatic stellate cells enhanced the proliferation and survival of CD133 + SPCs as well as their endothelial and hematopoietic function while promoting an anti-inflammatory profile. We demonstrated that the interaction between CD133 + SPCs from patients with ESLD and hepatic stellate cells induces significant functional changes in both cellular types that may be instrumental for the improvement of liver function in cirrhotic patients undergoing cell therapy. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

The Caenorhabditis elegans rsd-2 and rsd-6 genes are required for chromosome functions during exposure to unfavorable environments.

PubMed

Han, Wang; Sundaram, Prema; Kenjale, Himanshu; Grantham, James; Timmons, Lisa

2008-04-01

In Caenorhabditis elegans, exogenous dsRNA can elicit systemic RNAi, a process that requires the function of many genes. Considering that the activities of many of these genes are also required for normal development, it is surprising that exposure to high concentrations of dsRNA does not elicit adverse consequences to animals. Here, we report inducible phenotypes in attenuated C. elegans strains reared in environments that include nonspecific dsRNA and elevated temperature. Under these conditions, chromosome integrity is compromised in RNAi-defective strains harboring mutations in rsd-2 or rsd-6. Specifically, rsd-2 mutants display defects in transposon silencing, while meiotic chromosome disjunction is affected in rsd-6 mutants. RSD-2 proteins localize to multiple cellular compartments, including the nucleolus and cytoplasmic compartments that, in part, are congruent with calreticulin and HAF-6. We considered that the RNAi defects in rsd-2 mutants might have relevance to membrane-associated functions; however, endomembrane compartmentalization and endocytosis/exocytosis markers in rsd-2 and rsd-6 mutants appear normal. The mutants also possess environmentally sensitive defects in cell-autonomous RNAi elicited from transgene-delivered dsRNAs. Thus, the ultimate functions of rsd-2 and rsd-6 in systemic RNAi are remarkably complex and environmentally responsive.

[Effect of electroacupuncture on cellular structure of hippocampus in splenic asthenia pedo-rats].

PubMed

Yang, Zhuo-xin; Zhuo, Yuan-yuan; Yu, Hai-bo; Wang, Ning

2010-02-01

To observe the effect of electroacupuncture (EA) on hippocampal structure in splenic asthenia pedo-rats. A total of 15 SD male rats were randomly assigned to normal control group (n=5), model group (n=5) and EA group (n=5). Splenic asthenic syndrome model was established by intragastric administration of rhubarb and intraperitoneal injection of Reserpine for 14 d. EA (1 mA, 3 Hz/iS Hz) was applied to bilateral "Zusanli" (ST 36) and "Sanyinjiao" (SP 6) for 20 mm, once a day for 14 days. The cellular structure of hippocampus was observed by light microscope and transmission electron microscope. Optical microscopic observation showed that in normal control group, the cellular nucleus was distinct, and the granular cell layer well-arranged and tight. In model group, the intracellular space was widened, and the granular cell layer was out of order in the arrangement. In EA group, the celluldr nucleus and the granular cell layer were nearly normal. Results of the electronic microscope showed that cells in model group had a karyopyknosis with irregular appearance and clear incisure, and some of them presented dissolving and necrotic phenomena; and those in EA group were milder in injury, had nearly-normal nucleus with visible nucleoli and relatively-intact nuclear membrane. Regarding the cellular plasma, in comparison with rich normal organelles of control group, the mitochondria in model group were swelling, with vague, dissolved and broken cristae, while in EA group, majority of the organelles were well-kept, and slightly dissolved mitochondrial cristae found. In regard to the synaptic structure, in comparison with control group, synaptic apomorphosis and swelling mitochondria were found in model group While in EA group, milder swelling and hydropic degeneration were seen. Different from the distinct pre- and post-synaptic membrane and synaptic vesicles of control group, while those in EA group were nearly-normal. electroacupunture can effectively relieve splenasthenic syndrome induced pathohistological changes of neurons of the hippocampus in the rat.

The immune system which adversely alter thyroid functions: a review on the concept of autoimmunity.

PubMed

Mansourian, Azad Reza

2010-08-15

The immune system protect individual from many pathogens exists within our environment and in human body, by destroying them through molecular and cellular mechanism of B and T cells of immune system. Autoimmunity is an adverse relation of immune system against non- foreign substances leaving behind either alters the normal function or destroying the tissue involved. Autoimmunity occur in genetically predispose persons with familial connections. The autoimmunity to the thyroid gland mainly consists of Hashimato thyroiditis and Grave's disease, the two end of spectrum in thyroid function of hypo and hyperactivity, respectively. The thyroid stimulating hormone receptor, thyroglobuline, enzymes of thyroid hormones synthesis are targeted by autoantibodies and cell- mediated reactions. The aim of this review is to explore the studies reported on the autoimmunity to the thyroid gland.

Mediator complex dependent regulation of cardiac development and disease.

PubMed

Grueter, Chad E

2013-06-01

Cardiovascular disease (CVD) is a leading cause of morbidity and mortality. The risk factors for CVD include environmental and genetic components. Human mutations in genes involved in most aspects of cardiovascular function have been identified, many of which are involved in transcriptional regulation. The Mediator complex serves as a pivotal transcriptional regulator that functions to integrate diverse cellular signals by multiple mechanisms including recruiting RNA polymerase II, chromatin modifying proteins and non-coding RNAs to promoters in a context dependent manner. This review discusses components of the Mediator complex and the contribution of the Mediator complex to normal and pathological cardiac development and function. Enhanced understanding of the role of this core transcriptional regulatory complex in the heart will help us gain further insights into CVD. Copyright © 2013. Production and hosting by Elsevier Ltd.

Breaking into the epithelial apical–junctional complex — news from pathogen hackers

PubMed Central

Vogelmann, Roger; Amieva, Manuel R; Falkow, Stanley; Nelson, W James

2012-01-01

The epithelial apical–junctional complex is a key regulator of cellular functions. In addition, it is an important target for microbial pathogens that manipulate the cell to survive, proliferate and sometimes persist within a host. Out of a myriad of potential molecular targets, some bacterial and viral pathogens have selected a subset of protein targets at the apical–junctional complex of epithelial cells. Studying how microbes use these targets also teaches us about the inherent physiological properties of host molecules in the context of normal junctional structure and function. Thus, we have learned that three recently uncovered components of the apical–junctional complex of the Ig superfamily — junctional adhesion molecule, Nectin and the coxsackievirus and adenovirus receptor — are important regulators of junction structure and function and represent critical targets of microbial virulence gene products. PMID:15037310

Breaking into the epithelial apical-junctional complex--news from pathogen hackers.

PubMed

Vogelmann, Roger; Amieva, Manuel R; Falkow, Stanley; Nelson, W James

2004-02-01

The epithelial apical-junctional complex is a key regulator of cellular functions. In addition, it is an important target for microbial pathogens that manipulate the cell to survive, proliferate and sometimes persist within a host. Out of a myriad of potential molecular targets, some bacterial and viral pathogens have selected a subset of protein targets at the apical-junctional complex of epithelial cells. Studying how microbes use these targets also teaches us about the inherent physiological properties of host molecules in the context of normal junctional structure and function. Thus, we have learned that three recently uncovered components of the apical-junctional complex of the Ig superfamily--junctional adhesion molecule, Nectin and the coxsackievirus and adenovirus receptor--are important regulators of junction structure and function and represent critical targets of microbial virulence gene products.

[Cellular architecture of papillary and nonpapillary transitional cell carcinoma].

PubMed

Moriyama, M

1989-07-01

To characterize the cellular architecture of papillary and nonpapillary transitional cell carcinoma. 2 normal ureters, 6 papillary bladder cancers and 5 nonpapillary bladder cancers were subjected to light and electron microscopic study as well as three dimensional reconstruction by 0.5 microns thick serial sections. Normal urothelium consisted of three cell layers of the basal, intermediate and superficial cells, each of which was morphologically characterized in terms of cell shape and development of cell organelles. Over 90% of the epithelial cells were proved to be connected to the uniform basement membrane directly or with long, fine cytoplasmic processes, forming hemidesmosomes at the junctional portion. Papillary tumors had, as a rule, the same cellular architecture as that of normal epithelium in terms of the regularity of cellular polarity, arrangement and differentiation, and the connection to the basement membrane. But, in G2 tumors, the connection between the intermediate and superficial cells and the basement membrane failed to be confirmed in 7 to 44% of the cells, suggesting the heterogeneity of the tumors. In contrast, nonpapillary tumors showed a high irregularity of the cellular architecture in both lesions of stromal and intra-epithelial invasion. The development of the basement membrane was indefinite, often showing thinning or disruption where occasional cytoplasmic protrusion of the tumor cells into the lamina propria was found. Nearly all of the intermediate and superficial cells in the intraepithelial lesions proved not to communicate with the basement membrane. The present results indicate distinct differences of cellular architecture between the papillary and nonpapillary urothelial tumors, which may reflect not only the growth pattern but also the biological behaviour of the individual tumors.

Mitochondrial Fission and Autophagy in the Normal and Diseased Heart

PubMed Central

Iglewski, Myriam; Hill, Joseph A.; Lavandero, Sergio; Rothermel, Beverly A.

2011-01-01

Sustained hypertension promotes structural, functional and metabolic remodeling of cardiomyocyte mitochondria. As long-lived, postmitotic cells, cardiomyocytes turn over mitochondria continuously to compensate for changes in energy demands and to remove damaged organelles. This process involves fusion and fission of existing mitochondria to generate new organelles and separate old ones for degradation via autophagy. Autophagy is a lysosome-dependent proteolytic pathway capable of processing cellular components, including organelles and protein aggregates. Autophagy can be either nonselective or selective and contributes to remodeling of the myocardium under stress. Fission of mitochondria, loss of membrane potential, and ubiquitination are emerging as critical steps that direct selective autophagic degradation of mitochondria. This review discusses the molecular mechanisms controlling mitochondrial dynamics, including fission, fusion, transport, and degradation. Furthermore, it examines recent studies revealing the importance of these processes in normal and diseased heart. PMID:20865352

AMPKα2 Suppresses Murine Embryonic Fibroblast Transformation and Tumorigenesis

PubMed Central

Phoenix, Kathryn N.; Devarakonda, Charan V.; Fox, Melissa M.; Stevens, Laura E.

2012-01-01

AMP-activated kinase (AMPK) is a key metabolic sensor and stress signaling kinase. AMPK activity is known to suppress anabolic processes such as protein and lipid biosynthesis and promote energy-producing pathways including fatty acid oxidation, resulting in increased cellular energy. In addition, AMPK localizes to centrosomes during cell division, plays a role in cellular polarization, and directly targets p53, affecting apoptosis. Two distinct catalytic AMPKα isoforms exist: α1 and α2. Multiple reports indicate that both common and distinct functions exist for each of the 2 α isoforms. AMPK activation has been shown to repress tumor growth, and it has been suggested that AMPK may function as a metabolic tumor suppressor. To evaluate the potential role of each of the AMPKα isoforms in modulating cellular transformation, susceptibility to Ras-induced transformation was evaluated in normal murine embryonic fibroblasts (MEFs) obtained from genetically deleted AMPKα1- or AMPKα2-null mice. This study demonstrated that while AMPKα1 is the dominant AMPK isoform expressed in MEFs, only the AMPKα2-null MEFs displayed increased susceptibility to H-RasV12 transformation in vitro and tumorigenesis in vivo. Conversely, AMPKα1-null MEFs, which demonstrated compensation with increased expression of AMPKα2, displayed minimal transformation susceptibility, decreased cell survival, decreased cell proliferation, and increased apoptosis. Finally, this study demonstrates that AMPKα2 was selectively responsible for targeting p53, thus contributing to the suppression of transformation and tumorigenic mechanisms. PMID:22893790

From Evolution to Revolution: miRNAs as Pharmacological Targets for Modulating Cholesterol Efflux and Reverse Cholesterol Transport

PubMed Central

Dávalos, Alberto; Fernández-Hernando, Carlos

2013-01-01

There has been strong evolutionary pressure to ensure that an animal cell maintain levels of cholesterol within tight limits for normal function. Imbalances in cellular cholesterol levels are a major player in the development of different pathologies associated to dietary excess. Although epidemiological studies indicate that elevated levels of high-density lipoprotein (HDL)-cholesterol reduce the risk of cardiovascular disease, recent genetic evidence and pharmacological therapies to raise HDL levels do not support their beneficial effects. Cholesterol efflux as the first and probably the most important step in reverse cholesterol transport is an important biological process relevant to HDL function. Small non-coding RNAs (microRNAs), post-transcriptional control different aspects of cellular cholesterol homeostasis including cholesterol efflux. miRNA families miR-33, miR-758, miR-10b, miR-26 and miR-106b directly modulates cholesterol efflux by targeting the ATP-binding cassette transporter A1 (ABCA1). Pre-clinical studies with anti-miR therapies to inhibit some of these miRNAs have increased cellular cholesterol efflux, reverse cholesterol transport and reduce pathologies associated to dyslipidemia. Although miRNAs as therapy have benefits from existing antisense technology, different obstacles need to be solved before we incorporate such research into clinical care. Here we focus on the clinical potential of miRNAs as therapeutic target to increase cholesterol efflux and reverse cholesterol transport as a new alternative to ameliorate cholesterol-related pathologies. PMID:23435093

Mutations in SNX14 Cause a Distinctive Autosomal-Recessive Cerebellar Ataxia and Intellectual Disability Syndrome

PubMed Central

Thomas, Anna C.; Williams, Hywel; Setó-Salvia, Núria; Bacchelli, Chiara; Jenkins, Dagan; O’Sullivan, Mary; Mengrelis, Konstantinos; Ishida, Miho; Ocaka, Louise; Chanudet, Estelle; James, Chela; Lescai, Francesco; Anderson, Glenn; Morrogh, Deborah; Ryten, Mina; Duncan, Andrew J.; Pai, Yun Jin; Saraiva, Jorge M.; Ramos, Fabiana; Farren, Bernadette; Saunders, Dawn; Vernay, Bertrand; Gissen, Paul; Straatmaan-Iwanowska, Anna; Baas, Frank; Wood, Nicholas W.; Hersheson, Joshua; Houlden, Henry; Hurst, Jane; Scott, Richard; Bitner-Glindzicz, Maria; Moore, Gudrun E.; Sousa, Sérgio B.; Stanier, Philip

2014-01-01

Intellectual disability and cerebellar atrophy occur together in a large number of genetic conditions and are frequently associated with microcephaly and/or epilepsy. Here we report the identification of causal mutations in Sorting Nexin 14 (SNX14) found in seven affected individuals from three unrelated consanguineous families who presented with recessively inherited moderate-severe intellectual disability, cerebellar ataxia, early-onset cerebellar atrophy, sensorineural hearing loss, and the distinctive association of progressively coarsening facial features, relative macrocephaly, and the absence of seizures. We used homozygosity mapping and whole-exome sequencing to identify a homozygous nonsense mutation and an in-frame multiexon deletion in two families. A homozygous splice site mutation was identified by Sanger sequencing of SNX14 in a third family, selected purely by phenotypic similarity. This discovery confirms that these characteristic features represent a distinct and recognizable syndrome. SNX14 encodes a cellular protein containing Phox (PX) and regulator of G protein signaling (RGS) domains. Weighted gene coexpression network analysis predicts that SNX14 is highly coexpressed with genes involved in cellular protein metabolism and vesicle-mediated transport. All three mutations either directly affected the PX domain or diminished SNX14 levels, implicating a loss of normal cellular function. This manifested as increased cytoplasmic vacuolation as observed in cultured fibroblasts. Our findings indicate an essential role for SNX14 in neural development and function, particularly in development and maturation of the cerebellum. PMID:25439728

Mutations in SNX14 cause a distinctive autosomal-recessive cerebellar ataxia and intellectual disability syndrome.

PubMed

Thomas, Anna C; Williams, Hywel; Setó-Salvia, Núria; Bacchelli, Chiara; Jenkins, Dagan; O'Sullivan, Mary; Mengrelis, Konstantinos; Ishida, Miho; Ocaka, Louise; Chanudet, Estelle; James, Chela; Lescai, Francesco; Anderson, Glenn; Morrogh, Deborah; Ryten, Mina; Duncan, Andrew J; Pai, Yun Jin; Saraiva, Jorge M; Ramos, Fabiana; Farren, Bernadette; Saunders, Dawn; Vernay, Bertrand; Gissen, Paul; Straatmaan-Iwanowska, Anna; Baas, Frank; Wood, Nicholas W; Hersheson, Joshua; Houlden, Henry; Hurst, Jane; Scott, Richard; Bitner-Glindzicz, Maria; Moore, Gudrun E; Sousa, Sérgio B; Stanier, Philip

2014-11-06

Intellectual disability and cerebellar atrophy occur together in a large number of genetic conditions and are frequently associated with microcephaly and/or epilepsy. Here we report the identification of causal mutations in Sorting Nexin 14 (SNX14) found in seven affected individuals from three unrelated consanguineous families who presented with recessively inherited moderate-severe intellectual disability, cerebellar ataxia, early-onset cerebellar atrophy, sensorineural hearing loss, and the distinctive association of progressively coarsening facial features, relative macrocephaly, and the absence of seizures. We used homozygosity mapping and whole-exome sequencing to identify a homozygous nonsense mutation and an in-frame multiexon deletion in two families. A homozygous splice site mutation was identified by Sanger sequencing of SNX14 in a third family, selected purely by phenotypic similarity. This discovery confirms that these characteristic features represent a distinct and recognizable syndrome. SNX14 encodes a cellular protein containing Phox (PX) and regulator of G protein signaling (RGS) domains. Weighted gene coexpression network analysis predicts that SNX14 is highly coexpressed with genes involved in cellular protein metabolism and vesicle-mediated transport. All three mutations either directly affected the PX domain or diminished SNX14 levels, implicating a loss of normal cellular function. This manifested as increased cytoplasmic vacuolation as observed in cultured fibroblasts. Our findings indicate an essential role for SNX14 in neural development and function, particularly in development and maturation of the cerebellum. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Enhanced Medial Collateral Ligament Healing using Mesenchymal Stem Cells: Dosage Effects on Cellular Response and Cytokine Profile

PubMed Central

Saether, Erin E.; Chamberlain, Connie S.; Leiferman, Ellen M.; Kondratko-Mittnacht, Jaclyn R.; Li, Wan Ju; Brickson, Stacey L.; Vanderby, Ray

2013-01-01

Mesenchymal stem cells (MSCs) have potential therapeutic applications for musculoskeletal injuries due to their ability to differentiate into several tissue cell types and modulate immune and inflammatory responses. These immune-modulatory properties were examined in vivo during early stage rat medial collateral ligament healing. Two different cell doses (low dose 1×106 or high dose 4×106 MSCs) were administered at the time of injury and compared with normal ligament healing at days 5 and 14 post-injury. At both times, the high dose MSC group demonstrated a significant decrease in M2 macrophages compared to controls. At day 14, fewer M1 macrophages were detected in the low dose group compared to the high dose group. These results, along with significant changes in procollagen I, proliferating cells, and endothelialization suggest that MSCs can alter the cellular response during healing in a dose-dependent manner. The higher dose ligaments also had increased expression of several pro-inflammatory cytokines at day 5 (IL-1β, IFNγ, IL-2) and increased expression of IL-12 at day 14. Mechanical testing at day 14 revealed increased failure strength and stiffness in low dose ligaments compared to controls. Based on these improved mechanical properties, MSCs enhanced functional healing when applied at a lower dose. Different doses of MSCs uniquely affected the cellular response and cytokine expression in healing ligaments. Interestingly, the lower dose of cells proved to be most effective in improving functional properties. PMID:24174129

Alginic acid cell entrapment: a novel method for measuring in vivo macrophage cholesterol homeostasis

PubMed Central

Sontag, Timothy J.; Chellan, Bijoy; Bhanvadia, Clarissa V.; Getz, Godfrey S.; Reardon, Catherine A.

2015-01-01

Macrophage conversion to atherosclerotic foam cells is partly due to the balance of uptake and efflux of cholesterol. Cholesterol efflux from cells by HDL and its apoproteins for subsequent hepatic elimination is known as reverse cholesterol transport. Numerous methods have been developed to measure in vivo macrophage cholesterol efflux. Most methods do not allow for macrophage recovery for analysis of changes in cellular cholesterol status. We describe a novel method for measuring cellular cholesterol balance using the in vivo entrapment of macrophages in alginate, which retains incorporated cells while being permeable to lipoproteins. Recipient mice were injected subcutaneously with CaCl2 forming a bubble into which a macrophage/alginate suspension was injected, entrapping the macrophages. Cells were recovered after 24 h. Cellular free and esterified cholesterol mass were determined enzymatically and normalized to cellular protein. Both normal and cholesterol loaded macrophages undergo measureable changes in cell cholesterol when injected into WT and apoA-I-, LDL-receptor-, or apoE-deficient mice. Cellular cholesterol balance is dependent on initial cellular cholesterol status, macrophage cholesterol transporter expression, and apolipoprotein deficiency. Alginate entrapment allows for the in vivo measurement of macrophage cholesterol homeostasis and is a novel platform for investigating the role of genetics and therapeutic interventions in atherogenesis. PMID:25465389

Putting tumours in context

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

Bissell, Mina J.; Radisky, Derek

2001-10-01

The interactions between cancer cells and their micro- and macroenvironment create a context that promotes tumor growth and protects it from immune attack. The functional association of cancer cells with their surrounding tissues forms a new 'organ' that changes as malignancy progresses. Investigation of this process might provide new insights into the mechanisms of tumorigenesis and could also lead to new therapeutic targets. Under normal conditions, ORGANS are made up of TISSUES that exchange information with other cell types via cell-cell contact, cytokines and the EXTRACELLULAR MATRIX (ECM). The ECM, which is produced by collaboration between STROMAL fibroblasts and EPITHELIALmore » cells, provides structural scaffolding for cells, as well as contextual information. The endothelial vasculature provides nutrients and oxygen, and cells of the immune system combat pathogens and remove apoptotic cells. Epithelial cells associate into intact, polarized sheets. These tissues communicate through a complex network of interactions: physically, through direct contact or through the intervening ECM, and biochemically, through both soluble and insoluble signalling molecules. In combination, these interactions provide the information that is necessary to maintain cellular differentiation and to create complex tissue structures. Occasionally, the intercellular signals that define the normal context become disrupted. Alterations in epithelial tissues can lead to movement of epithelial sheets and proliferation - for example, after activation of mesenchymal fibroblasts due to wounding.Normally, these conditions are temporary and reversible, but when inflammation is sustained, an escalating feedback loop ensues.Under persistent inflammatory conditions, continual upregulation of enzymes such as matrix metalloproteinases (MMPs) by stromal fibroblasts can disrupt the ECM, and invading immune cells can overproduce factors that promote abnormal proliferation. As this process progresses, the normal organization of the organ is replaced by a functional disorder. If there are pre-existing epithelial cells within this changing context that possess tumorigenic potential, they can start to proliferate. Alternatively, the abnormal interactions might lead to genomic instability within the epithelial cells and the acquisition of tumorigenic potential. The proliferating cancer cells can then interact with their microenvironment and enhance the abnormal interactions. At this point, the tumor has become its own organ, with a distinct context that now defines all its cellular responses. Here, we will examine how the mechanisms that contribute to the normal context also act to suppress developing tumors, how disruption of this context initiates and supports the process of tumorigenicity, and how some cells with a tumorigenic genotype can become phenotypically normal if the context is appropriately manipulated.« less

Investigating cell mechanics with atomic force microscopy

PubMed Central

Haase, Kristina; Pelling, Andrew E.

2015-01-01

Transmission of mechanical force is crucial for normal cell development and functioning. However, the process of mechanotransduction cannot be studied in isolation from cell mechanics. Thus, in order to understand how cells ‘feel’, we must first understand how they deform and recover from physical perturbations. Owing to its versatility, atomic force microscopy (AFM) has become a popular tool to study intrinsic cellular mechanical properties. Used to directly manipulate and examine whole and subcellular reactions, AFM allows for top-down and reconstitutive approaches to mechanical characterization. These studies show that the responses of cells and their components are complex, and largely depend on the magnitude and time scale of loading. In this review, we generally describe the mechanotransductive process through discussion of well-known mechanosensors. We then focus on discussion of recent examples where AFM is used to specifically probe the elastic and inelastic responses of single cells undergoing deformation. We present a brief overview of classical and current models often used to characterize observed cellular phenomena in response to force. Both simple mechanistic models and complex nonlinear models have been used to describe the observed cellular behaviours, however a unifying description of cell mechanics has not yet been resolved. PMID:25589563

Smoking, p53 Mutation, and Lung Cancer

PubMed Central

Gibbons, Don L.; Byers, Lauren A.; Kurie, Jonathan M.

2014-01-01

This issue marks the 50th Anniversary of the release of the U.S. Surgeon General’s Report on Smoking and Health. Perhaps no other singular event has done more to highlight the effects of smoking on the development of cancer. Tobacco exposure is the leading cause of cancers involving the oral cavity, conductive airways and the lung. Owing to the many carcinogens in tobacco smoke, smoking-related malignancies have a high genome-wide burden of mutations, including in the gene encoding for p53. The p53 protein is the most frequently mutated tumor suppressor in cancer, responsible for a range of critical cellular functions that are compromised by the presence of a mutation. Herein we review the epidemiologic connection between tobacco exposure and cancer, the molecular basis of p53 mutation in lung cancer, and the normal molecular and cellular roles of p53 that are abrogated during lung tumor development and progression as defined by in vitro and in vivo studies. We also consider the therapeutic potential of targeting mutant p53 in a clinical setting based upon the cellular role of mutant p53 and data from genetic murine models. PMID:24442106

Cellular redox dysfunction in the development of cardiovascular diseases.

PubMed

Kanaan, Georges N; Harper, Mary-Ellen

2017-11-01

To meet its exceptionally high energy demands, the heart relies largely on fatty acid oxidation, which then drives the oxidative phosphorylation system in mitochondria. Each day, this system produces about 6kg of ATP to sustain heart function. Fatty acid oxidation is sometimes associated with high rates of mitochondrial reactive oxygen species (ROS) production. By definition, ROS are singlet electron intermediates formed during the partial reduction of oxygen to water and they include radical and non-radical intermediates like superoxide, hydrogen peroxide and hydroxyl radical. Superoxide can also interact with nitric oxide to produce peroxynitrite that in turn can give rise to other radical or non-radical reactive nitrogen species (RNS) like nitrogen dioxide, dinitrogen trioxide and others. While mitochondrial and cellular functions can be impaired by ROS if they accumulate, under normal physiological conditions ROS are important signaling molecules in the cardiovascular system. A fine balance between ROS production and antioxidant systems, including glutathione redox, is essential in the heart; otherwise the ensuing damage can contribute to pathogenic processes, which can culminate in endothelial dysfunction, atherosclerosis, hypertension, cardiac hypertrophy, arrhythmias, myocardial ischemia/reperfusion damage, and heart failure. Here we provide a succinct review of recent findings. Copyright © 2017 Elsevier B.V. All rights reserved.

Mammalian Cell-Based Sensor System

NASA Astrophysics Data System (ADS)

Banerjee, Pratik; Franz, Briana; Bhunia, Arun K.

Use of living cells or cellular components in biosensors is receiving increased attention and opens a whole new area of functional diagnostics. The term "mammalian cell-based biosensor" is designated to biosensors utilizing mammalian cells as the biorecognition element. Cell-based assays, such as high-throughput screening (HTS) or cytotoxicity testing, have already emerged as dependable and promising approaches to measure the functionality or toxicity of a compound (in case of HTS); or to probe the presence of pathogenic or toxigenic entities in clinical, environmental, or food samples. External stimuli or changes in cellular microenvironment sometimes perturb the "normal" physiological activities of mammalian cells, thus allowing CBBs to screen, monitor, and measure the analyte-induced changes. The advantage of CBBs is that they can report the presence or absence of active components, such as live pathogens or active toxins. In some cases, mammalian cells or plasma membranes are used as electrical capacitors and cell-cell and cell-substrate contact is measured via conductivity or electrical impedance. In addition, cytopathogenicity or cytotoxicity induced by pathogens or toxins resulting in apoptosis or necrosis could be measured via optical devices using fluorescence or luminescence. This chapter focuses mainly on the type and applications of different mammalian cell-based sensor systems.

Downregulated microRNA-510-5p acts as a tumor suppressor in renal cell carcinoma.

PubMed

Chen, Duqun; Li, Yuchi; Yu, Zuhu; Li, Yifan; Su, Zhengming; Ni, Liangchao; Yang, Shangqi; Gui, Yaoting; Lai, Yongqing

2015-08-01

MicroRNA (miR)-510-5p has been demonstrated to be involved in a number of types of malignancy; however, the function of miR-510-5p in renal cancer remains unclear. The present study aimed to determine the expression of miR-510-5p in renal cell carcinoma (RCC) specimens and analyzed the impact of miR-510-5p on renal cancer by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, wound scratch and apoptosis assays. The results showed that miR-510-5p was significantly downregulated in RCC specimens compared with normal renal specimens. Overexpression of miR-510-5p by synthetic mature mimics reduced cell proliferation and migration and induced an increase in cell apoptosis, indicating that miR-510-5p may act as a tumor suppressor in RCC. The present study firstly revealed that downregulated miR-510-5p functioned as a tumor suppressor by reducing cellular proliferation and migration, and inducing apoptosis in RCC. Further research is required to define target genes of miR-510-5p to determine the cellular mechanism of miR-510-5p in the carcinogenesis of RCC.

The innate capacity of proteins to protect against reactive radical species.

PubMed

Hamdy, Omar M; Alizadeh, Arman; Julian, Ryan R

2015-08-07

Maintaining redox homeostasis, or the balance of oxidant and antioxidant forces, is essential for proper cellular functioning in biology. Although the antioxidant nature of many small molecules such as vitamin c and glutathione have been thoroughly investigated, contributions to redox homeostasis from larger biomolecules have received less attention. Evidence has shown that some proteins are antioxidant (in a non-catalytic sense), but large scale examination of this property for a diverse set of proteins has proven difficult. Herein, radical-directed dissociation mass spectrometry (RDD-MS) is used to examine the antioxidant capacity of a series of proteins with diverse biological roles, persistence intervals, and localizations. Digestion of these proteins reveals that all contain antioxidant peptide regions. Examination of the amino acid content of the antioxidant peptides does not reveal significant differences relative to normal peptides, suggesting that sequence may be more important than residue content. Sequence homology analysis across organisms reveals that antioxidant regions are frequently conserved, although many of these regions are also known to have other functions which may have influenced evolutionary pressure. Regardless of the origin, it is clear that many proteins may play secondary roles as sacrificial antioxidants within the cellular milieu.

Blood biomarkers for brain injury: What are we measuring?

PubMed Central

Kawata, Keisuke; Liu, Charles Y.; Merkel, Steven F.; Ramirez, Servio H.; Tierney, Ryan T.; Langford, Dianne

2016-01-01

Accurate diagnosis for mild traumatic brain injury (mTBI) remains challenging, as prognosis and return-to-play/work decisions are based largely on patient reports. Numerous investigations have identified and characterized cellular factors in the blood as potential biomarkers for TBI, in the hope that these factors may be used to gauge the severity of brain injury. None of these potential biomarkers have advanced to use in the clinical setting. Some of the most extensively studied blood biomarkers for TBI include S100β, neuron-specific enolase, glial fibrillary acidic protein, and Tau. Understanding the biological function of each of these factors may be imperative to achieve progress in the field. We address the basic question: what are we measuring? This review will discuss blood biomarkers in terms of cellular origin, normal and pathological function, and possible reasons for increased blood levels. Considerations in the selection, evaluation, and validation of potential biomarkers will also be addressed, along with mechanisms that allow brain-derived proteins to enter the bloodstream after TBI. Lastly, we will highlight perspectives and implications for repetitive neurotrauma in the field of blood biomarkers for brain injury. PMID:27181909

Time-resolved optical imaging provides a molecular snapshot of altered metabolic function in living human cancer cell models

NASA Astrophysics Data System (ADS)

Sud, Dhruv; Zhong, Wei; Beer, David G.; Mycek, Mary-Ann

2006-05-01

A fluorescence lifetime imaging microscopy (FLIM) method was developed and applied to investigate metabolic function in living human normal esophageal (HET-1) and Barrett’s adenocarcinoma (SEG-1) cells. In FLIM, image contrast is based on fluorophore excited state lifetimes, which reflect local biochemistry and molecular activity. Unique FLIM system attributes, including variable ultrafast time gating (≥ 200 ps), wide spectral tunability (337.1 - 960 nm), large temporal dynamic range (≥ 600 ps), and short data acquisition and processing times (15 s), enabled the study of two key molecules consumed at the termini of the oxidative phosphorylation pathway, NADH and oxygen, in living cells under controlled and calibrated environmental conditions. NADH is an endogenous cellular fluorophore detectable in living human tissues that has been shown to be a quantitative biomarker of dysplasia in the esophagus. Lifetime calibration of an oxygen-sensitive, ruthenium-based cellular stain enabled in vivo oxygen level measurements with a resolution of 8 μM over the entire physiological range (1 - 300 μM). Starkly higher intracellular oxygen and NADH levels in living SEG-1 vs. HET-1 cells were detected by FLIM and attributed to altered metabolic pathways in malignant cells.

High-fat diet induces cardiac remodelling and dysfunction: assessment of the role played by SIRT3 loss.

PubMed

Zeng, Heng; Vaka, Venkata Ramana; He, Xiaochen; Booz, George W; Chen, Jian-Xiong

2015-08-01

Mitochondrial dysfunction plays an important role in obesity-induced cardiac impairment. SIRT3 is a mitochondrial protein associated with increased human life span and metabolism. This study investigated the functional role of SIRT3 in obesity-induced cardiac dysfunction. Wild-type (WT) and SIRT3 knockout (KO) mice were fed a normal diet (ND) or high-fat diet (HFD) for 16 weeks. Body weight, fasting glucose levels, reactive oxygen species (ROS) levels, myocardial capillary density, cardiac function and expression of hypoxia-inducible factor (HIF)-1α/-2α were assessed. HFD resulted in a significant reduction in SIRT3 expression in the heart. Both HFD and SIRT3 KO mice showed increased ROS formation, impaired HIF signalling and reduced capillary density in the heart. HFD induced cardiac hypertrophy and impaired cardiac function. SIRT3 KO mice fed HFD showed greater ROS production and a further reduction in cardiac function compared to SIRT3 KO mice on ND. Thus, the adverse effects of HFD on cardiac function were not attributable to SIRT3 loss alone. However, HFD did not further reduce capillary density in SIRT3 KO hearts, implicating SIRT3 loss in HFD-induced capillary rarefaction. Our study demonstrates the importance of SIRT3 in preserving heart function and capillary density in the setting of obesity. Thus, SIRT3 may be a potential therapeutic target for obesity-induced heart failure. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Increased cFLIP expression in thymic epithelial tumors blocks autophagy via NF-κB signalling.

PubMed

Belharazem, Djeda; Grass, Albert; Paul, Cornelia; Vitacolonna, Mario; Schalke, Berthold; Rieker, Ralf J; Körner, Daniel; Jungebluth, Philipp; Simon-Keller, Katja; Hohenberger, Peter; Roessner, Eric M; Wiebe, Karsten; Gräter, Thomas; Kyriss, Thomas; Ott, German; Geserick, Peter; Leverkus, Martin; Ströbel, Philipp; Marx, Alexander

2017-10-27

The anti-apoptotic cellular FLICE-like inhibitory protein cFLIP plays a pivotal role in normal tissues homoeostasis and the development of many tumors, but its role in normal thymus (NT), thymomas and thymic carcinomas (TC) is largely unknown. Expression, regulation and function of cFLIP were analyzed in biopsies of NT, thymomas, thymic squamous cell carcinomas (TSCC), thymic epithelial cells (TECs) derived thereof and in the TC line 1889c by qRT-PCR, western blot, shRNA techniques, and functional assays addressing survival, senescence and autophagy. More than 90% of thymomas and TSCCs showed increased cFLIP expression compared to NT. cFLIP expression declined with age in NTs but not in thymomas. During short term culture cFLIP expression levels declined significantly slower in neoplastic than non-neoplastic primary TECs. Down-regulation of cFLIP by shRNA or NF-κB inhibition accelerated senescence and induced autophagy and cell death in neoplastic TECs. The results suggest a role of cFLIP in the involution of normal thymus and the development of thymomas and TSCC. Since increased expression of cFLIP is a known tumor escape mechanism, it may serve as tissue-based biomarker in future clinical trials, including immune checkpoint inhibitor trials in the commonly PD-L1 high thymomas and TCs.

Using ToxCast data to reconstruct dynamic cell state ...

EPA Pesticide Factsheets

AbstractBackground. High-throughput in vitro screening is an important tool for evaluating the potential biological activity of the thousands of existing chemicals in commerce and the hundreds more introduced each year. Among the assay technologies available, high-content imaging (HCI) allows multiplexed measurements of cellular phenotypic changes induced by chemical exposures. For a large chemical inventory having limited concentration-time series data, the deconvolution of cellular response profiles into transitive or irrevocable state trajectories is an important consideration. Objectives. Our goal was to analyze temporal and concentration-related cellular changes measured using HCI to identify the “tipping point” at which the cells did not show recovery towards a normal phenotypic state. Methods. The effects of 976 chemicals (ToxCast Phase I and II) were evaluated using HCI as a function of concentration and time in HepG2 cells over a 72-hr exposure period to concentrations ranging from 0.4- to 200 µM. The cellular endpoints included nuclear p53 accumulation, JNK, markers of oxidative stress, cytoskeletal changes, mitochondrial energization and density, cell viability and cell cycle progression. A novel computational model was developed to interpret dynamic multidimensional system responses as cell-state trajectories. Results. Analysis of cell-state trajectories showed that HepG2 cells were resilient to the effects of 178 chemicals up to the highest co

Chlorella protects against hydrogen peroxide-induced pancreatic β-cell damage.

PubMed

Lin, Chia-Yu; Huang, Pei-Jane; Chao, Che-Yi

2014-12-01

Oxidative stress has been implicated in the etiology of pancreatic β-cell dysfunction and diabetes. Studies have shown that chlorella could be important in health promotion or disease prevention through its antioxidant capacity. However, whether chlorella has a cytoprotective effect in pancreatic β-cells remains to be elucidated. We investigated the protective effects of chlorella on H2O2-induced oxidative damage in INS-1 (832/13) cells. Chlorella partially restored cell viability after H2O2 toxicity. To further investigate the effects of chlorella on mitochondria function and cellular oxidative stress, we analyzed mitochondria membrane potential, ATP concentrations, and cellular levels of reactive oxygen species (ROS). Chlorella prevented mitochondria disruption and maintained cellular ATP levels after H2O2 toxicity. It also normalized intracellular levels of ROS to that of control in the presence of H2O2. Chlorella protected cells from apoptosis as indicated by less p-Histone and caspase 3 activation. In addition, chlorella not only enhanced glucose-stimulated insulin secretion (GSIS), but also partially restored the reduced GSIS after H2O2 toxicity. Our results suggest that chlorella is effective in amelioration of cellular oxidative stress and destruction, and therefore protects INS-1 (832/13) cells from H2O2-induced apoptosis and increases insulin secretion. Chlorella should be studied for use in the prevention or treatment of diabetes.

Cellular retinoic acid bioavailability in various pathologies and its therapeutic implication.

PubMed

Osanai, Makoto

2017-06-01

Retinoic acid (RA), an active metabolite of vitamin A, is a critical signaling molecule in various cell types. We found that RA depletion caused by expression of the RA-metabolizing enzyme CYP26A1 promotes carcinogenesis, implicating CYP26A1 as a candidate oncogene. Several studies of CYP26s have suggested that the biological effect of RA on target cells is primarily determined by "cellular RA bioavailability", which is defined as the RA level in an individual cell, rather than by the serum concentration of RA. Consistently, stellate cells store approximately 80% of vitamin A in the body, and the state of cellular RA bioavailability regulates their function. Based on the similarities between stellate cells and astrocytes, we demonstrated that retinal astrocytes regulate tight junction-based endothelial integrity in a paracrine manner. Since diabetic retinopathy is characterized by increased vascular permeability in its early pathogenesis, RA normalized retinal astrocytes that are compromised in diabetes, resulting in suppression of vascular leakiness. RA also attenuated the loss of the epithelial barrier in murine experimental colitis. The concept of "cellular RA bioavailability" in various diseases will be directed at understanding various pathologies caused by RA insufficiency, implying the potential feasibility of a therapeutic strategy targeting the stellate cell system. © 2017 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.

Characteristics of Middle School Students Learning Actions in Outdoor Mathematical Activities with the Cellular Phone

ERIC Educational Resources Information Center

Daher, Wajeeh; Baya'a, Nimer

2012-01-01

Learning in the cellular phone environment enables utilizing the multiple functions of the cellular phone, such as mobility, availability, interactivity, verbal and voice communication, taking pictures or recording audio and video, measuring time and transferring information. These functions together with mathematics-designated cellular phone…

The Landscape of mtDNA Modifications in Cancer: A Tale of Two Cities.

PubMed

Hertweck, Kate L; Dasgupta, Santanu

2017-01-01

Mitochondria from normal and cancerous cells represent a tale of two cities, wherein both execute similar processes but with different cellular and molecular effects. Given the number of reviews currently available which describe the functional implications of mitochondrial mutations in cancer, this article focuses on documenting current knowledge in the abundance and distribution of somatic mitochondrial mutations, followed by elucidation of processes which affect the fate of mutations in cancer cells. The conclusion includes an overview of translational implications for mtDNA mutations, as well as recommendations for future research uniting mitochondrial variants and tumorigenesis.

[The prevention and treatment of suppurative-inflammatory complications in the bronchopulmonary system during prolonged artificial ventilation].

PubMed

Mozhaev, G A; Tikhonovskiĭ, I Iu

1992-01-01

The use of physical methods, namely low frequency magnetic field in critically ill patients under respiratory therapy made it possible to prevent and in case of their development to effectively treat pyoinflammatory bronchopulmonary complications that accompany prolonged controlled lung ventilation. The results obtained were due to the elimination of an unfavourable effect of controlled lung ventilation on natural resistance and immune response of the respiratory tract because of normalization of physicochemical properties of the tracheobronchial tree secretion, enhanced functional capacities of phagocytes, repaired bonds between cellular and humoral local immunity in the lungs.

Modeling integrated cellular machinery using hybrid Petri-Boolean networks.

PubMed

Berestovsky, Natalie; Zhou, Wanding; Nagrath, Deepak; Nakhleh, Luay

2013-01-01

The behavior and phenotypic changes of cells are governed by a cellular circuitry that represents a set of biochemical reactions. Based on biological functions, this circuitry is divided into three types of networks, each encoding for a major biological process: signal transduction, transcription regulation, and metabolism. This division has generally enabled taming computational complexity dealing with the entire system, allowed for using modeling techniques that are specific to each of the components, and achieved separation of the different time scales at which reactions in each of the three networks occur. Nonetheless, with this division comes loss of information and power needed to elucidate certain cellular phenomena. Within the cell, these three types of networks work in tandem, and each produces signals and/or substances that are used by the others to process information and operate normally. Therefore, computational techniques for modeling integrated cellular machinery are needed. In this work, we propose an integrated hybrid model (IHM) that combines Petri nets and Boolean networks to model integrated cellular networks. Coupled with a stochastic simulation mechanism, the model simulates the dynamics of the integrated network, and can be perturbed to generate testable hypotheses. Our model is qualitative and is mostly built upon knowledge from the literature and requires fine-tuning of very few parameters. We validated our model on two systems: the transcriptional regulation of glucose metabolism in human cells, and cellular osmoregulation in S. cerevisiae. The model produced results that are in very good agreement with experimental data, and produces valid hypotheses. The abstract nature of our model and the ease of its construction makes it a very good candidate for modeling integrated networks from qualitative data. The results it produces can guide the practitioner to zoom into components and interconnections and investigate them using such more detailed mathematical models.

Modeling Integrated Cellular Machinery Using Hybrid Petri-Boolean Networks

PubMed Central

Berestovsky, Natalie; Zhou, Wanding; Nagrath, Deepak; Nakhleh, Luay

2013-01-01

The behavior and phenotypic changes of cells are governed by a cellular circuitry that represents a set of biochemical reactions. Based on biological functions, this circuitry is divided into three types of networks, each encoding for a major biological process: signal transduction, transcription regulation, and metabolism. This division has generally enabled taming computational complexity dealing with the entire system, allowed for using modeling techniques that are specific to each of the components, and achieved separation of the different time scales at which reactions in each of the three networks occur. Nonetheless, with this division comes loss of information and power needed to elucidate certain cellular phenomena. Within the cell, these three types of networks work in tandem, and each produces signals and/or substances that are used by the others to process information and operate normally. Therefore, computational techniques for modeling integrated cellular machinery are needed. In this work, we propose an integrated hybrid model (IHM) that combines Petri nets and Boolean networks to model integrated cellular networks. Coupled with a stochastic simulation mechanism, the model simulates the dynamics of the integrated network, and can be perturbed to generate testable hypotheses. Our model is qualitative and is mostly built upon knowledge from the literature and requires fine-tuning of very few parameters. We validated our model on two systems: the transcriptional regulation of glucose metabolism in human cells, and cellular osmoregulation in S. cerevisiae. The model produced results that are in very good agreement with experimental data, and produces valid hypotheses. The abstract nature of our model and the ease of its construction makes it a very good candidate for modeling integrated networks from qualitative data. The results it produces can guide the practitioner to zoom into components and interconnections and investigate them using such more detailed mathematical models. PMID:24244124

SOD1 Overexpression Preserves Baroreflex Control of Heart Rate with an Increase of Aortic Depressor Nerve Function

PubMed Central

Hatcher, Jeffrey; Gu, He; Cheng, Zixi (Jack)

2016-01-01

Overproduction of reactive oxygen species (ROS), such as the superoxide radical (O2 ∙−), is associated with diseases which compromise cardiac autonomic function. Overexpression of SOD1 may offer protection against ROS damage to the cardiac autonomic nervous system, but reductions of O2 ∙− may interfere with normal cellular functions. We have selected the C57B6SJL-Tg (SOD1)2 Gur/J mouse as a model to determine whether SOD1 overexpression alters cardiac autonomic function, as measured by baroreflex sensitivity (BRS) and aortic depressor nerve (ADN) recordings, as well as evaluation of baseline heart rate (HR) and mean arterial pressure (MAP). Under isoflurane anesthesia, C57 wild-type and SOD1 mice were catheterized with an arterial pressure transducer and measurements of HR and MAP were taken. After establishing a baseline, hypotension and hypertension were induced by injection of sodium nitroprusside (SNP) and phenylephrine (PE), respectively, and ΔHR versus ΔMAP were recorded as a measure of baroreflex sensitivity (BRS). SNP and PE treatment were administered sequentially after a recovery period to measure arterial baroreceptor activation by recording aortic depressor nerve activity. Our findings show that overexpression of SOD1 in C57B6SJL-Tg (SOD1)2 Gur/J mouse preserved the normal HR, MAP, and BRS but enhanced aortic depressor nerve function. PMID:26823951

Interrogation of Mammalian Protein Complex Structure, Function, and Membership Using Genome-Scale Fitness Screens.

PubMed

Pan, Joshua; Meyers, Robin M; Michel, Brittany C; Mashtalir, Nazar; Sizemore, Ann E; Wells, Jonathan N; Cassel, Seth H; Vazquez, Francisca; Weir, Barbara A; Hahn, William C; Marsh, Joseph A; Tsherniak, Aviad; Kadoch, Cigall

2018-05-23

Protein complexes are assemblies of subunits that have co-evolved to execute one or many coordinated functions in the cellular environment. Functional annotation of mammalian protein complexes is critical to understanding biological processes, as well as disease mechanisms. Here, we used genetic co-essentiality derived from genome-scale RNAi- and CRISPR-Cas9-based fitness screens performed across hundreds of human cancer cell lines to assign measures of functional similarity. From these measures, we systematically built and characterized functional similarity networks that recapitulate known structural and functional features of well-studied protein complexes and resolve novel functional modules within complexes lacking structural resolution, such as the mammalian SWI/SNF complex. Finally, by integrating functional networks with large protein-protein interaction networks, we discovered novel protein complexes involving recently evolved genes of unknown function. Taken together, these findings demonstrate the utility of genetic perturbation screens alone, and in combination with large-scale biophysical data, to enhance our understanding of mammalian protein complexes in normal and disease states. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Functional activation of PPARγ in human upper aerodigestive cancer cell lines.

PubMed

Wright, Simon K; Wuertz, Beverly R; Harris, George; Abu Ghazallah, Raed; Miller, Wendy A; Gaffney, Patrick M; Ondrey, Frank G

2017-01-01

Upper aerodigestive cancer is an aggressive malignancy with relatively stagnant long-term survival rates over 20 yr. Recent studies have demonstrated that exploitation of PPARγ pathways may be a novel therapy for cancer and its prevention. We tested whether PPARγ is expressed and inducible in aerodigestive carcinoma cells and whether it is present in human upper aerodigestive tumors. Human oral cancer CA-9-22 and NA cell lines were treated with the PPAR activators eicosatetraynoic acid (ETYA), 15-deoxy-δ- 12,14-prostaglandin J2 (PG-J2), and the thiazolidinedione, ciglitazone, and evaluated for their ability to functionally activate PPARγ luciferase reporter gene constructs. Cellular proliferation and clonogenic potential after PPARγ ligand treatment were also evaluated. Aerodigestive cancer specimens and normal tissues were evaluated for PPARγ expression on gene expression profiling and immunoblotting. Functional activation of PPARγ reporter gene constructs and increases in PPARγ protein were confirmed in the nuclear compartment after PPARγ ligand treatment. Significant decreases in cell proliferation and clonogenic potential resulted from treatment. Lipid accumulation was induced by PPARγ activator treatment. 75% of tumor specimens and 100% of normal control tissues expressed PPARγ RNA, and PPARγ protein was confirmed in 66% of tumor specimens analyzed by immunoblotting. We conclude PPARγ can be functionally activated in upper aerodigestive cancer and that its activation downregulates several features of the neoplastic phenotype. PPARγ expression in human upper aerodigestive tract tumors and normal cells potentially legitimizes it as a novel intervention target in this disease. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Expression of cellular prion protein in the frontal and occipital lobe in Alzheimer's disease, diffuse Lewy body disease, and in normal brain: an immunohistochemical study.

PubMed

Rezaie, Payam; Pontikis, Charlie C; Hudson, Lance; Cairns, Nigel J; Lantos, Peter L

2005-08-01

Cellular prion protein (PrP(c)) is a glycoprotein expressed at low to moderate levels within the nervous system. Recent studies suggest that PrP(c) may possess neuroprotective functions and that its expression is upregulated in certain neurodegenerative disorders. We investigated whether PrP(c) expression is altered in the frontal and occipital cortex in two well-characterized neurodegenerative disorders--Alzheimer's disease (AD) and diffuse Lewy body disease (DLBD)--compared with that in normal human brain using immunohistochemistry and computerized image analysis. The distribution of PrP(c) was further tested for correlation with glial reactivity. We found that PrP(c) was localized mainly in the gray matter (predominantly in neurons) and expressed at higher levels within the occipital cortex in the normal human brain. Image analysis revealed no significant variability in PrP(c) expression between DLBD and control cases. However, blood vessels within the white matter of DLBD cases showed immunoreactivity to PrP(c). By contrast, this protein was differentially expressed in the frontal and occipital cortex of AD cases; it was markedly overexpressed in the former and significantly reduced in the latter. Epitope specificity of antibodies appeared important when detecting PrP(c). The distribution of PrP(c) did not correlate with glial immunoreactivity. In conclusion, this study supports the proposal that regional changes in expression of PrP(c) may occur in certain neurodegenerative disorders such as AD, but not in other disorders such as DLBD.

Cell Signaling and Neurotoxicity: 3H-Arachidonic acid release (Phospholipase A2) in cerebellar granule neurons

EPA Science Inventory

Cell signaling is a complex process which controls basic cellular activities and coordinates actions to maintain normal cellular homeostasis. Alterations in signaling processes have been associated with neurological diseases such as Alzheimer's and cerebellar ataxia, as well as, ...

Bmi-1: At the crossroads of physiological and pathological biology

PubMed Central

Bhattacharya, Resham; Mustafi, Soumyajit Banerjee; Street, Mark; Dey, Anindya; Dwivedi, Shailendra Kumar Dhar

2015-01-01

Bmi-1 is a member of the Polycomb Repressor Complex1 that mediates gene silencing by regulating chromatin structure and is indispensable for self-renewal of both normal and cancer stem cells. Despite three decades of research that have elucidated the transcriptional regulation, post-translational modifications and functions of Bmi-1 in regulating the DNA damage response, cellular bioenergetics, and pathologies, the entire potential of a protein with such varied function remains to be realized. This review attempts to synthesize the current knowledge on Bmi-1 with an emphasis on its role in both normal physiology and cancer. Additionally, since cancer stem cells are emerging as a new paradigm for therapy resistance, the role of Bmi-1 in this perspective is also highlighted. The wide spectrum of malignancies that implicate Bmi-1 as a signature for stemness and oncogenesis also make it a suitable candidate for therapy. Nonetheless new approaches are vitally needed to further characterize physiological roles of Bmi-1 with the long-term goal of using Bmi-1 as a prognostic marker and a therapeutic target. PMID:26448339

What We Should Know Before Using Tissue Engineering Techniques to Repair Injured Tendons: A Developmental Biology Perspective

PubMed Central

Liu, Chia-Feng; Aschbacher-Smith, Lindsey; Barthelery, Nicolas J.; Dyment, Nathaniel; Butler, David

2011-01-01

Tendons connect muscles to bones, and serve as the transmitters of force that allow all the movements of the body. Tenocytes are the basic cellular units of tendons, and produce the collagens that form the hierarchical fiber system of the tendon. Tendon injuries are common, and difficult to repair, particularly in the case of the insertion of tendon into bone. Successful attempts at cell-based repair therapies will require an understanding of the normal development of tendon tissues, including their differentiated regions such as the fibrous mid-section and fibrocartilaginous insertion site. Many genes are known to be involved in the formation of tendon. However, their functional roles in tendon development have not been fully characterized. Tissue engineers have attempted to generate functional tendon tissue in vitro. However, a lack of knowledge of normal tendon development has hampered these efforts. Here we review studies focusing on the developmental mechanisms of tendon development, and discuss the potential applications of a molecular understanding of tendon development to the treatment of tendon injuries. PMID:21314435

NF-κB Signaling Pathway and its Potential as a Target for Therapy in Lymphoid Neoplasms

PubMed Central

Yu, Li; Li, Ling; Medeiros, L. Jeffrey; Young, Ken H.

2016-01-01

The NF-κB pathway, a critical regulator of apoptosis, plays a key role in many normal cellular functions. Genetic alterations and other mechanisms leading to constitutive activation of the NF-κB pathway contribute to cancer development, progression and therapy resistance by activation of downstream anti-apoptotic pathways, unfavorable microenvironment interactions, and gene dysregulation. Not surprisingly, given its importance to normal and cancer cell function, the NF-κB pathway has emerged as a target for therapy. In the review, we present the physiologic role of the NF-κB pathway and recent advances in better understanding of the pathologic roles of the NF-κB pathway in major types of lymphoid neoplasms. We also provide an update of clinical trials that use NF-κB pathway inhibitors. These trials are exploring the clinical efficiency of combining NF-κB pathway inhibitors with various agents that target diverse mechanisms of action with the goal being to optimize novel therapeutic opportunities for targeting oncogenic pathways to eradicate cancer cells. PMID:27773462

Autophagy: a new player in skeletal maintenance?

PubMed

Hocking, Lynne J; Whitehouse, Caroline; Helfrich, Miep H

2012-07-01

Imbalances between bone resorption and formation lie at the root of disorders such as osteoporosis, Paget's disease of bone (PDB), and osteopetrosis. Recently, genetic and functional studies have implicated proteins involved in autophagic protein degradation as important mediators of bone cell function in normal physiology and in pathology. Autophagy is the conserved process whereby aggregated proteins, intracellular pathogens, and damaged organelles are degraded and recycled. This process is important both for normal cellular quality control and in response to environmental or internal stressors, particularly in terminally-differentiated cells. Autophagic structures can also act as hubs for the spatial organization of recycling and synthetic process in secretory cells. Alterations to autophagy (reduction, hyperactivation, or impairment) are associated with a number of disorders, including neurodegenerative diseases and cancers, and are now being implicated in maintenance of skeletal homoeostasis. Here, we introduce the topic of autophagy, describe the new findings that are starting to emerge from the bone field, and consider the therapeutic potential of modifying this pathway for the treatment of age-related bone disorders. Copyright © 2012 American Society for Bone and Mineral Research.

Breast cancer mitosis detection in histopathological images with spatial feature extraction

NASA Astrophysics Data System (ADS)

Albayrak, Abdülkadir; Bilgin, Gökhan

2013-12-01

In this work, cellular mitosis detection in histopathological images has been investigated. Mitosis detection is very expensive and time consuming process. Development of digital imaging in pathology has enabled reasonable and effective solution to this problem. Segmentation of digital images provides easier analysis of cell structures in histopathological data. To differentiate normal and mitotic cells in histopathological images, feature extraction step is very crucial step for the system accuracy. A mitotic cell has more distinctive textural dissimilarities than the other normal cells. Hence, it is important to incorporate spatial information in feature extraction or in post-processing steps. As a main part of this study, Haralick texture descriptor has been proposed with different spatial window sizes in RGB and La*b* color spaces. So, spatial dependencies of normal and mitotic cellular pixels can be evaluated within different pixel neighborhoods. Extracted features are compared with various sample sizes by Support Vector Machines using k-fold cross validation method. According to the represented results, it has been shown that separation accuracy on mitotic and non-mitotic cellular pixels gets better with the increasing size of spatial window.

A gene involved in control of human cellular senescence on human chromosome 1q

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

Hensler, P.J.; Pereira-Smith, O.M.; Annab, L.A.

1994-04-01

Normal cells in culture exhibit limited division potential and have been used as a model for cellular senescence. In contrast, tumor-derived or carcinogen- or virus-transformed cells are capable of indefinite division. Fusion of normal human diploid fibroblasts with immortal human cells yielded hybrids having limited life spans, indicating that cellular senescence was dominant. Fusions of various immortal human cell lines with each other led to the identification of four complementation groups for indefinite division. The purpose of this study was to determine whether human chromosome 1 could complement the recessive immortal defect of human cell lines assigned to one ofmore » the four complementation groups. Using microcell fusion, the authors introduced a single normal human chromosome 1 into immortal human cell lines representing the complementation groups and determined that it caused loss of proliferative potential of an osteosarcoma-derived cell line (TE85), a cytomegalovirus-transformed lung fibroblast cell line (CMV-Mj-HEL-1), and a Ki-ras[sup +]-transformed derivative of TE85 (143B TK[sup [minus

Metabolic activation and nucleic acid binding of acetaminophen and related arylamine substrates by the respiratory burst of human granulocytes.

PubMed

Corbett, M D; Corbett, B R; Hannothiaux, M H; Quintana, S J

1989-01-01

Following stimulation with phorbol myristate acetate, human granulocytes were found to incorporate acetaminophen, p-phenetidine, p-aminophenol, and p-chloroaniline into cellular DNA and RNA. Phenacetin was not incorporated into nucleic acid or metabolized by such activated granulocytes. None of the substrates gave nucleic acid binding if the granulocyte cultures were not induced to undergo the respiratory burst. Additional studies on the binding of acetaminophen to DNA and RNA were made by use of both ring-14C-labeled and carbonyl-14C-labeled forms of this substrate. The finding that equivalent amounts of these two labeled acetaminophen substrates were bound to cellular DNA demonstrated that the intact acetaminophen molecule was incorporated into DNA. On the other hand, the finding that excess ring-14C-labeled acetaminophen was incorporated into cellular RNA implies partial hydrolysis of the acetaminophen substrate prior to RNA binding. Evidence was presented which strongly indicates that the nucleic acid binding of the substrates was covalent in nature. The inability of the respiratory burst to result in the binding of phenacetin to nucleic acid suggests that arylamides are not normally activated or metabolized by activated granulocytes. Acetaminophen is an exception to the recalcitrance of arylamides to such bioactivation processes because it also possesses the phenolic functional group, which, like the arylamine group, is oxidized by certain reactive oxygen species. Myeloperoxidase appears to be much more important in the binding of acetaminophen to DNA than it is in the DNA binding of arylamines in general. The role of the respiratory burst in causing the bioactivation of certain arylamines, which are not normally genotoxic via the more usual microsomal activation pathways, was extended to include certain amide substrates such as acetaminophen.

The effect of glucagon-like peptide-1 in the management of diabetes mellitus: cellular and molecular mechanisms.

PubMed

Lotfy, Mohamed; Singh, Jaipaul; Rashed, Hameed; Tariq, Saeed; Zilahi, Erika; Adeghate, Ernest

2014-11-01

Incretins, such as glucagon-like peptide-1 (GLP)-1, have been shown to elevate plasma insulin concentration. The purpose of this study is to investigate the cellular and molecular basis of the beneficial effects of GLP-1. Normal and diabetic male Wistar rats were treated with GLP-1 (50 ng/kg body weight) for 10 weeks. At the end of the experiment, pancreatic tissues were taken for immunohistochemistry, immunoelectron microscopy and real-time polymerase chain reaction studies. Samples of blood were retrieved from the animals for the measurement of enzymes and insulin. The results show that treatment of diabetic rats with GLP-1 caused significant (P < 0.05) reduction in body weight gain and blood glucose level. GLP-1 (10(-12)-10(-6) M) induced significant (P < 0.01) dose-dependent increases in insulin release from the pancreas of normal and diabetic rats compared to basal. Diabetes-induced abnormal liver (aspartate aminotransferase and alanine aminotransferase) and kidney (blood urea nitrogen and uric acid) parameters were corrected in GLP-1-treated rats compared to controls. GLP-1 treatment induced significant (P < 0.05) elevation in the expression of pancreatic duodenal homeobox-1, heat shock protein-70, glutathione peroxidase, insulin receptor and GLP-1-receptor genes in diabetic animals compared to controls. GLP-1 is present in pancreatic beta cells and significantly (P < 0.05) increased the number of insulin-, glutathione reductase- and catalase-immunoreactive islet cells. The results of this study show that GLP-1 is co-localized with insulin and seems to exert its beneficial effects by increasing cellular concentrations of endogenous antioxidant genes and other genes involved in the maintenance of pancreatic beta cell structure and function.

Notch3 protein expression in skin fibroblasts from CADASIL patients.

PubMed

Qualtieri, Antonio; Ungaro, Carmine; Bagalà, Angelo; Bianchi, Silvia; Pantoni, Leonardo; Moccia, Marcello; Mazzei, Rosalucia

2018-07-15

CADASIL is an inherited cerebrovascular disease caused by mutations in the NOTCH3 gene. Notch signaling is involved in a broad spectrum of function, from the cell proliferation to apoptosis. Thus far, because the molecular mechanism underlying the pathological alterations remains unclear and taking into account that fibroblasts contribute to the integrity of the vasculature, our aims was to establish whether fibroblasts, in subjects carrying different NOTCH3 mutations, show abnormalities in the protein expression. We performed the investigation on skin fibroblasts in culture obtained from three CADASIL patients and normal subjects. The patients were genetically characterized, and carried a p.R61W, a p.C174T, and p.R103X, mutation respectively. Notch3 expression was first evaluated on fibroblasts by immunofluorescence analysis, then western blot on cellular extract was utilized to validate the immunofluorescence results. The Notch3 immunoreactivity was clearly detected along the cellular body and in the cellular nuclei of the control fibroblasts. We observed a marked, statistically significant, reduction of the fluorescence immunoreactivity in the fibroblasts from patient with the classical C174T cysteine mutation and a less pronounced reduction in the other two subject's samples with respect to the normal controls. These data were confirmed by the immunoblot analysis. Our results show that the investigated three NOTCH3 mutations are associated with a reduction of the levels of Notch3 expression in vitro. Because the smooth muscle cells appear to be predominantly involved in this cerebrovascular disease, our result, despite the limitation of the sample size examinated, clearly suggest that also fibroblasts, directly involved in making the vascular basal lamina and in maintaining the vascular integrity, may play an important role in the mechanism responsible for the disease. Copyright © 2018 Elsevier B.V. All rights reserved.

Heat stress-induced nuclear transport mediated by Hikeshi confers nuclear function of Hsp70s.

PubMed

Imamoto, Naoko

2018-06-01

The prime feature of eukaryotic cells is the separation of the intracellular space into two compartments, the nucleus and the cytoplasm. Active nuclear transport is crucial for the maintenance of this separation. In this report, we focus on a nuclear transport receptor named Hikeshi, which mediates the heat stress-induced nuclear import of 70-kDa heat shock proteins (Hsp70s), and discuss how the same protein can function differently depending on the cellular compartment in which it is localized. Hsp70 is a molecular chaperone that is predominantly localized in the cytoplasm under normal conditions but is known to accumulate in the nucleus under conditions of heat stress. Although the reported function of Hsp70 is mostly attributed to its molecular function in the cytoplasm, the functions of Hsp70 may extend beyond molecular chaperone activity in the nucleus. Copyright © 2018 The Author. Published by Elsevier Ltd.. All rights reserved.

Biochemical and Cellular Assessment of Acetabular Chondral Flaps Identified During Hip Arthroscopy.

PubMed

Hariri, Sanaz; Truntzer, Jeremy; Smith, Robert Lane; Safran, Marc R

2015-06-01

To analyze chondral flaps debrided during hip arthroscopy to determine their biochemical and cellular composition. Thirty-one full-thickness acetabular chondral flaps were collected during hip arthroscopy. Biochemical analysis was undertaken in 21 flaps from 20 patients, and cellular viability was determined in 10 flaps from 10 patients. Biochemical analysis included concentrations of (1) DNA (an indicator of chondrocyte content), (2) hydroxyproline (an indicator of collagen content), and (3) glycosaminoglycan (an indicator of chondrocyte biosynthesis). Higher values for these parameters indicated more healthy tissue. The flaps were examined to determine the percentage of viable chondrocytes. The percentage of acetabular chondral flap specimens that had concentrations within 1 SD of the mean values reported in previous normal cartilage studies was 38% for DNA, 0% for glycosaminoglycan, and 43% for hydroxyproline. The average cellular viability of our acetabular chondral flap specimens was 39% (SD, 14%). Only 2 of the 10 specimens had more than half the cells still viable. There was no correlation between (1) the gross examination of the joint or knowledge of the patient's demographic characteristics and symptoms and (2) biochemical properties and cell viability of the flap, with one exception: a degenerative appearance of the surrounding cartilage correlated with a higher hydroxyproline concentration. Although full-thickness acetabular chondral flaps can appear normal grossly, the biochemical properties and percentage of live chondrocytes in full-thickness chondral flaps encountered in hip arthroscopy show that this tissue is not normal. There has been recent interest in repairing chondral flaps encountered during hip arthroscopy. These data suggest that acetabular chondral flaps are not biochemically and cellularly normal. Although these flaps may still be valuable mechanically and/or as a scaffold in some conductive or inductive capacity, further study is required to assess the clinical benefit of repair. Copyright © 2015 Arthroscopy Association of North America. Published by Elsevier Inc. All rights reserved.

Identification of a molecular recognition feature in the E1A oncoprotein that binds the SUMO conjugase UBC9 and likely interferes with polySUMOylation.

PubMed

Yousef, A F; Fonseca, G J; Pelka, P; Ablack, J N G; Walsh, C; Dick, F A; Bazett-Jones, D P; Shaw, G S; Mymryk, J S

2010-08-19

Hub proteins have central roles in regulating cellular processes. By targeting a single cellular hub, a viral oncogene may gain control over an entire module in the cellular interaction network that is potentially comprised of hundreds of proteins. The adenovirus E1A oncoprotein is a viral hub that interacts with many cellular hub proteins by short linear motifs/molecular recognition features (MoRFs). These interactions transform the architecture of the cellular protein interaction network and virtually reprogram the cell. To identify additional MoRFs within E1A, we screened portions of E1A for their ability to activate yeast pseudohyphal growth or differentiation. This identified a novel functional region within E1A conserved region 2 comprised of the sequence EVIDLT. This MoRF is necessary and sufficient to bind the N-terminal region of the SUMO conjugase UBC9, which also interacts with SUMO noncovalently and is involved in polySUMOylation. Our results suggest that E1A interferes with polySUMOylation, but not with monoSUMOylation. These data provide the first insight into the consequences of the interaction of E1A with UBC9, which was initially described in 1996. We further demonstrate that polySUMOylation regulates pseudohyphal growth and promyelocytic leukemia body reorganization by E1A. In conclusion, the interaction of the E1A oncogene with UBC9 mimics the normal binding between SUMO and UBC9 and represents a novel mechanism to modulate polySUMOylation.

Future Targets for Female Sexual Dysfunction.

PubMed

Farmer, Melissa; Yoon, Hana; Goldstein, Irwin

2016-08-01

Female sexual function reflects a dynamic interplay of central and peripheral nervous, vascular, and endocrine systems. The primary challenge in the development of novel treatments for female sexual dysfunction is the identification and targeted modulation of excitatory sexual circuits using pharmacologic treatments that facilitate the synthesis, release, and/or receptor binding of neurochemicals, peptides, and hormones that promote female sexual function. To develop an evidence-based state-of-the-art consensus report that critically integrates current knowledge of the therapeutic potential for known molecular and cellular targets to facilitate the physiologic processes underlying female sexual function. State-of-the-art review representing the opinions of international experts developed in a consensus process during a 1-year period. Expert opinion was established by grading the evidence-based medical literature, intensive internal committee discussion, public presentation, and debate. Scientific investigation is urgently needed to expand knowledge and foster development of future treatments that maintain genital tissue integrity, enhance genital physiologic responsiveness, and optimize positive subjective appraisal of internal and external sexual cues. This article critically condenses the current knowledge of therapeutic manipulation of molecular and cellular targets within biological systems responsible for female sexual physiologic function. Future treatment targets include pharmacologic modulation of emotional learning circuits, restoration of normal tactile sensation, growth factor therapy, gene therapy, stem cell-based therapies, and regenerative medicine. Concurrent use of centrally and peripherally acting therapies could optimize treatment response. Copyright © 2016 International Society for Sexual Medicine. Published by Elsevier Inc. All rights reserved.

MRF Family Genes Are Involved in Translation Control, Especially under Energy-Deficient Conditions, and Their Expression and Functions Are Modulated by the TOR Signaling Pathway[OPEN

PubMed Central

Lee, Du-Hwa; Park, Seung Jun; Ahn, Chang Sook

2017-01-01

Dynamic control of protein translation in response to the environment is essential for the survival of plant cells. Target of rapamycin (TOR) coordinates protein synthesis with cellular energy/nutrient availability through transcriptional modulation and phosphorylation of the translation machinery. However, mechanisms of TOR-mediated translation control are poorly understood in plants. Here, we report that Arabidopsis thaliana MRF (MA3 DOMAIN-CONTAINING TRANSLATION REGULATORY FACTOR) family genes encode translation regulatory factors under TOR control, and their functions are particularly important in energy-deficient conditions. Four MRF family genes (MRF1-MRF4) are transcriptionally induced by dark and starvation (DS). Silencing of multiple MRFs increases susceptibility to DS and treatment with a TOR inhibitor, while MRF1 overexpression decreases susceptibility. MRF proteins interact with eIF4A and cofractionate with ribosomes. MRF silencing decreases translation activity, while MRF1 overexpression increases it, accompanied by altered ribosome patterns, particularly in DS. Furthermore, MRF deficiency in DS causes altered distribution of mRNAs in sucrose gradient fractions and accelerates rRNA degradation. MRF1 is phosphorylated in vivo and phosphorylated by S6 kinases in vitro. MRF expression and MRF1 ribosome association and phosphorylation are modulated by cellular energy status and TOR activity. We discuss possible mechanisms of the function of MRF family proteins under normal and energy-deficient conditions and their functional link with the TOR pathway. PMID:29084871

Growth factors in the anterior segment: role in tissue maintenance, wound healing and ocular pathology.

PubMed

Klenkler, Bettina; Sheardown, Heather

2004-11-01

A number of growth factors and their associated receptors, including epidermal growth factor, transforming growth factor-beta, keratinocyte growth factor, hepatocyte growth factor, fibroblast growth factor and platelet-derived growth factor have been detected in the anterior segment of the eye. On binding to cellular receptors, these factors activate signalling cascades, which regulate functions including mitosis, differentiation, motility and apoptosis. Production of growth factors by corneal cells and their presence in the tear fluid and aqueous humour is essential for maintenance and renewal of normal tissue in the anterior eye and the prevention of undesirable immune or angiogenic reactions. Growth factors also play a vital role in corneal wound healing, mediating the proliferation of epithelial and stromal tissue and affecting the remodelling of the extracellular matrix (ECM). These functions depend on a complex interplay between growth factors of different types, the ECM, and regulatory mechanisms of the affected cells. Imbalances may lead to deficient wound healing and various ocular pathologies, including edema, neovascularization and glaucoma. Growth factors may be targeted in therapeutic ophthalmic applications, through exogenous application or selective inhibition, and may be used to elicit specific cellular responses to ophthalmic materials. A thorough understanding of the mechanism and function of growth factors and their actions in the complex environment of the anterior eye is required for these purposes. Growth factors, their function and mechanisms of action as well as the interplay between different growth factors based on recent in vitro and in vivo studies are presented.

Longitudinal assessment of retinal structure and function reveals a rod-cone degeneration in a guinea pig model initially presented as night blind.

PubMed

Racine, Julie; Joly, Sandrine; Lachapelle, Pierre

2011-08-01

We have previously reported a naturally occurring retinopathy in a population of guinea pigs, where the affected animals presented a defect of the rod-mediated vision. The purpose of this study was to investigate if the mutants were affected with a stationary or degenerative retinopathy and to identify the cellular origin of this unique disorder. Electroretinogram (ERG) [postnatal day 1 (P1) to P450], light (LM) and electron microscopy (EM) [P5, P150, P450], and immunohistochemistry [P30, P150, P450] were evaluated from normal and mutant animals. Irrespective of age, the scotopic ERGs of mutants could only be evoked by bright flashes, and the resulting ERGs were of photopic waveform. Interestingly, the amplitude of the cone and the rod/cone a-waves was always of smaller amplitude in mutants, but this difference tended to decrease with age. In contrast, the b-waves were of larger amplitude than normal in photopic ERGs obtained prior to age 25 (days) and prior to age 10 for rod/cone ERGs. LM revealed, in mutants, an absence of the outer segment layer (OSL) with a reduction in the outer nuclear layer (ONL) thickness. EM disclosed the presence of cone outer segment (OS) while no rod OS could be evidenced. Immunohistochemistry revealed the presence of rhodopsin, both cone opsins as well as normal synaptophysin immunoreactivity. Finally, neither the retinal structure nor the function in the mutants achieved normal development. Results suggest that mutant animals are suffering from a degenerative retinal disorder that affects the structure and function of rods and cones.

Chronic kidney disease alters lipid trafficking and inflammatory responses in macrophages: effects of liver X receptor agonism.

PubMed

Kaseda, Ryohei; Tsuchida, Yohei; Yang, Hai-Chun; Yancey, Patricia G; Zhong, Jianyong; Tao, Huan; Bian, Aihua; Fogo, Agnes B; Linton, Mac Rae F; Fazio, Sergio; Ikizler, Talat Alp; Kon, Valentina

2018-01-27

Our aim was to evaluate lipid trafficking and inflammatory response of macrophages exposed to lipoproteins from subjects with moderate to severe chronic kidney disease (CKD), and to investigate the potential benefits of activating cellular cholesterol transporters via liver X receptor (LXR) agonism. LDL and HDL were isolated by sequential density gradient ultracentrifugation of plasma from patients with stage 3-4 CKD and individuals without kidney disease (HDL CKD and HDL Cont , respectively). Uptake of LDL, cholesterol efflux to HDL, and cellular inflammatory responses were assessed in human THP-1 cells. HDL effects on inflammatory markers (MCP-1, TNF-α, IL-1β), Toll-like receptors-2 (TLR-2) and - 4 (TLR-4), ATP-binding cassette class A transporter (ABCA1), NF-κB, extracellular signal regulated protein kinases 1/2 (ERK1/2) were assessed by RT-PCR and western blot before and after in vitro treatment with an LXR agonist. There was no difference in macrophage uptake of LDL isolated from CKD versus controls. By contrast, HD CKD was significantly less effective than HDL Cont in accepting cholesterol from cholesterol-enriched macrophages (median 20.8% [IQR 16.1-23.7] vs control (26.5% [IQR 19.6-28.5]; p = 0.008). LXR agonist upregulated ABCA1 expression and increased cholesterol efflux to HDL of both normal and CKD subjects, although the latter continued to show lower efflux capacity. HDL CKD increased macrophage cytokine response (TNF-α, MCP-1, IL-1β, and NF-κB) versus HDL Cont . The heightened cytokine response to HDL CKD was further amplified in cells treated with LXR agonist. The LXR-augmentation of inflammation was associated with increased TLR-2 and TLR-4 and ERK1/2. Moderate to severe impairment in kidney function promotes foam cell formation that reflects impairment in cholesterol acceptor function of HDL CKD . Activation of cellular cholesterol transporters by LXR agonism improves but does not normalize efflux to HDL CKD . However, LXR agonism actually increases the pro-inflammatory effects of HDL CKD through activation of TLRs and ERK1/2 pathways.

Is the urothelium intelligent?

PubMed

Birder, L A; Kanai, A J; Cruz, F; Moore, K; Fry, C H

2010-04-01

The urothelium separates the urinary tract lumen from underlying tissues of the tract wall. Previously considered as merely an effective barrier between these two compartments it is now recognized as a more active tissue that senses and transduces information about physical and chemical conditions within the urinary tract, such as luminal pressure, urine composition, etc. To understand this sensory function it is useful to consider the urothelium and suburothelium as a functional unit; containing uroepithelial cells, afferent and efferent nerve fibers and suburothelial interstitial cells. This structure responds to alterations in its external environment through the release of diffusible agents, such as ATP and acetylcholine, and eventually modulates the activity of afferent nerves and underlying smooth muscles. This review considers different stresses the urothelium/suburothelium responds to; the particular chemicals released; the cellular receptors that are consequently affected; and how nerve and muscle function is modulated. Brief consideration is also to regional differences in the urothelium/suburothelium along the urinary tract. The importance of different pathways in relaying sensory information in the normal urinary tract, or whether they are significant only in pathological conditions is also discussed. An operational definition of intelligence is used, whereby a system (urothelium/suburothelium) responds to external changes, to maximize the possibility of the urinary tract achieving its normal function. If so, the urothelium can be regarded as intelligent. The advantage of this approach is that input-output functions can be mathematically formulated, and the importance of different components contributing to abnormal urinary tract function can be calculated. (c) 2010 Wiley-Liss, Inc.

Whole Exome Sequencing and Heterologous Cellular Electrophysiology Studies Elucidate a Novel Loss-of-Function Mutation in the CACNA1A-Encoded Neuronal P/Q-Type Calcium Channel in a Child With Congenital Hypotonia and Developmental Delay.

PubMed

Weyhrauch, Derek L; Ye, Dan; Boczek, Nicole J; Tester, David J; Gavrilova, Ralitza H; Patterson, Marc C; Wieben, Eric D; Ackerman, Michael J

2016-02-01

A 4-year-old boy born at 37 weeks' gestation with intrauterine growth retardation presented with developmental delay with pronounced language and gross motor delay, axial hypotonia, and dynamic hypertonia of the extremities. Investigations including the Minnesota Newborn Screen, thyroid stimulating hormone/thyroxin, and inborn errors of metabolism screening were negative. Cerebral magnetic resonance imaging and spectroscopy were normal. Genetic testing was negative for coagulopathy, Smith-Lemli-Opitz, fragile X, and Prader-Willi/Angelman syndromes. Whole genome array analysis was unremarkable. Whole exome sequencing was performed through a commercial testing laboratory to elucidate the underlying etiology for the child's presentation. A de novo mutation was hypothesized. In attempt to establish pathogenicity of our candidate variant, cellular electrophysiologic functional analysis of the putative de novo mutation was performed using patch-clamp technology. Whole exome sequencing revealed a p.P1353L variant in the CACNA1A gene, which encodes for the α1-subunit of the brain-specific P/Q-type calcium channel (CaV2.1). This presynaptic high-voltage-gated channel couples neuronal excitation to the vesicular release of neurotransmitter and is implicated in several neurologic disorders. DNA Sanger sequencing confirmed that the de novo mutation was absent in both parents and present in the child only. Electrophysiologic analysis of P1353L-CACNA1A demonstrated near complete loss of function, with a 95% reduction in peak current density. Whole exome sequencing coupled with cellular electrophysiologic functional analysis of a de novoCACNA1A missense mutation has elucidated the probable underlying pathophysiologic mechanism responsible for the child's phenotype. Genetic testing of CACNA1A in patients with congenital hypotonia and developmental delay may be warranted. Copyright © 2016. Published by Elsevier Inc.

Fibromodulin deficiency reduces collagen structural network but not glycosaminoglycan content in a syngeneic model of colon carcinoma.

PubMed

Olsson, P Olof; Kalamajski, Sebastian; Maccarana, Marco; Oldberg, Åke; Rubin, Kristofer

2017-01-01

Tumor barrier function in carcinoma represents a major challenge to treatment and is therefore an attractive target for increasing drug delivery. Variables related to tumor barrier include aberrant blood vessels, high interstitial fluid pressure, and the composition and structure of the extracellular matrix. One of the proteins associated with dense extracellular matrices is fibromodulin, a collagen fibrillogenesis modulator expressed in tumor stroma but scarce in normal loose connective tissues. Here, we investigated the effects of fibromodulin on stroma ECM in a syngeneic murine colon carcinoma model. We show that fibromodulin deficiency decreased collagen fibril thickness but glycosaminoglycan content and composition were unchanged. Furthermore, vascular density, pericyte coverage and macrophage amount were unaffected. Fibromodulin can therefore be a unique effector of dense collagen matrix assembly in tumor stroma and, without affecting other major matrix components or the cellular composition, can function as a main agent in tumor barrier function.

[Structure and function of suburothelial myofibroblasts in the human urinary bladder under normal and pathological conditions].

PubMed

Neuhaus, J; Heinrich, M; Schlichting, N; Oberbach, A; Fitzl, G; Schwalenberg, T; Horn, L-C; Stolzenburg, J-U

2007-09-01

Myofibroblasts play a pivotal role in numerous pathological alterations. Clarification of the structure and function and of the cellular plasticity of this cell type in the bladder may lead to new insights into the pathogenesis of lower urinary tract disorders. Bladder biopsies from patients with bladder carcinoma and interstitial cystitis were used to analyse the morphology and receptor expression using confocal immunofluorescence and electron microscopy. Cytokine effects and coupling behavior were tested in cultured myofibroblasts and detrusor smooth muscle cells. Myofibroblasts are in close contact with the suburothelial capillary network. They express Cx43 and form functional syncytia. The expression of muscarinic and purinergic receptors is highly variable. Dye coupling experiments showed differences to detrusor myocytes. Upregulation of smooth muscle cell alpha-actin and/or transdifferentiation into smooth muscle cells may contribute to the etiology of urge incontinence. A multi-step model is presented as a working hypothesis.

NF-κB Essential Modulator (NEMO) Is Critical for Thyroid Function.

PubMed

Reale, Carla; Iervolino, Anna; Scudiero, Ivan; Ferravante, Angela; D'Andrea, Luca Egildo; Mazzone, Pellegrino; Zotti, Tiziana; Leonardi, Antonio; Roberto, Luca; Zannini, Mariastella; de Cristofaro, Tiziana; Shanmugakonar, Muralitharan; Capasso, Giovambattista; Pasparakis, Manolis; Vito, Pasquale; Stilo, Romania

2016-03-11

The I-κB kinase (IKK) subunit NEMO/IKKγ (NEMO) is an adapter molecule that is critical for canonical activation of NF-κB, a pleiotropic transcription factor controlling immunity, differentiation, cell growth, tumorigenesis, and apoptosis. To explore the functional role of canonical NF-κB signaling in thyroid gland differentiation and function, we have generated a murine strain bearing a genetic deletion of the NEMO locus in thyroid. Here we show that thyrocyte-specific NEMO knock-out mice gradually develop hypothyroidism after birth, which leads to reduced body weight and shortened life span. Histological and molecular analysis indicate that absence of NEMO in thyrocytes results in a dramatic loss of the thyroid gland cellularity, associated with down-regulation of thyroid differentiation markers and ongoing apoptosis. Thus, NEMO-dependent signaling is essential for normal thyroid physiology. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Seasonal differences in the physiology of Carcinus maenas (Crustacea: Decapoda) from estuaries with varying levels of anthropogenic contamination

NASA Astrophysics Data System (ADS)

Dissanayake, Awantha; Galloway, Tamara S.; Jones, Malcolm B.

2011-07-01

This study reports the seasonal variability in aspects of the physiology of the shore crab Carcinus maenas from three estuaries in South-west England, each with varying anthropogenic inputs: Avon Estuary ('relatively low' impact), Yealm Estuary ('intermediate' impact) and Plym Estuary ('relatively high' impact). Crabs collected over 12 months from the Avon had a significantly 'lower' physiological condition in winter and spring compared to summer and autumn; in particular, haemocyte phagocytic capability (a general indicator of immune function) was significantly higher in winter and spring compared to summer and autumn, and total haemolymph antioxidant status (an indicator of oxidative stress) was significantly lower in winter compared to the remainder of the year. Potentially, shore crabs may be more susceptible to the effects of contaminant exposure, such as increased immunotoxicity (thus, reduction of immune function) and/or oxyradicals (or reactive oxygen species) exposure) especially in seasons of increased susceptibility i.e. summer/autumn (lower phagocytic capability) and winter (lowest antioxidant function). As the Avon was taken to represent the 'reference' site, this pattern is considered to reflect the 'normal' seasonal variability in shore crab physiology. Shore crab physiological condition from the 'relatively high' impact estuary (Plym) revealed increased cellular viability and antioxidant status in autumn and winter compared with that of the 'standard' pattern (Avon) However, crabs from the intermediate impact estuary (Yealm) only demonstrated significant physiological differences in summer as shown by a lower cellular viability. All crabs had been exposed to PAHs (confirmed by the presence of PAH metabolites in their urine) which may account for the observed differences in shore crab physiology. In conclusion, to aid understanding of the potential contaminant impacts on biota it is imperative that the 'normal' seasonal variability of physiological condition be established. Biological effects-based monitoring studies should therefore be employed seasonally to potentially highlight 'windows of sensitivity' to contaminant impact.

A large shRNA library approach identifies lncRNA Ntep as an essential regulator of cell proliferation

PubMed Central

Beermann, Julia; Kirste, Dominique; Iwanov, Katharina; Lu, Dongchao; Kleemiß, Felix; Kumarswamy, Regalla; Schimmel, Katharina; Bär, Christian; Thum, Thomas

2018-01-01

The mammalian cell cycle is a complex and tightly controlled event. Myriads of different control mechanisms are involved in its regulation. Long non-coding RNAs (lncRNA) have emerged as important regulators of many cellular processes including cellular proliferation. However, a more global and unbiased approach to identify lncRNAs with importance for cell proliferation is missing. Here, we present a lentiviral shRNA library-based approach for functional lncRNA profiling. We validated our library approach in NIH3T3 (3T3) fibroblasts by identifying lncRNAs critically involved in cell proliferation. Using stringent selection criteria we identified lncRNA NR_015491.1 out of 3842 different RNA targets represented in our library. We termed this transcript Ntep (non-coding transcript essential for proliferation), as a bona fide lncRNA essential for cell cycle progression. Inhibition of Ntep in 3T3 and primary fibroblasts prevented normal cell growth and expression of key fibroblast markers. Mechanistically, we discovered that Ntep is important to activate P53 concomitant with increased apoptosis and cell cycle blockade in late G2/M. Our findings suggest Ntep to serve as an important regulator of fibroblast proliferation and function. In summary, our study demonstrates the applicability of an innovative shRNA library approach to identify long non-coding RNA functions in a massive parallel approach. PMID:29099486

Staying young at heart: autophagy and adaptation to cardiac aging.

PubMed

Leon, Leonardo J; Gustafsson, Åsa B

2016-06-01

Aging is a predominant risk factor for developing cardiovascular disease. Therefore, the cellular processes that contribute to aging are attractive targets for therapeutic interventions that can delay or prevent the development of age-related diseases. Our understanding of the underlying mechanisms that contribute to the decline in cell and tissue functions with age has greatly advanced over the past decade. Classical hallmarks of aging cells include increased levels of reactive oxygen species, DNA damage, accumulation of dysfunctional organelles, oxidized proteins and lipids. These all contribute to a progressive decline in the normal physiological function of the cell and to the onset of age-related conditions. A major cause of the aging process is progressive loss of cellular quality control. Autophagy is an important quality control pathway and is necessary to maintain cardiac homeostasis and to adapt to stress. A reduction in autophagy has been observed in a number of aging models and there is compelling evidence that enhanced autophagy delays aging and extends life span. Enhancing autophagy counteracts age-associated accumulation of protein aggregates and damaged organelles in cells. In this review, we discuss the functional role of autophagy in maintaining homeostasis in the heart, and how a decline is associated with accelerated cardiac aging. We also evaluate therapeutic approaches being researched in an effort to maintain a healthy young heart. Copyright © 2015 Elsevier Ltd. All rights reserved.

Estrogen biology: new insights into GPER function and clinical opportunities.

PubMed

Prossnitz, Eric R; Barton, Matthias

2014-05-25

Estrogens play an important role in the regulation of normal physiology, aging and many disease states. Although the nuclear estrogen receptors have classically been described to function as ligand-activated transcription factors mediating genomic effects in hormonally regulated tissues, more recent studies reveal that estrogens also mediate rapid signaling events traditionally associated with G protein-coupled receptors. The G protein-coupled estrogen receptor GPER (formerly GPR30) has now become recognized as a major mediator of estrogen's rapid cellular effects throughout the body. With the discovery of selective synthetic ligands for GPER, both agonists and antagonists, as well as the use of GPER knockout mice, significant advances have been made in our understanding of GPER function at the cellular, tissue and organismal levels. In many instances, the protective/beneficial effects of estrogen are mimicked by selective GPER agonism and are absent or reduced in GPER knockout mice, suggesting an essential or at least parallel role for GPER in the actions of estrogen. In this review, we will discuss recent advances and our current understanding of the role of GPER and the activity of clinically used drugs, such as SERMs and SERDs, in physiology and disease. We will also highlight novel opportunities for clinical development towards GPER-targeted therapeutics, for molecular imaging, as well as for theranostic approaches and personalized medicine. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Advancing haematopoietic stem and progenitor cell biology through single-cell profiling.

PubMed

Hamey, Fiona K; Nestorowa, Sonia; Wilson, Nicola K; Göttgens, Berthold

2016-11-01

Haematopoietic stem and progenitor cells (HSPCs) sit at the top of the haematopoietic hierarchy, and their fate choices need to be carefully controlled to ensure balanced production of all mature blood cell types. As cell fate decisions are made at the level of the individual cells, recent technological advances in measuring gene and protein expression in increasingly large numbers of single cells have been rapidly adopted to study both normal and pathological HSPC function. In this review we emphasise the importance of combining the correct computational models with single-cell experimental techniques, and illustrate how such integrated approaches have been used to resolve heterogeneities in populations, reconstruct lineage differentiation, identify regulatory relationships and link molecular profiling to cellular function. © 2016 Federation of European Biochemical Societies.

The solid-state signaling pathway from extracellular matrix to nuclear matrix: The critical role of three-dimensional architecture for functional differentiation

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

Lelievre, S.; Bissell, M.J.

Breast cells are useful experimental subjects for cell biologists because the mammary gland is one of the few tissues that undergoes dramatic changes in form and function after adulthood. Recently, the study in our laboratory of a human breast tumor progression series has allowed for the analysis of changes in cellular architecture (including nuclear architecture) when phenotypically normal cells become tumorigenic. This research aims to participate in the battle against breast cancer by helping to understand tumor progression and to identify new therapeutic markers for cancer treatment. This article explores the advantages and challenges of using high resolution X-ray computedmore » microtomography for the study of 3-dimensional organization of breast tissue architecture.« less

Pathomimetic cancer avatars for live-cell imaging of protease activity

PubMed Central

Ji, Kyungmin; Heyza, Joshua; Cavallo-Medved, Dora; Sloane, Bonnie F.

2016-01-01

Proteases are essential for normal physiology as well as multiple diseases, e.g., playing a causative role in cancer progression, including in tumor angiogenesis, invasion, and metastasis. Identification of dynamic alterations in protease activity may allow us to detect early stage cancers and to assess the efficacy of anti-cancer therapies. Despite the clinical importance of proteases in cancer progression, their functional roles individually and within the context of complex protease networks have not yet been well defined. These gaps in our understanding might be addressed with: 1) accurate and sensitive tools and methods to directly identify changes in protease activities in live cells, and 2) pathomimetic avatars for cancer that recapitulate in vitro the tumor in the context of its cellular and non-cellular microenvironment. Such avatars should be designed to facilitate mechanistic studies that can be translated to animal models and ultimately the clinic. Here, we will describe basic principles and recent applications of live-cell imaging for identification of active proteases. The avatars optimized by our laboratory are three-dimensional (3D) human breast cancer models in a matrix of reconstituted basement membrane (rBM). They are designated mammary architecture and microenvironment engineering (MAME) models as they have been designed to mimic the structural and functional interactions among cell types in the normal and cancerous human breast. We have demonstrated the usefulness of these pathomimetic avatars for following dynamic and temporal changes in cell:cell interactions and quantifying changes in protease activity associated with these interactions in real-time (4D). We also briefly describe adaptation of the avatars to custom-designed and fabricated tissue architecture and microenvironment engineering (TAME) chambers that enhance our ability to analyze concomitant changes in the malignant phenotype and the associated tumor microenvironment. PMID:26375517

Pathomimetic cancer avatars for live-cell imaging of protease activity.

PubMed

Ji, Kyungmin; Heyza, Joshua; Cavallo-Medved, Dora; Sloane, Bonnie F

2016-03-01

Proteases are essential for normal physiology as well as multiple diseases, e.g., playing a causative role in cancer progression, including in tumor angiogenesis, invasion, and metastasis. Identification of dynamic alterations in protease activity may allow us to detect early stage cancers and to assess the efficacy of anti-cancer therapies. Despite the clinical importance of proteases in cancer progression, their functional roles individually and within the context of complex protease networks have not yet been well defined. These gaps in our understanding might be addressed with: 1) accurate and sensitive tools and methods to directly identify changes in protease activities in live cells, and 2) pathomimetic avatars for cancer that recapitulate in vitro the tumor in the context of its cellular and non-cellular microenvironment. Such avatars should be designed to facilitate mechanistic studies that can be translated to animal models and ultimately the clinic. Here, we will describe basic principles and recent applications of live-cell imaging for identification of active proteases. The avatars optimized by our laboratory are three-dimensional (3D) human breast cancer models in a matrix of reconstituted basement membrane (rBM). They are designated mammary architecture and microenvironment engineering (MAME) models as they have been designed to mimic the structural and functional interactions among cell types in the normal and cancerous human breast. We have demonstrated the usefulness of these pathomimetic avatars for following dynamic and temporal changes in cell:cell interactions and quantifying changes in protease activity associated with these interactions in real-time (4D). We also briefly describe adaptation of the avatars to custom-designed and fabricated tissue architecture and microenvironment engineering (TAME) chambers that enhance our ability to analyze concomitant changes in the malignant phenotype and the associated tumor microenvironment. Copyright © 2015. Published by Elsevier B.V.

Production of cattle lacking prion protein

PubMed Central

Richt, Jürgen A; Kasinathan, Poothappillai; Hamir, Amir N; Castilla, Joaquin; Sathiyaseelan, Thillai; Vargas, Francisco; Sathiyaseelan, Janaki; Wu, Hua; Matsushita, Hiroaki; Koster, Julie; Kato, Shinichiro; Ishida, Isao; Soto, Claudio; Robl, James M; Kuroiwa, Yoshimi

2010-01-01

Prion diseases are caused by propagation of misfolded forms of the normal cellular prion protein PrPC, such as PrPBSE in bovine spongiform encephalopathy (BSE) in cattle and PrPCJD in Creutzfeldt-Jakob disease (CJD) in humans1. Disruption of PrPC expression in mice, a species that does not naturally contract prion diseases, results in no apparent developmental abnormalities2–5. However, the impact of ablating PrPC function in natural host species of prion diseases is unknown. Here we report the generation and characterization of PrPC-deficient cattle produced by a sequential gene-targeting system6. At over 20 months of age, the cattle are clinically, physiologically, histopathologically, immunologically and reproductively normal. Brain tissue homogenates are resistant to prion propagation in vitro as assessed by protein misfolding cyclic amplification7. PrPC-deficient cattle may be a useful model for prion research and could provide industrial bovine products free of prion proteins. PMID:17195841

Adenomatous Polyposis Coli-Mediated Accumulation of Abasic DNA Lesions Lead to Cigarette Smoke Condensate-Induced Neoplastic Transformation of Normal Breast Epithelial Cells1

PubMed Central

Jaiswal, Aruna S; Panda, Harekrushna; Pampo, Christine A; Siemann, Dietmar W; Gairola, C Gary; Hromas, Robert; Narayan, Satya

2013-01-01

Adenomatous polyposis coli (APC) is a multifunctional protein having diverse cellular functions including cell migration, cell-cell adhesion, cell cycle control, chromosomal segregation, and apoptosis. Recently, we found a new role of APC in base excision repair (BER) and showed that it interacts with DNA polymerase β and 5′-flap endonuclease 1 and interferes in BER. Previously, we have also reported that cigarette smoke condensate (CSC) increases expression of APC and enhances the growth of normal human breast epithelial (MCF10A) cells in vitro. In the present study, using APC overexpression and knockdown systems, we have examined the molecular mechanisms by which CSC and its major component, Benzo[α]pyrene, enhances APC-mediated accumulation of abasic DNA lesions, which is cytotoxic and mutagenic in nature, leading to enhanced neoplastic transformation of MCF10A cells in an orthotopic xenograft model. PMID:23555190

Exosomes: an overview of biogenesis, composition and role in ovarian cancer

PubMed Central

2014-01-01

Exosomes are tiny membrane-bound vesicles that are over produced by most proliferating cell types during normal and pathological states. Their levels are up-regulated during pregnancy and disease states such as cancer. Exosomes contain a wide variety of proteins, lipids, RNAs, non-transcribed RNAs, microRNAs and small RNAs that are representative to their cellular origin and shuttle from a donor cell to a recipient cell. From intercellular communication to tumor proliferation, exosomes carry out a diverse range of functions, both helpful and harmful. Useful as biomarkers, exosomes may be applicable in diagnostic assessments as well as cell-free anti-tumor vaccines. Exosomes of ovarian cancer contain different set of proteins and miRNAs compared to exosomes of normal, cancer-free individuals. These molecules may be used as multiple “barcode” for the development of a diagnostic tool for early detection of ovarian cancer. PMID:24460816

L1 Antibodies Block Lymph Node Fibroblastic Reticular Matrix Remodeling In Vivo

PubMed Central

Di Sciullo, Gino; Donahue, Tim; Schachner, Melitta; Bogen, Steven A.

1998-01-01

L1 is an immunoglobulin superfamily adhesion molecule highly expressed on neurons and involved in cell motility, neurite outgrowth, axon fasciculation, myelination, and synaptic plasticity. L1 is also expressed by nonneural cells, but its function outside of the nervous system has not been studied extensively. We find that administration of an L1 monoclonal antibody in vivo disrupts the normal remodeling of lymph node reticular matrix during an immune response. Ultrastructural examination reveals that reticular fibroblasts in mice treated with L1 monoclonal antibodies fail to spread and envelop collagen fibers with their cellular processes. The induced defect in the remodeling of the fibroblastic reticular system results in the loss of normal nodal architecture, collapsed cortical sinusoids, and macrophage accumulation in malformed sinuses. Surprisingly, such profound architectural abnormalities have no detectable effects on the primary immune response to protein antigens. PMID:9625755

Cellular senescence and organismal aging.

PubMed

Jeyapalan, Jessie C; Sedivy, John M

2008-01-01

Cellular senescence, first observed and defined using in vitro cell culture studies, is an irreversible cell cycle arrest which can be triggered by a variety of factors. Emerging evidence suggests that cellular senescence acts as an in vivo tumor suppression mechanism by limiting aberrant proliferation. It has also been postulated that cellular senescence can occur independently of cancer and contribute to the physiological processes of normal organismal aging. Recent data have demonstrated the in vivo accumulation of senescent cells with advancing age. Some characteristics of senescent cells, such as the ability to modify their extracellular environment, could play a role in aging and age-related pathology. In this review, we examine current evidence that links cellular senescence and organismal aging.

Cellular senescence and organismal aging

PubMed Central

Jeyapalan, Jessie C.; Sedivy, John M.

2012-01-01

Cellular senescence, first observed and defined using in vitro cell culture studies, is an irreversible cell cycle arrest which can be triggered by a variety of factors. Emerging evidence suggests that cellular senescence acts as an in vivo tumor suppression mechanism by limiting aberrant proliferation. It has also been postulated that cellular senescence can occur independently of cancer and contribute to the physiological processes of normal organismal aging. Recent data have demonstrated the in vivo accumulation of senescent cells with advancing age. Some characteristics of senescent cells, such as the ability to modify their extracellular environment, could play a role in aging and age related pathology. In this review, we examine current evidence that links cellular senescence and organismal aging. PMID:18502472

Simultaneous cryo X-ray ptychographic and fluorescence microscopy of green algae

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

Deng, Junjing; Vine, David J.; Chen, Si

Trace metals play important roles in normal and in disease-causing biological functions. X-ray fluorescence microscopy reveals trace elements with no dependence on binding affinities (unlike with visible light fluorophores) and with improved sensitivity relative to electron probes. However, X-ray fluorescence is not very sensitive for showing the light elements that comprise the majority of cellular material. Here we show that X-ray ptychography can be combined with fluorescence to image both cellular structure and trace element distribution in frozen-hydrated cells at cryogenic temperatures, with high structural and chemical fidelity. Ptychographic reconstruction algorithms deliver phase and absorption contrast images at a resolutionmore » beyond that of the illuminating lens or beam size. Using 5.2-keV X-rays, we have obtained sub-30-nm resolution structural images and ~90-nm-resolution fluorescence images of several elements in frozen-hydrated green algae. Finally, this combined approach offers a way to study the role of trace elements in their structural context.« less

Simultaneous cryo X-ray ptychographic and fluorescence microscopy of green algae

DOE PAGES

Deng, Junjing; Vine, David J.; Chen, Si; ...

2015-02-24

Trace metals play important roles in normal and in disease-causing biological functions. X-ray fluorescence microscopy reveals trace elements with no dependence on binding affinities (unlike with visible light fluorophores) and with improved sensitivity relative to electron probes. However, X-ray fluorescence is not very sensitive for showing the light elements that comprise the majority of cellular material. Here we show that X-ray ptychography can be combined with fluorescence to image both cellular structure and trace element distribution in frozen-hydrated cells at cryogenic temperatures, with high structural and chemical fidelity. Ptychographic reconstruction algorithms deliver phase and absorption contrast images at a resolutionmore » beyond that of the illuminating lens or beam size. Using 5.2-keV X-rays, we have obtained sub-30-nm resolution structural images and ~90-nm-resolution fluorescence images of several elements in frozen-hydrated green algae. Finally, this combined approach offers a way to study the role of trace elements in their structural context.« less

Simultaneous cryo X-ray ptychographic and fluorescence microscopy of green algae

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

Deng, Junjing; Vine, David J.; Chen, Si

Trace metals play important roles in normal and in disease-causing biological functions. X-ray fluorescence microscopy reveals trace elements with no dependence on binding affinities (unlike with visible light fluorophores) and with improved sensitivity relative to electron probes. However, X-ray fluorescence is not very sensitive for showing the light elements that comprise the majority of cellular material. Here we show that X-ray ptychography can be combined with fluorescence to image both cellular structure and trace element distribution in frozen-hydrated cells at cryogenic temperatures, with high structural and chemical fidelity. Ptychographic reconstruction algorithms deliver phase and absorption contrast images at a resolutionmore » beyond that of the illuminating lens or beam size. Using 5.2-keV X-rays, we have obtained sub-30-nm resolution structural images and similar to 90-nm-resolution fluorescence images of several elements in frozen-hydrated green algae. This combined approach offers a way to study the role of trace elements in their structural context.« less

Dual-Functional Nanographene Oxide as Cancer-Targeted Drug-Delivery System to Selectively Induce Cancer-Cell Apoptosis.

PubMed

Zhou, Binwei; Huang, Yanyu; Yang, Fang; Zheng, Wenjie; Chen, Tianfeng

2016-04-05

Construction of bioresponsive drug-delivery nanosystems could enhance the anticancer efficacy of anticancer agents and reduce their toxic side effects. Herein, by using transferrin (Tf) as a surface decorator, we constructed a cancer-targeted nanographene oxide (NGO) nanosystem for use in drug delivery. This nanosystem (Tf-NGO@HPIP) drastically enhanced the cellular uptake, retention, and anticancer efficacy of loaded drugs but showed much lower toxicity to normal cells. The nanosystem was internalized through receptor-mediated endocytosis and triggered pH-dependent drug release in acidic environments and in the presence of cellular enzymes. Moreover, Tf-NGO@HPIP effectively induced cancer-cell apoptosis through activation of superoxide-mediated p53 and MAPK pathways along with inactivation of ERK and AKT. Taken together, this study demonstrates a good strategy for the construction of bioresponsive NGO drug-delivery nanosystems and their use as efficient anticancer drug carriers. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Altered proliferation and networks in neural cells derived from idiopathic autistic individuals.

PubMed

Marchetto, Maria C; Belinson, Haim; Tian, Yuan; Freitas, Beatriz C; Fu, Chen; Vadodaria, Krishna; Beltrao-Braga, Patricia; Trujillo, Cleber A; Mendes, Ana P D; Padmanabhan, Krishnan; Nunez, Yanelli; Ou, Jing; Ghosh, Himanish; Wright, Rebecca; Brennand, Kristen; Pierce, Karen; Eichenfield, Lawrence; Pramparo, Tiziano; Eyler, Lisa; Barnes, Cynthia C; Courchesne, Eric; Geschwind, Daniel H; Gage, Fred H; Wynshaw-Boris, Anthony; Muotri, Alysson R

2017-06-01

Autism spectrum disorders (ASD) are common, complex and heterogeneous neurodevelopmental disorders. Cellular and molecular mechanisms responsible for ASD pathogenesis have been proposed based on genetic studies, brain pathology and imaging, but a major impediment to testing ASD hypotheses is the lack of human cell models. Here, we reprogrammed fibroblasts to generate induced pluripotent stem cells, neural progenitor cells (NPCs) and neurons from ASD individuals with early brain overgrowth and non-ASD controls with normal brain size. ASD-derived NPCs display increased cell proliferation because of dysregulation of a β-catenin/BRN2 transcriptional cascade. ASD-derived neurons display abnormal neurogenesis and reduced synaptogenesis leading to functional defects in neuronal networks. Interestingly, defects in neuronal networks could be rescued by insulin growth factor 1 (IGF-1), a drug that is currently in clinical trials for ASD. This work demonstrates that selection of ASD subjects based on endophenotypes unraveled biologically relevant pathway disruption and revealed a potential cellular mechanism for the therapeutic effect of IGF-1.

A cellular, molecular, and pharmacological basis for appendage regeneration in mice.

PubMed

Leung, Thomas H; Snyder, Emily R; Liu, Yinghua; Wang, Jing; Kim, Seung K

2015-10-15

Regenerative medicine aims to restore normal tissue architecture and function. However, the basis of tissue regeneration in mammalian solid organs remains undefined. Remarkably, mice lacking p21 fully regenerate injured ears without discernable scarring. Here we show that, in wild-type mice following tissue injury, stromal-derived factor-1 (Sdf1) is up-regulated in the wound epidermis and recruits Cxcr4-expressing leukocytes to the injury site. In p21-deficient mice, Sdf1 up-regulation and the subsequent recruitment of Cxcr4-expressing leukocytes are significantly diminished, thereby permitting scarless appendage regeneration. Lineage tracing demonstrates that this regeneration derives from fate-restricted progenitor cells. Pharmacological or genetic disruption of Sdf1-Cxcr4 signaling enhances tissue repair, including full reconstitution of tissue architecture and all cell types. Our findings identify signaling and cellular mechanisms underlying appendage regeneration in mice and suggest new therapeutic approaches for regenerative medicine. © 2015 Leung et al.; Published by Cold Spring Harbor Laboratory Press.

Raf kinase inhibitory protein: a signal transduction modulator and metastasis suppressor.

PubMed

Granovsky, Alexey E; Rosner, Marsha Rich

2008-04-01

Cells have a multitude of controls to maintain their integrity and prevent random switching from one biological state to another. Raf Kinase Inhibitory Protein (RKIP), a member of the phosphatidylethanolamine binding protein (PEBP) family, is representative of a new class of modulators of signaling cascades that function to maintain the "yin yang" or balance of biological systems. RKIP inhibits MAP kinase (Raf-MEK-ERK), G protein-coupled receptor (GPCR) kinase and NFkappaB signaling cascades. Because RKIP targets different kinases dependent upon its state of phosphorylation, RKIP also acts to integrate crosstalk initiated by multiple environmental stimuli. Loss or depletion of RKIP results in disruption of the normal cellular stasis and can lead to chromosomal abnormalities and disease states such as cancer. Since RKIP and the PEBP family have been reviewed previously, the goal of this analysis is to provide an update and highlight some of the unique features of RKIP that make it a critical player in the regulation of cellular signaling processes.

Effect of simulated microgravity and shear stress on microcin B17 production by Escherichia coli and on its excretion into the medium

NASA Technical Reports Server (NTRS)

Fang, A.; Pierson, D. L.; Koenig, D. W.; Mishra, S. K.; Demain, A. L.

1997-01-01

Production of the antibacterial polypeptide microcin B17 (MccB17) by Escherichia coli ZK650 was inhibited by simulated microgravity. The site of MccB17 accumulation was found to be different, depending on whether the organism was grown in shaking flasks or in rotating bioreactors designed to establish a simulated microgravity environment. In flasks, the accumulation was cellular, but in the reactors, virtually all the microcin was found in the medium. The change from a cellular site to an extracellular one was apparently not a function of gravity, since extracellular production occurred in these bioreactors, irrespective of whether they were operated in the simulated microgravity or normal gravity mode. More probably, excretion is due to the much lower degree of shear stress in the bioreactors. Addition of even a single glass bead to the 50-ml medium volume in the bioreactor created enough shear to change the site of MccB17 accumulation from the medium to the cells.

The Polarity Protein Partitioning-defective 1 (PAR-1) Regulates Dendritic Spine Morphogenesis through Phosphorylating Postsynaptic Density Protein 95 (PSD-95)*

PubMed Central

Wu, Qian; DiBona, Victoria L.; Bernard, Laura P.; Zhang, Huaye

2012-01-01

The polarity protein PAR-1 plays an essential role in many cellular contexts, including embryogenesis, asymmetric cell division, directional migration, and epithelial morphogenesis. Despite its known importance in different cellular processes, the role of PAR-1 in neuronal morphogenesis is less well understood. In particular, its role in the morphogenesis of dendritic spines, which are sites of excitatory synaptic inputs, has been unclear. Here, we show that PAR-1 is required for normal spine morphogenesis in hippocampal neurons. We further show that PAR-1 functions through phosphorylating the synaptic scaffolding protein PSD-95 in this process. Phosphorylation at a conserved serine residue in the KXGS motif in PSD-95 regulates spine morphogenesis, and a phosphomimetic mutant of this site can rescue the defects of kinase-dead PAR-1. Together, our findings uncover a role of PAR-1 in spine morphogenesis in hippocampal neurons through phosphorylating PSD-95. PMID:22807451

Contribution of cellular retinol-binding protein type 1 to retinol metabolism during mouse development.

PubMed

Matt, Nicolas; Schmidt, Carsten K; Dupé, Valérie; Dennefeld, Christine; Nau, Heinz; Chambon, Pierre; Mark, Manuel; Ghyselinck, Norbert B

2005-05-01

Within cells, retinol (ROL) is bound to cytoplasmic proteins (cellular retinol-binding proteins [CRBPs]), whose proposed function is to protect it from unspecific enzymes through channeling to retinoid-metabolizing pathways. We show that, during development, ROL and retinyl ester levels are decreased in CRBP type 1 (CRBP1) -deficient embryos and fetuses by 50% and 80%, respectively. The steady state level of retinoic acid (RA) is also decreased but to a lesser extent. However, CRBP1-null fetuses do not exhibit the abnormalities characteristic of a vitamin A-deficiency syndrome. Neither CRBP1 deficiency alters the expression patterns of RA-responding genes during development, nor does CRBP1 availability modify the expression of an RA-dependent gene in primary embryonic fibroblasts treated with ROL. Therefore, CRBP1 is required in prenatal life to maintain normal amounts of ROL and to ensure its efficient storage but seems of secondary importance for RA synthesis, at least under conditions of maternal vitamin A sufficiency. Copyright 2005 Wiley-Liss, Inc.

More to NAD+ than meets the eye: A regulator of metabolic pools and gene expression in Arabidopsis.

PubMed

Gakière, Bertrand; Fernie, Alisdair R; Pétriacq, Pierre

2018-01-05

Since its discovery more than a century ago, nicotinamide adenine dinucleotide (NAD + ) is recognised as a fascinating cornerstone of cellular metabolism. This ubiquitous energy cofactor plays vital roles in metabolic pathways and regulatory processes, a fact emphasised by the essentiality of a balanced NAD + metabolism for normal plant growth and development. Research on the role of NAD in plants has been predominantly carried out in the model plant Arabidopsis thaliana (Arabidopsis) with emphasis on the redox properties and cellular signalling functions of the metabolite. This review examines the current state of knowledge concerning how NAD can regulate both metabolic pools and gene expression in Arabidopsis. Particular focus is placed on recent studies highlighting the complexity of metabolic regulations involving NAD, more particularly in the mitochondrial compartment, and of signalling roles with respect to interactions with environmental fluctuations most specifically those involving plant immunity. Copyright © 2018 Elsevier Inc. All rights reserved.

Deficiency of Huntingtin Has Pleiotropic Effects in the Social Amoeba Dictyostelium discoideum

PubMed Central

Myre, Michael A.; Lumsden, Amanda L.; Thompson, Morgan N.; Wasco, Wilma; MacDonald, Marcy E.; Gusella, James F.

2011-01-01

Huntingtin is a large HEAT repeat protein first identified in humans, where a polyglutamine tract expansion near the amino terminus causes a gain-of-function mechanism that leads to selective neuronal loss in Huntington's disease (HD). Genetic evidence in humans and knock-in mouse models suggests that this gain-of-function involves an increase or deregulation of some aspect of huntingtin's normal function(s), which remains poorly understood. As huntingtin shows evolutionary conservation, a powerful approach to discovering its normal biochemical role(s) is to study the effects caused by its deficiency in a model organism with a short life-cycle that comprises both cellular and multicellular developmental stages. To facilitate studies aimed at detailed knowledge of huntingtin's normal function(s), we generated a null mutant of hd, the HD ortholog in Dictyostelium discoideum. Dictyostelium cells lacking endogenous huntingtin were viable but during development did not exhibit the typical polarized morphology of Dictyostelium cells, streamed poorly to form aggregates by accretion rather than chemotaxis, showed disorganized F-actin staining, exhibited extreme sensitivity to hypoosmotic stress, and failed to form EDTA-resistant cell–cell contacts. Surprisingly, chemotactic streaming could be rescued in the presence of the bivalent cations Ca2+ or Mg2+ but not pulses of cAMP. Although hd − cells completed development, it was delayed and proceeded asynchronously, producing small fruiting bodies with round, defective spores that germinated spontaneously within a glassy sorus. When developed as chimeras with wild-type cells, hd − cells failed to populate the pre-spore region of the slug. In Dictyostelium, huntingtin deficiency is compatible with survival of the organism but renders cells sensitive to low osmolarity, which produces pleiotropic cell autonomous defects that affect cAMP signaling and as a consequence development. Thus, Dictyostelium provides a novel haploid organism model for genetic, cell biological, and biochemical studies to delineate the functions of the HD protein. PMID:21552328

Drosophila Uri, a PP1α binding protein, is essential for viability, maintenance of DNA integrity and normal transcriptional activity

PubMed Central

Kirchner, Jasmin; Vissi, Emese; Gross, Sascha; Szoor, Balazs; Rudenko, Andrey; Alphey, Luke; White-Cooper, Helen

2008-01-01

Background Protein phosphatase 1 (PP1) is involved in diverse cellular processes, and is targeted to substrates via interaction with many different protein binding partners. PP1 catalytic subunits (PP1c) fall into PP1α and PP1β subfamilies based on sequence analysis, however very few PP1c binding proteins have been demonstrated to discriminate between PP1α and PP1β. Results URI (unconventional prefoldin RPB5 interactor) is a conserved molecular chaperone implicated in a variety of cellular processes, including the transcriptional response to nutrient signalling and maintenance of DNA integrity. We show that Drosophila Uri binds PP1α with much higher affinity than PP1β, and that this ability to discriminate between PP1c forms is conserved to humans. Most Uri is cytoplasmic, however we found some protein associated with active RNAPII on chromatin. We generated a uri loss of function allele, and show that uri is essential for viability in Drosophila. uri mutants have transcriptional defects, reduced cell viability and differentiation in the germline, and accumulate DNA damage in their nuclei. Conclusion Uri is the first PP1α specific binding protein to be described in Drosophila. Uri protein plays a role in transcriptional regulation. Activity of uri is required to maintain DNA integrity and cell survival in normal development. PMID:18412953

Detection of early changes in lung cell cytology by flow-systems analysis techniques

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

Steinkamp, J.A.; Hansen, K.M.; Wilson, J.S.

1976-12-01

This report summarizes results of continuing experiments to develop cytological and biochemical indicators for estimating damage to respiratory cells in test animals exposed by inhalation to toxic agents associated with nonnuclear energy production, the specific goal being the application of advanced multiparameter flow-systems technologies to the detection of early atypical cellular changes in lung epithelium. Normal Syrian hamster lung cell samples composed of macrophages, leukocytes, ciliated columnar cells, and epithelial cells were stained with fluorescent dyes specific for different biochemical parameters and were analyzed in liquid suspension as they flowed through a chamber intersecting a laser beam of exciting light.more » Multiple sensors measured the total or two-color fluorescence and light scatter on a cell-by-cell basis. Cellular parameters proportional to optical measurements (i.e., cell size, DNA content, total protein, nonspecific esterase activity, nuclear and cytoplasmic diameters) were displayed as frequency distribution histograms. Lung cell samples were also separated according to various cytological parameters and identified microscopically. The basic operating features of the methodology are discussed briefly, along with specific examples of preliminary results illustrating the initial characterization of exfoliated pulmonary cells from normal hamsters. As the flow technology is adapted further to the analysis of respiratory cells, measurements of changes in physical and biochemical properties as a function of exposure to toxic agents will be performed.« less

Juvenile-onset loss of lipid-raft domains in attractin-deficient mice

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

Azouz, Abdallah; Gunn, Teresa M.; Duke-Cohan, Jonathan S.

2007-02-15

Mutations at the attractin (Atrn) locus in mice result in altered pigmentation on an agouti background, higher basal metabolic rate and juvenile-onset hypomyelination leading to neurodegeneration, while studies on human immune cells indicate a chemotaxis regulatory function. The underlying biochemical defect remains elusive. In this report we identify a role for attractin in plasma membrane maintenance. In attractin's absence there is a decline in plasma membrane glycolipid-enriched rafts from normal levels at 8 weeks to a complete absence by 24 weeks. The structural integrity of lipid rafts depends upon cholesterol and sphingomyelin, and can be identified by partitioning within ofmore » ganglioside GM{sub 1}. Despite a significant fall in cellular cholesterol with maturity, and a lesser fall in both membrane and total cellular GM{sub 1}, these parameters lag behind raft loss, and are normal when hypomyelination/neurodegeneration has already begun thus supporting consequence rather than cause. These findings can be recapitulated in Atrn-deficient cell lines propagated in vitro. Further, signal transduction through complex membrane receptor assemblies is not grossly disturbed despite the complete absence of lipid rafts. We find these results compatible with a role for attractin in plasma membrane maintenance and consistent with the proposal that the juvenile-onset hypomyelination and neurodegeneration represent a defect in attractin-mediated raft-dependent myelin biogenesis.« less

Insulin resistance in patients with recurrent pregnancy loss is associated with lymphocyte population aberration.

PubMed

Yan, Yan; Bao, Shihua; Sheng, Shile; Wang, Liuliu; Tu, Weiyan

2017-12-01

This study was designed to investigate the relationship of insulin resistance (IR) and cellular immune abnormalities associated with women with recurrent pregnancy loss (RPL). Women with RPL were divided into two groups according to their homeostasis model assessment for IR (HOMA-IR) scores. The IR group received metformin approximately 3 months before pregnancy. The percentage of lymphocyte subsets and other blood biochemical indices were tested. The HOMA-IR and fasting serum insulin levels were related to the percentage of lymphocyte subsets. The women with RPL had higher CD3 + and CD3 + CD4 + cell levels while CD56 + CD16 + cell levels were lower. A higher likelihood of cellular immune abnormalities was observed. Women with normal lymphocyte subsets had normal pregnancy outcomes. Metformin significantly downregulated CD3 + and CD3 + CD4 + cells and improved pregnancy outcomes. IR was associated with cellular immune abnormalities in RPL. The data suggests that metformin affected the immune/inflammatory response, which may regulate the cellular immune balance and improve pregnancy outcomes. Abbreviations RPL: recurrent pregnancy loss; IR insulin resistance; HOMA-IR: homeostasis model assessment for IR.

Robust Selection Algorithm (RSA) for Multi-Omic Biomarker Discovery; Integration with Functional Network Analysis to Identify miRNA Regulated Pathways in Multiple Cancers.

PubMed

Sehgal, Vasudha; Seviour, Elena G; Moss, Tyler J; Mills, Gordon B; Azencott, Robert; Ram, Prahlad T

2015-01-01

MicroRNAs (miRNAs) play a crucial role in the maintenance of cellular homeostasis by regulating the expression of their target genes. As such, the dysregulation of miRNA expression has been frequently linked to cancer. With rapidly accumulating molecular data linked to patient outcome, the need for identification of robust multi-omic molecular markers is critical in order to provide clinical impact. While previous bioinformatic tools have been developed to identify potential biomarkers in cancer, these methods do not allow for rapid classification of oncogenes versus tumor suppressors taking into account robust differential expression, cutoffs, p-values and non-normality of the data. Here, we propose a methodology, Robust Selection Algorithm (RSA) that addresses these important problems in big data omics analysis. The robustness of the survival analysis is ensured by identification of optimal cutoff values of omics expression, strengthened by p-value computed through intensive random resampling taking into account any non-normality in the data and integration into multi-omic functional networks. Here we have analyzed pan-cancer miRNA patient data to identify functional pathways involved in cancer progression that are associated with selected miRNA identified by RSA. Our approach demonstrates the way in which existing survival analysis techniques can be integrated with a functional network analysis framework to efficiently identify promising biomarkers and novel therapeutic candidates across diseases.

MicroRNAs in Leukemias: Emerging Diagnostic Tools and Therapeutic Targets

PubMed Central

Mian, Yousaf A.; Zeleznik-Le, Nancy J.

2010-01-01

MicroRNAs (miRNA) are small non-coding RNAs of ~22 nucleotides that regulate the translation and stability of mRNA to control different functions of the cell. Misexpression of miRNA has been linked to disruption of normal cellular functions, which results in various disorders including cancers such as leukemias. MicroRNA involvement in disease has been the subject of much attention and is increasing our current understanding of disease biology. Such linkages have been determined by high-throughput studies, which provide a framework for characterizing differential miRNA expression levels correlating to different cytogenetic abnormalities and their corresponding malignancies. In addition, functional studies of particular miRNAs have begun to define the effects of miRNA on predicted mRNA targets. It is clear that miRNAs can serve as molecular markers of leukemias and the hope is that they can also serve as new therapeutic targets. Studies are beginning to elucidate how to deliver therapeutic antagonists to attenuate overexpressed miRNAs and to replace underexpressed miRNAs. In this review, we: i) discuss the current understanding of miRNA function and expression in normal hematopoiesis, ii) provide examples of miRNAs that are misregulated in leukemias, and iii) evaluate the current status and potential future directions for the burgeoning field of antisense oligonucleotides and other therapeutic attempts to intervene in miRNA disregulation in leukemias. PMID:20370647

Normalizing glycosphingolipids restores function in CD4+ T cells from lupus patients

PubMed Central

McDonald, Georgia; Deepak, Shantal; Miguel, Laura; Hall, Cleo J.; Isenberg, David A.; Magee, Anthony I.; Butters, Terry; Jury, Elizabeth C.

2014-01-01

Patients with the autoimmune rheumatic disease systemic lupus erythematosus (SLE) have multiple defects in lymphocyte signaling and function that contribute to disease pathogenesis. Such defects could be attributed to alterations in metabolic processes, including abnormal control of lipid biosynthesis pathways. Here, we reveal that CD4+ T cells from SLE patients displayed an altered profile of lipid raft–associated glycosphingolipids (GSLs) compared with that of healthy controls. In particular, lactosylceramide, globotriaosylceramide (Gb3), and monosialotetrahexosylganglioside (GM1) levels were markedly increased. Elevated GSLs in SLE patients were associated with increased expression of liver X receptor β (LXRβ), a nuclear receptor that controls cellular lipid metabolism and trafficking and influences acquired immune responses. Stimulation of CD4+ T cells isolated from healthy donors with synthetic and endogenous LXR agonists promoted GSL expression, which was blocked by an LXR antagonist. Increased GSL expression in CD4+ T cells was associated with intracellular accumulation and accelerated trafficking of GSL, reminiscent of cells from patients with glycolipid storage diseases. Inhibition of GSL biosynthesis in vitro with a clinically approved inhibitor (N-butyldeoxynojirimycin) normalized GSL metabolism, corrected CD4+ T cell signaling and functional defects, and decreased anti-dsDNA antibody production by autologous B cells in SLE patients. Our data demonstrate that lipid metabolism defects contribute to SLE pathogenesis and suggest that targeting GSL biosynthesis restores T cell function in SLE. PMID:24463447

Normalizing glycosphingolipids restores function in CD4+ T cells from lupus patients.

PubMed

McDonald, Georgia; Deepak, Shantal; Miguel, Laura; Hall, Cleo J; Isenberg, David A; Magee, Anthony I; Butters, Terry; Jury, Elizabeth C

2014-02-01

Patients with the autoimmune rheumatic disease systemic lupus erythematosus (SLE) have multiple defects in lymphocyte signaling and function that contribute to disease pathogenesis. Such defects could be attributed to alterations in metabolic processes, including abnormal control of lipid biosynthesis pathways. Here, we reveal that CD4+ T cells from SLE patients displayed an altered profile of lipid raft-associated glycosphingolipids (GSLs) compared with that of healthy controls. In particular, lactosylceramide, globotriaosylceramide (Gb3), and monosialotetrahexosylganglioside (GM1) levels were markedly increased. Elevated GSLs in SLE patients were associated with increased expression of liver X receptor β (LXRβ), a nuclear receptor that controls cellular lipid metabolism and trafficking and influences acquired immune responses. Stimulation of CD4+ T cells isolated from healthy donors with synthetic and endogenous LXR agonists promoted GSL expression, which was blocked by an LXR antagonist. Increased GSL expression in CD4+ T cells was associated with intracellular accumulation and accelerated trafficking of GSL, reminiscent of cells from patients with glycolipid storage diseases. Inhibition of GSL biosynthesis in vitro with a clinically approved inhibitor (N-butyldeoxynojirimycin) normalized GSL metabolism, corrected CD4+ T cell signaling and functional defects, and decreased anti-dsDNA antibody production by autologous B cells in SLE patients. Our data demonstrate that lipid metabolism defects contribute to SLE pathogenesis and suggest that targeting GSL biosynthesis restores T cell function in SLE.

Addiction and the brain antireward system.

PubMed

Koob, George F; Le Moal, Michel

2008-01-01

A neurobiological model of the brain emotional systems has been proposed to explain the persistent changes in motivation that are associated with vulnerability to relapse in addiction, and this model may generalize to other psychopathology associated with dysregulated motivational systems. In this framework, addiction is conceptualized as a cycle of decreased function of brain reward systems and recruitment of antireward systems that progressively worsen, resulting in the compulsive use of drugs. Counteradaptive processes, such as opponent process, that are part of the normal homeostatic limitation of reward function fail to return within the normal homeostatic range and are hypothesized to repeatedly drive the allostatic state. Excessive drug taking thus results in not only the short-term amelioration of the reward deficit but also suppression of the antireward system. However, in the long term, there is worsening of the underlying neurochemical dysregulations that ultimately form an allostatic state (decreased dopamine and opioid peptide function, increased corticotropin-releasing factor activity). This allostatic state is hypothesized to be reflected in a chronic deviation of reward set point that is fueled not only by dysregulation of reward circuits per se but also by recruitment of brain and hormonal stress responses. Vulnerability to addiction may involve genetic comorbidity and developmental factors at the molecular, cellular, or neurocircuitry levels that sensitize the brain antireward systems.

Neurologic manifestations of major electrolyte abnormalities.

PubMed

Diringer, M

2017-01-01

The brain operates in an extraordinarily intricate environment which demands precise regulation of electrolytes. Tight control over their concentrations and gradients across cellular compartments is essential and when these relationships are disturbed neurologic manifestations may develop. Perturbations of sodium are the electrolyte disturbances that most often lead to neurologic manifestations. Alterations in extracellular fluid sodium concentrations produce water shifts that lead to brain swelling or shrinkage. If marked or rapid they can result in profound changes in brain function which are proportional to the degree of cerebral edema or contraction. Adaptive mechanisms quickly respond to changes in cell size by either increasing or decreasing intracellular osmoles in order to restore size to normal. Unless cerebral edema has been severe or prolonged, correction of sodium disturbances usually restores function to normal. If the rate of correction is too rapid or overcorrection occurs, however, new neurologic manifestations may appear as a result of osmotic demyelination syndrome. Disturbances of magnesium, phosphate and calcium all may contribute to alterations in sensorium. Hypomagnesemia and hypocalcemia can lead to weakness, muscle spasms, and tetany; the weakness from hypophosphatemia and hypomagnesemia can impair respiratory function. Seizures can be seen in cases with very low concentrations of sodium, magnesium, calcium, and phosphate. © 2017 Elsevier B.V. All rights reserved.

Reconstructing impairment of secretory ameloblast function in porcine teeth by analysis of morphological alterations in dental enamel

PubMed Central

Witzel, Carsten; Kierdorf, Uwe; Dobney, Keith; Ervynck, Anton; Vanpoucke, Sofie; Kierdorf, Horst

2006-01-01

We studied the relationship between the macroscopic appearance of hypoplastic defects in the dental enamel of wild boar and domestic pigs, and microstructural enamel changes, at both the light and the scanning electron microscopic levels. Deviations from normal enamel microstructure were used to reconstruct the functional and related morphological changes of the secretory ameloblasts caused by the action of stress factors during amelogenesis. The deduced reaction pattern of the secretory ameloblasts can be grouped in a sequence of increasingly severe impairments of cell function. The reactions ranged from a slight enhancement of the periodicity of enamel matrix secretion, over a temporary reduction in the amount of secreted enamel matrix, with reduction of the distal portion of the Tomes' process, to either a temporary or a definite cessation of matrix formation. The results demonstrate that analysis of structural changes in dental enamel allows a detailed reconstruction of the reaction of secretory ameloblasts to stress events, enabling an assessment of duration and intensity of these events. Analysing the deviations from normal enamel microstructure provides a deeper insight into the cellular changes underlying the formation of hypoplastic enamel defects than can be achieved by mere inspection of tooth surface characteristics alone. PMID:16822273

Programmed cell death promotes male sterility in the functional dioecious Opuntia stenopetala (Cactaceae)

PubMed Central

Flores-Rentería, Lluvia; Orozco-Arroyo, Gregorio; Cruz-García, Felipe; García-Campusano, Florencia; Alfaro, Isabel; Vázquez-Santana, Sonia

2013-01-01

Background and Aims The sexual separation in dioecious species has interested biologists for decades; however, the cellular mechanism leading to unisexuality has been poorly understood. In this study, the cellular changes that lead to male sterility in the functionally dioecious cactus, Opuntia stenopetala, are described. Methods The spatial and temporal patterns of programmed cell death (PCD) were determined in the anthers of male and female flowers using scanning electron microscopy analysis and histological observations, focusing attention on the transition from bisexual to unisexual development. In addition, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assays were used as an indicator of DNA fragmentation to corroborate PCD. Key results PCD was detected in anthers of both female and male flowers, but their patterns differed in time and space. Functionally male individuals developed viable pollen, and normal development involved PCD on each layer of the anther wall, which occurred progressively from the inner (tapetum) to the outer layer (epidermis). Conversely, functional female individuals aborted anthers by premature and displaced PCD. In anthers of female flowers, the first signs of PCD, such as a nucleus with irregular shape, fragmented and condensed chromatin, high vacuolization and condensed cytoplasm, occurred at the microspore mother cell stage. Later these features were observed simultaneously in all anther wall layers, connective tissue and filament. Neither pollen formation nor anther dehiscence was detected in female flowers of O. stenopetala due to total anther disruption. Conclusions Temporal and spatial changes in the patterns of PCD are responsible for male sterility of female flowers in O. stenopetala. Male fertility requires the co-ordination of different events, which, when altered, can lead to male sterility and to functionally unisexual individuals. PCD could be a widespread mechanism in the determination of functionally dioecious species. PMID:23877075

Programmed cell death promotes male sterility in the functional dioecious Opuntia stenopetala (Cactaceae).

PubMed

Flores-Rentería, Lluvia; Orozco-Arroyo, Gregorio; Cruz-García, Felipe; García-Campusano, Florencia; Alfaro, Isabel; Vázquez-Santana, Sonia

2013-09-01

The sexual separation in dioecious species has interested biologists for decades; however, the cellular mechanism leading to unisexuality has been poorly understood. In this study, the cellular changes that lead to male sterility in the functionally dioecious cactus, Opuntia stenopetala, are described. The spatial and temporal patterns of programmed cell death (PCD) were determined in the anthers of male and female flowers using scanning electron microscopy analysis and histological observations, focusing attention on the transition from bisexual to unisexual development. In addition, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assays were used as an indicator of DNA fragmentation to corroborate PCD. PCD was detected in anthers of both female and male flowers, but their patterns differed in time and space. Functionally male individuals developed viable pollen, and normal development involved PCD on each layer of the anther wall, which occurred progressively from the inner (tapetum) to the outer layer (epidermis). Conversely, functional female individuals aborted anthers by premature and displaced PCD. In anthers of female flowers, the first signs of PCD, such as a nucleus with irregular shape, fragmented and condensed chromatin, high vacuolization and condensed cytoplasm, occurred at the microspore mother cell stage. Later these features were observed simultaneously in all anther wall layers, connective tissue and filament. Neither pollen formation nor anther dehiscence was detected in female flowers of O. stenopetala due to total anther disruption. Temporal and spatial changes in the patterns of PCD are responsible for male sterility of female flowers in O. stenopetala. Male fertility requires the co-ordination of different events, which, when altered, can lead to male sterility and to functionally unisexual individuals. PCD could be a widespread mechanism in the determination of functionally dioecious species.

Downregulation of Metabolic Activity Increases Cell Survival Under Hypoxic Conditions: Potential Applications for Tissue Engineering

PubMed Central

Kim, Jaehyun; Andersson, Karl-Erik; Jackson, John D.; Lee, Sang Jin; Atala, Anthony

2014-01-01

A major challenge to the success of cell-based implants for tissue regeneration is an insufficient supply of oxygen before host vasculature is integrated into the implants, resulting in premature cell death and dysfunction. Whereas increasing oxygenation to the implants has been a major focus in the field, our strategy is aimed at lowering oxygen consumption by downregulating cellular metabolism of cell-based implants. Adenosine, which is a purine nucleoside that functions as an energy transferring molecule, has been reported to increase under hypoxia, resulting in reducing the adenosine triphosphate (ATP) demands of the Na+/K+ ATPase. In the present study, we investigated whether adenosine could be used to downregulate cellular metabolism to achieve prolonged survival under hypoxic conditions. Murine myoblasts (C2C12) lacking a self-survival mechanism were treated with adenosine under 0.1% hypoxic stress. The cells, cultured in the presence of 5 mM adenosine, maintained their viability under hypoxia, and regained their normal growth and function of forming myotubes when transferred to normoxic conditions at day 11 without further supply of adenosine, whereas nontreated cells failed to survive. An increase in adenosine concentrations shortened the onset of reproliferation after transfer to normoxic conditions. This increase correlated with an increase in metabolic downregulation during the early phase of hypoxia. A higher intracellular ATP level was observed in adenosine-treated cells throughout the duration of hypoxia. This strategy of increasing cell survival under hypoxic conditions through downregulating cellular metabolism may be utilized for cell-based tissue regeneration applications as well as protecting tissues against hypoxic injuries. PMID:24524875

Mutation in GNE Downregulates Peroxiredoxin IV Altering ER Redox Homeostasis.

PubMed

Chanana, Pratibha; Padhy, Gayatri; Bhargava, Kalpana; Arya, Ranjana

2017-12-01

GNE myopathy is a rare neuromuscular genetic disorder characterized by early adult onset and muscle weakness due to mutation in sialic acid biosynthetic enzyme, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). More than 180 different GNE mutations are known all over the world with unclear pathomechanism. Although hyposialylation of glycoproteins is speculated to be the major cause, but cellular mechanism leading to loss of muscle mass has not yet been deciphered. Besides sialic acid biosynthesis, GNE affects other cellular functions such as cell adhesion and apoptosis. In order to understand the effect of mutant GNE protein on cellular functions, differential proteome profile of HEK293 cells overexpressing pathologically relevant recombinant mutant GNE protein (D207V and V603L) was analyzed. These cells, along with vector control and wild-type GNE-overexpressing cells, were subjected to two-dimensional gel electrophoresis coupled with mass spectrometry (MALDI-TOF/TOF MS/MS). In the study, 10 differentially expressed proteins were identified. Progenesis same spots software revealed downregulation of peroxiredoxin IV (PrdxIV), an ER-resident H 2 O 2 sensor that regulates neurogenesis. Significant reduction in mRNA and protein levels of PrdxIV was observed in GNE mutant cell lines compared with vector control. However, neither total reactive oxygen species was altered nor H 2 O 2 accumulation was observed in GNE mutant cell lines. Interestingly, ER redox state was significantly affected due to reduced normal GNE enzyme activity. Our study indicates that downregulation of PrdxIV affects ER redox state that may contribute to misfolding and aggregation of proteins in GNE myopathy.

Increased Cysteine Biosynthesis Capacity of Transgenic Tobacco Overexpressing an O-Acetylserine(thiol) Lyase Modifies Plant Responses to Oxidative Stress1

PubMed Central

Youssefian, Shohab; Nakamura, Michimi; Orudgev, Emin; Kondo, Noriaki

2001-01-01

O-Acetylserine(thiol) lyase (OASTL), a key enzyme of plant sulfur metabolism, catalyzes the formation of Cys from sulfide and O-acetylserine. The biosynthesis of Cys is regarded as the exclusive function of sulfur reduction in plants, and a key limiting step in the production of glutathione (GSH), a thiol implicated in various cellular functions, including sulfur transport, gene expression, scavenging of reactive oxygen species (ROS), and resistance to biotic and abiotic stresses. To examine whether an increased capacity for cysteine (Cys) biosynthesis alters cellular responses to such stresses, we studied the differential changes in thiol levels and ROS scavenging of transgenic tobacco (Nicotiana tabacum) plants expressing the wheat (Triticum aestivum) OASTL gene, cys1, to SO2 and to the ROS generator, methyl viologen. Intracellular Cys and GSH contents were generally higher in cys1 transgenics than in controls under normal growth conditions, but became especially elevated in transgenic plants after SO2 exposure. An examination of differences in the ROS scavenging system of the transgenic plants also demonstrated the specific accumulation of Cu/Zn superoxide dismutase transcripts, known to be induced by Cys or GSH, and elevated cellular superoxide dismutase activities. The transgenic plants accordingly showed dramatic reductions in the extent of both foliar and photooxidative damage in response to acute SO2, as well as reduced levels of chlorosis and membrane damage following methyl viologen treatment. Overall, our results imply that OASTL plays a pivotal role in the synthesis of Cys and GSH that are required for regulation of plant responses to oxidative stress. PMID:11457951

Expression of SET Protein in the Ovaries of Patients with Polycystic Ovary Syndrome

PubMed Central

Boqun, Xu; Xiaonan, Dai; YuGui, Cui; Lingling, Gao; Xue, Dai; Gao, Chao; Feiyang, Diao; Jiayin, Liu; Gao, Li; Li, Mei; Zhang, Yuan; Ma, Xiang

2013-01-01

Background. We previously found that expression of SET gene was up-regulated in polycystic ovaries by using microarray. It suggested that SET may be an attractive candidate regulator involved in the pathophysiology of polycystic ovary syndrome (PCOS). In this study, expression and cellular localization of SET protein were investigated in human polycystic and normal ovaries. Method. Ovarian tissues, six normal ovaries and six polycystic ovaries, were collected during transsexual operation and surgical treatment with the signed consent form. The cellular localization of SET protein was observed by immunohistochemistry. The expression levels of SET protein were analyzed by Western Blot. Result. SET protein was expressed predominantly in the theca cells and oocytes of human ovarian follicles in both PCOS ovarian tissues and normal ovarian tissues. The level of SET protein expression in polycystic ovaries was triple higher than that in normal ovaries (P < 0.05). Conclusion. SET was overexpressed in polycystic ovaries more than that in normal ovaries. Combined with its localization in theca cells, SET may participate in regulating ovarian androgen biosynthesis and the pathophysiology of hyperandrogenism in PCOS. PMID:23861679

Expression of SET Protein in the Ovaries of Patients with Polycystic Ovary Syndrome.

PubMed

Boqun, Xu; Xiaonan, Dai; Yugui, Cui; Lingling, Gao; Xue, Dai; Gao, Chao; Feiyang, Diao; Jiayin, Liu; Gao, Li; Li, Mei; Zhang, Yuan; Ma, Xiang

2013-01-01

Background. We previously found that expression of SET gene was up-regulated in polycystic ovaries by using microarray. It suggested that SET may be an attractive candidate regulator involved in the pathophysiology of polycystic ovary syndrome (PCOS). In this study, expression and cellular localization of SET protein were investigated in human polycystic and normal ovaries. Method. Ovarian tissues, six normal ovaries and six polycystic ovaries, were collected during transsexual operation and surgical treatment with the signed consent form. The cellular localization of SET protein was observed by immunohistochemistry. The expression levels of SET protein were analyzed by Western Blot. Result. SET protein was expressed predominantly in the theca cells and oocytes of human ovarian follicles in both PCOS ovarian tissues and normal ovarian tissues. The level of SET protein expression in polycystic ovaries was triple higher than that in normal ovaries (P < 0.05). Conclusion. SET was overexpressed in polycystic ovaries more than that in normal ovaries. Combined with its localization in theca cells, SET may participate in regulating ovarian androgen biosynthesis and the pathophysiology of hyperandrogenism in PCOS.

Acute Fasting Regulates Retrograde Synaptic Enhancement through a 4E-BP-Dependent Mechanism.

PubMed

Kauwe, Grant; Tsurudome, Kazuya; Penney, Jay; Mori, Megumi; Gray, Lindsay; Calderon, Mario R; Elazouzzi, Fatima; Chicoine, Nicole; Sonenberg, Nahum; Haghighi, A Pejmun

2016-12-21

While beneficial effects of fasting on organismal function and health are well appreciated, we know little about the molecular details of how fasting influences synaptic function and plasticity. Our genetic and electrophysiological experiments demonstrate that acute fasting blocks retrograde synaptic enhancement that is normally triggered as a result of reduction in postsynaptic receptor function at the Drosophila larval neuromuscular junction (NMJ). This negative regulation critically depends on transcriptional enhancement of eukaryotic initiation factor 4E binding protein (4E-BP) under the control of the transcription factor Forkhead box O (Foxo). Furthermore, our findings indicate that postsynaptic 4E-BP exerts a constitutive negative input, which is counteracted by a positive regulatory input from the Target of Rapamycin (TOR). This combinatorial retrograde signaling plays a key role in regulating synaptic strength. Our results provide a mechanistic insight into how cellular stress and nutritional scarcity could acutely influence synaptic homeostasis and functional stability in neural circuits. Copyright © 2016 Elsevier Inc. All rights reserved.

Biological pathways involved in the development of inflammatory bowel disease.

PubMed

Zemljic, Mateja; Pejkovic, Bozena; Krajnc, Ivan; Lipovsek, Saska

2014-10-01

Apoptosis, autophagy and necrosis are three distinct functional types of the mammalian cell death network. All of them are characterized by a number of cell's morphological changes. The inappropriate induction of cell death is involved in the pathogenesis of a number of diseases.Pathogenesis of inflammatory bowel diseases (ulcerative colitis, Crohn's disease) includes an abnormal immunological response to disturbed intestinal microflora. One of the most important reason in pathogenesis of chronic inflammatory disease and subsequent multiple organ pathology is a barrier function of the gut, regulating cellular viability. Recent findings have begun to explain the mechanisms by which intestinal epithelial cells are able to survive in such an environment and how loss of normal regulatory processes may lead to inflammatory bowel disease (IBD).This review focuses on the regulation of biological pathways in development and homeostasis in IBD. Better understanding of the physiological functions of biological pathways and their influence on inflammation, immunity, and barrier function will simplify our expertice of homeostasis in the gastrointestinal tract and in upgrading diagnosis and treatment.

Novel riboflavin transporter family RFVT/SLC52: identification, nomenclature, functional characterization and genetic diseases of RFVT/SLC52.

PubMed

Yonezawa, Atsushi; Inui, Ken-ichi

2013-01-01

Riboflavin, a water-soluble vitamin also known as vitamin B2, is essential for normal cellular functions. Riboflavin transporters play important roles in its homeostasis. Recently, three novel riboflavin transporters were identified, and designated as RFT1, RFT2 and RFT3. Because the RFTs did not show similarity to other SLC transporters, and RFT1 and RFT3 are similar in sequence and function, they were assigned into a new SLC family, SLC52. Subsequently, RFT1/GPR172B, RFT3/GPR172A and RFT2/C20orf54 were renamed as RFVT1/SLC52A1, RFVT2/SLC52A2 and RFVT3/SLC52A3, respectively. In this review, we summarize recent findings on the cloning, nomenclature, functional characterization and genetic diseases of RFVT1/SLC52A1, RFVT2/SLC52A2 and RFVT3/SLC52A3. Copyright © 2012 Elsevier Ltd. All rights reserved.

Emerging concepts in smooth muscle contributions to airway structure and function: implications for health and disease

PubMed Central

2016-01-01

Airway structure and function are key aspects of normal lung development, growth, and aging, as well as of lung responses to the environment and the pathophysiology of important diseases such as asthma, chronic obstructive pulmonary disease, and fibrosis. In this regard, the contributions of airway smooth muscle (ASM) are both functional, in the context of airway contractility and relaxation, as well as synthetic, involving production and modulation of extracellular components, modulation of the local immune environment, cellular contribution to airway structure, and, finally, interactions with other airway cell types such as epithelium, fibroblasts, and nerves. These ASM contributions are now found to be critical in airway hyperresponsiveness and remodeling that occur in lung diseases. This review emphasizes established and recent discoveries that underline the central role of ASM and sets the stage for future research toward understanding how ASM plays a central role by being both upstream and downstream in the many interactive processes that determine airway structure and function in health and disease. PMID:27742732

In Situ Patrolling of Regulatory T Cells Is Essential for Protecting Autoimmune Exocrinopathy

PubMed Central

Ishimaru, Naozumi; Nitta, Takeshi; Arakaki, Rieko; Yamada, Akiko; Lipp, Martin; Takahama, Yousuke; Hayashi, Yoshio

2010-01-01

Background Migration of T cells, including regulatory T (Treg) cells, into the secondary lymph organs is critically controlled by chemokines and adhesion molecules. However, the mechanisms by which Treg cells regulate organ-specific autoimmunity via these molecules remain unclear. Although we previously reported autoimmune exocrinopathy resembling Sjögren's syndrome (SS) in the lacrimal and salivary glands from C-C chemokine receptor 7 (CCR7)-deficient mice, it is still unclear whether CCR7 signaling might specifically affect the dynamics and functions of Treg cells in vivo. We therefore investigated the cellular mechanism for suppressive function of Treg cells via CCR7 in autoimmunity using mouse models and human samples. Methods and Findings Patrolling Treg cells were detected in the exocrine organs such as lacrimal and salivary glands from normal mice that tend to be targets for autoimmunity while the Treg cells were almost undetectable in the exocrine glands of CCR7 −/− mice. In addition, we found the significantly increased retention of CD4+CD25+Foxp3+ Treg cells in the lymph nodes of CCR7 −/− mice with aging. Although Treg cell egress requires sphingosine 1-phosphate (S1P), chemotactic function to S1P of CCR7−/− Treg cells was impaired compared with that of WT Treg cells. Moreover, the in vivo suppression activity was remarkably diminished in CCR7 −/− Treg cells in the model where Treg cells were co-transferred with CCR7 −/− CD25-CD4+ T cells into Rag2 −/− mice. Finally, confocal analysis showed that CCR7+Treg cells were detectable in normal salivary glands while the number of CCR7+Treg cells was extremely decreased in the tissues from patients with Sjögren's syndrome. Conclusions These results indicate that CCR7 essentially governs the patrolling functions of Treg cells by controlling the traffic to the exocrine organs for protecting autoimmunity. Characterization of this cellular mechanism could have clinical implications by supporting development of new diagnosis or treatments for the organ-specific autoimmune diseases such as Sjögren's syndrome and clarifying how the local immune system regulates autoimmunity. PMID:20052419

Metabostemness: a new cancer hallmark.

PubMed

Menendez, Javier A; Alarcón, Tomás

2014-01-01

The acquisition of and departure from stemness in cancer tissues might not only be hardwired by genetic controllers, but also by the pivotal regulatory role of the cellular metabotype, which may act as a "starter dough" for cancer stemness traits. We have coined the term metabostemness to refer to the metabolic parameters causally controlling or functionally substituting the epitranscriptional orchestration of the genetic reprograming that redirects normal and tumor cells toward less-differentiated cancer stem cell (CSC) cellular states. Certain metabotypic alterations might operate as pivotal molecular events rendering a cell of origin susceptible to epigenetic rewiring required for the acquisition of aberrant stemness and, concurrently, of refractoriness to differentiation. The metabostemness attribute can remove, diminish, or modify the nature of molecular barriers present in Waddington's epigenetic landscapes, thus allowing differentiated cells to more easily (re)-enter into CSC cellular macrostates. Activation of the metabostemness trait can poise cells with chromatin states competent for rapid dedifferentiation while concomitantly setting the idoneous metabolic stage for later reprograming stimuli to finish the journey from non-cancerous into tumor-initiating cells. Because only a few permitted metabotypes will be compatible with the operational properties owned by CSC cellular states, the metabostemness property provides a new framework through which to pharmacologically resolve the apparently impossible problem of discovering drugs aimed to target the molecular biology of the cancer stemness itself. The metabostemness cancer hallmark generates a shifting oncology theory that should guide a new era of metabolo-epigenetic cancer precision medicine.

Metabostemness: A New Cancer Hallmark

PubMed Central

Menendez, Javier A.; Alarcón, Tomás

2014-01-01

The acquisition of and departure from stemness in cancer tissues might not only be hardwired by genetic controllers, but also by the pivotal regulatory role of the cellular metabotype, which may act as a “starter dough” for cancer stemness traits. We have coined the term metabostemness to refer to the metabolic parameters causally controlling or functionally substituting the epitranscriptional orchestration of the genetic reprograming that redirects normal and tumor cells toward less-differentiated cancer stem cell (CSC) cellular states. Certain metabotypic alterations might operate as pivotal molecular events rendering a cell of origin susceptible to epigenetic rewiring required for the acquisition of aberrant stemness and, concurrently, of refractoriness to differentiation. The metabostemness attribute can remove, diminish, or modify the nature of molecular barriers present in Waddington’s epigenetic landscapes, thus allowing differentiated cells to more easily (re)-enter into CSC cellular macrostates. Activation of the metabostemness trait can poise cells with chromatin states competent for rapid dedifferentiation while concomitantly setting the idoneous metabolic stage for later reprograming stimuli to finish the journey from non-cancerous into tumor-initiating cells. Because only a few permitted metabotypes will be compatible with the operational properties owned by CSC cellular states, the metabostemness property provides a new framework through which to pharmacologically resolve the apparently impossible problem of discovering drugs aimed to target the molecular biology of the cancer stemness itself. The metabostemness cancer hallmark generates a shifting oncology theory that should guide a new era of metabolo-epigenetic cancer precision medicine. PMID:25325014

Copper toxicity, oxidative stress, and antioxidant nutrients.

PubMed

Gaetke, Lisa M; Chow, Ching Kuang

2003-07-15

Copper (Cu) is an integral part of many important enzymes involved in a number of vital biological processes. Although normally bound to proteins, Cu may be released and become free to catalyze the formation of highly reactive hydroxyl radicals. Data obtained from in vitro and cell culture studies are largely supportive of Cu's capacity to initiate oxidative damage and interfere with important cellular events. Oxidative damage has been linked to chronic Cu-overload and/or exposure to excess Cu caused by accidents, occupational hazards, and environmental contamination. Additionally, Cu-induced oxidative damage has been implicated in disorders associated with abnormal Cu metabolism and neurodegenerative changes. Interestingly, a deficiency in dietary Cu also increases cellular susceptibility to oxidative damage. A number of nutrients have been shown to interact with Cu and alter its cellular effects. Vitamin E is generally protective against Cu-induced oxidative damage. While most in vitro or cell culture studies show that ascorbic acid aggravates Cu-induced oxidative damage, results obtained from available animal studies suggest that the compound is protective. High intakes of ascorbic acid and zinc may provide protection against Cu toxicity by preventing excess Cu uptake. Zinc also removes Cu from its binding site, where it may cause free radical formation. Beta-carotene, alpha-lipoic acid and polyphenols have also been shown to attenuate Cu-induced oxidative damage. Further studies are needed to better understand the cellular effects of this essential, but potentially toxic, trace mineral and its functional interaction with other nutrients.

Profiling cellular bioenergetics, glutathione levels, and caspase activities in stomach biopsies of patients with upper gastrointestinal symptoms

PubMed Central

Alfazari, Ali S; Al-Dabbagh, Bayan; Al-Dhaheri, Wafa; Taha, Mazen S; Chebli, Ahmad A; Fontagnier, Eva M; Koutoubi, Zaher; Kochiyi, Jose; Karam, Sherif M; Souid, Abdul-Kader

2015-01-01

AIM: To measure biochemical parameters in stomach biopsies and test their suitability as diagnostic biomarkers for gastritis and precancerous lesions. METHODS: Biopsies were obtained from the stomachs of two groups of patients (n = 40) undergoing fiber-optic endoscopy due to upper gastrointestinal symptoms. In the first group (n = 17), only the corpus region was examined. Biopsies were processed for microscopic examination and measurement of mitochondrial O2 consumption (cellular respiration), cellular adenosine triphosphate (ATP), glutathione (GSH), and caspase activity. In the second group of patients (n = 23), both corpus and antral regions were studied. Some biopsies were processed for microscopic examination, while the others were used for measurements of cellular respiration and GSH level. RESULTS: Microscopic examinations of gastric corpus biopsies from 17 patients revealed normal mucosae in 8 patients, superficial gastritis in 7 patients, and chronic atrophic gastritis in 1 patient. In patients with normal histology, the rate (mean ± SD) of cellular respiration was 0.17 ± 0.02 μmol/L O2 min-1 mg-1, ATP content was 487 ± 493 pmol/mg, and GSH was 469 ± 98 pmol/mg. Caspase activity was detected in 3 out of 8 specimens. The values of ATP and caspase activity were highly variable. The presence of superficial gastritis had insignificant effects on the measured biomarkers. In the patient with atrophic gastritis, cellular respiration was high and ATP was relatively low, suggesting uncoupling oxidative phosphorylation. In the second cohort of patients, the examined biopsies showed either normal or superficial gastritis. The rate of cellular respiration (O2. μmol/L min-1 mg-1) was slightly higher in the corpus than the antrum (0.18 ± 0.05 vs 0.15 ± 0.04, P = 0.019). The value of GSH was about the same in both tissues (310 ± 135 vs 322 ± 155, P = 0.692). CONCLUSION: The corpus mucosa was metabolically more active than the antrum tissue. The data in this study will help in understanding the pathophysiology of gastric mucosa. PMID:25593494

Prolonged Delirium Secondary to Hypoxic-ischemic Encephalopathy Following Cardiac Arrest

PubMed Central

Yogaratnam, Jegan; Jacob, Rajesh; Naik, Sandeep; Magadi, Harish

2013-01-01

Hypoxic-ischemic brain injury encompasses a complex constellation of pathophysiological and cellular brain injury induced by hypoxia, ischemia, cytotoxicity, or combinations of these mechanisms and can result in poor outcomes including significant changes in personality and cognitive impairments in memory, cognition, and attention. We report a case of a male patient with normal premorbid functioning who developed prolonged delirium following hypoxic-ischemic brain insults subsequent to cardiac arrest. The case highlights the importance of adopting a multidisciplinary treatment approach involving the coordinated care of medical and nursing teams to optimise management of patients suffering from such a debilitating organic brain syndrome. PMID:23678354

Drug targeting of oncogenic pathways in melanoma.

PubMed

Fecher, Leslie A; Amaravadi, Ravi K; Schuchter, Lynn M; Flaherty, Keith T

2009-06-01

Melanoma continues to be one of the most aggressive and morbid malignancies once metastatic. Overall survival for advanced unresectable melanoma has not changed over the past several decades. However, the presence of some long-term survivors of metastatic melanoma highlights the heterogeneity of this disease and the potential for improved outcomes. Current research is uncovering the molecular and genetic scaffolding of normal and aberrant cell function. The known oncogenic pathways in melanoma and the attempts to develop therapy for them are discussed. The targeting of certain cellular processes, downstream of the common genetic alterations, for which the issues of target and drug validation are somewhat distinct, are also highlighted.

Nonlinear cellular dynamics of keratinocytes in normal and psoriatic epidermis under action of UV radiation

NASA Astrophysics Data System (ADS)

Stolnitz, Mikhail M.; Medvedev, Boris A.; Gribko, Tatyana V.

2004-05-01

The semi-phenomenological model of epidermal cell dynamics is submitted. The model takes into account three types of basal layer keratinocytes (stem, transient amplifying, terminally differentiated), distribution of first two types cells on mitotic cycle stages and resting states, keratinocytes-lymphocytes interactions that provide a positive feedback loop, influence of more differentiated cells on their progenitors that provide a negative feedback loop. Simplified model are developed and its stationary solutions are received. The opportunity of interpretation of some received modes as corresponding to various stages of psoriasis is discussed. Influence of UV-radiation on transitions between various modes of epidermis functioning is qualitatively analyzed.

Adult mesenchymal stem cells and cell-based tissue engineering

PubMed Central

Tuan, Rocky S; Boland, Genevieve; Tuli, Richard

2003-01-01

The identification of multipotential mesenchymal stem cells (MSCs) derived from adult human tissues, including bone marrow stroma and a number of connective tissues, has provided exciting prospects for cell-based tissue engineering and regeneration. This review focuses on the biology of MSCs, including their differentiation potentials in vitro and in vivo, and the application of MSCs in tissue engineering. Our current understanding of MSCs lags behind that of other stem cell types, such as hematopoietic stem cells. Future research should aim to define the cellular and molecular fingerprints of MSCs and elucidate their endogenous role(s) in normal and abnormal tissue functions. PMID:12716446

Endocannabinoid signalling and the deteriorating brain

PubMed Central

Di Marzo, Vincenzo; Stella, Nephi; Zimmer, Andreas

2015-01-01

Ageing is characterized by the progressive impairment of physiological functions and increased risk of developing debilitating disorders, including chronic inflammation and neurodegenerative diseases. These disorders have common molecular mechanisms that can be targeted therapeutically. In the wake of the approval of the first cannabinoid-based drug for the symptomatic treatment of multiple sclerosis, we examine how endocannabinoid (eCB) signalling controls — and is affected by — normal ageing and neuroinflammatory and neurodegenerative disorders. We propose a conceptual framework linking eCB signalling to the control of the cellular and molecular hallmarks of these processes, and categorize the key components of endocannabinoid signalling that may serve as targets for novel therapeutics. PMID:25524120

Neuroimmune regulation of neurophysiology in the cerebellum.

PubMed

Gruol, Donna L

2013-06-01

Recent studies have established the existence of an innate immune system in the central nervous system (CNS) and implicated a critical role for this system in both normal and pathological processes. Astrocytes and microglia, normal components of the CNS, are the primary cell types that comprise the innate immune system of the CNS. Basic to their role during normal and adverse conditions is the production of neuroimmune factors such as cytokines and chemokines, which are signaling molecules that initiate or coordinate downstream cellular actions. During adverse conditions, cytokines and chemokines function in defensive and repair. However, if expression of these factors becomes dysregulated, abnormal CNS function can result. Both neurons and glial cells of the CNS express receptors for cytokines and chemokines, but the biological consequence of receptor activation has yet to be fully resolved. Our studies show that neuroadaptive changes are produced in primary cultures of rat cerebellar cells chronically treated with the cytokine interleukin-6 (IL-6) and in the cerebellum of transgenic mice that chronically express elevated levels of IL-6 in the CNS. In the cerebellum in culture and in vivo, the neuroadaptive changes included alterations in the level of expression of proteins involved in gene expression, signal transduction, and synaptic transmission. Associated with these changes were alterations in neuronal function. A comparison of results from the cultured cerebellar cells and cerebellum of the transgenic mice indicated that the effects of IL-6 can vary across neuronal types. However, alterations in mechanisms involved in Ca(2+) homeostasis were observed in all cell types studied. These results indicate that modifications in cerebellar function are likely to occur in disorders associated with elevated levels of IL-6 in the cerebellum.

Impaired cardiac contractile function in arginine:glycine amidinotransferase knockout mice devoid of creatine is rescued by homoarginine but not creatine

PubMed Central

Faller, Kiterie M E; Atzler, Dorothee; McAndrew, Debra J; Zervou, Sevasti; Whittington, Hannah J; Simon, Jillian N; Aksentijevic, Dunja; ten Hove, Michiel; Choe, Chi-un; Isbrandt, Dirk; Casadei, Barbara; Schneider, Jurgen E; Neubauer, Stefan; Lygate, Craig A

2018-01-01

Abstract Aims Creatine buffers cellular adenosine triphosphate (ATP) via the creatine kinase reaction. Creatine levels are reduced in heart failure, but their contribution to pathophysiology is unclear. Arginine:glycine amidinotransferase (AGAT) in the kidney catalyses both the first step in creatine biosynthesis as well as homoarginine (HA) synthesis. AGAT-/- mice fed a creatine-free diet have a whole body creatine-deficiency. We hypothesized that AGAT-/- mice would develop cardiac dysfunction and rescue by dietary creatine would imply causality. Methods and results Withdrawal of dietary creatine in AGAT-/- mice provided an estimate of myocardial creatine efflux of ∼2.7%/day; however, in vivo cardiac function was maintained despite low levels of myocardial creatine. Using AGAT-/- mice naïve to dietary creatine we confirmed absence of phosphocreatine in the heart, but crucially, ATP levels were unchanged. Potential compensatory adaptations were absent, AMPK was not activated and respiration in isolated mitochondria was normal. AGAT-/- mice had rescuable changes in body water and organ weights suggesting a role for creatine as a compatible osmolyte. Creatine-naïve AGAT-/- mice had haemodynamic impairment with low LV systolic pressure and reduced inotropy, lusitropy, and contractile reserve. Creatine supplementation only corrected systolic pressure despite normalization of myocardial creatine. AGAT-/- mice had low plasma HA and supplementation completely rescued all other haemodynamic parameters. Contractile dysfunction in AGAT-/- was confirmed in Langendorff perfused hearts and in creatine-replete isolated cardiomyocytes, indicating that HA is necessary for normal cardiac function. Conclusions Our findings argue against low myocardial creatine per se as a major contributor to cardiac dysfunction. Conversely, we show that HA deficiency can impair cardiac function, which may explain why low HA is an independent risk factor for multiple cardiovascular diseases. PMID:29236952