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Sample records for cell enolase level

  1. Normalization of Red Cell Enolase Level Following Allogeneic Bone Marrow Transplantation in a Child with Diamond-Blackfan Anemia

    OpenAIRE

    Park, Jeong A; Lim, Yeon Jung; Park, Hyeon Jin; Kong, Sun Young; Park, Byung Kiu; Ghim, Thad T.

    2010-01-01

    We describe a girl with Diamond-Blackfan anemia with accompanying red cell enolase deficiency. At the age of 9 yr old, the patient received allogeneic bone marrow transplantation from her HLA-identical sister who had normal red cell enolase activity. While the post transplant DNA analysis with short tandem repeat has continuously demonstrated a stable mixed chimerism on follow-up, the patient remains transfusion independent and continues to show a steady increase in red cell enolase activity ...

  2. Modulation of α-enolase post-translational modifications by dengue virus: increased secretion of the basic isoforms in infected hepatic cells.

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    Luiza M Higa

    Full Text Available Hepatic cells are major sites of dengue virus (DENV replication and liver injury constitutes a characteristic of severe forms of dengue. The role of hepatic cells in dengue pathogenesis is not well established, but since hepatocytes are the major source of plasma proteins, changes in protein secretion by these cells during infection might contribute to disease progression. Previously, we showed that DENV infection alters the secretion pattern of hepatic HepG2 cells, with α-enolase appearing as one of the major proteins secreted in higher levels by infected cells. ELISA analysis demonstrated that DENV infection modulates α-enolase secretion in HepG2 cells in a dose-dependent manner, but has no effect on its gene expression and on the intracellular content of the protein as assessed by PCR and western blot analyses, respectively. Two-dimensional western blots showed that both intracellular and secreted forms of α-enolase appear as five spots, revealing α-enolase isoforms with similar molecular weights but distinct isoeletric points. Remarkably, quantification of each spot content revealed that DENV infection shifts the isoform distribution pattern of secreted α-enolase towards the basic isoforms, whereas the intracellular protein remains unaltered, suggesting that post-translational modifications might be involved in α-enolase secretion by infected cells. These findings provide new insights into the mechanisms underlying α-enolase secretion by hepatic cells and its relationship with the role of liver in dengue pathogenesis. In addition, preliminary results obtained with plasma samples from DENV-infected patients suggest an association between plasma levels of α-enolase and disease severity. Since α-enolase binds plasminogen and modulates its activation, it is plausible to speculate the association of the increase in α-enolase secretion by infected hepatic cells with the haemostatic dysfunction observed in dengue patients including the

  3. Wheat enolase demonstrates potential as a non-toxic cryopreservation agent for liver and pancreatic cells.

    Science.gov (United States)

    Grondin, Mélanie; Chow-Shi-Yée, Mélanie; Ouellet, François; Averill-Bates, Diana A

    2015-05-01

    Cryopreservation is essential for long-term storage of cells and tissues, which can be used for clinical applications such as drug toxicity testing, human transplantation, reproductive, regenerative and transfusion medicine. It requires use of cryoprotectants (e.g. dimethyl disulfoxide (DMSO), glycerol) that protect cells and tissues from dehydration and damage caused by formation of intracellular ice during freezing. As an alternative to these cytotoxic cryoprotectants, we are developing new technology using natural substances produced by plants that survive freezing conditions. We previously showed that soluble protein extracts such as wheat protein extract (WPE) prepared from winter wheat plants can substitute for DMSO as a cryoprotectant for certain mammalian cell types. To identify novel cryoactive proteins, WPE was separated using different chromatographic procedures and cryoactive fractions were analyzed by mass spectrometry. The analysis revealed enolase as a potential wheat protein candidate. A recombinant enolase protein was prepared and was able to successfully cryopreserve rat hepatocytes and insulin-secreting INS832/13 pancreatic cells. Post-thaw cells had high viability and levels of metabolic activities. Cryopreserved cells were plateable and had good adherence and morphological properties. These results indicate that individual plant proteins such as enolase have promising potential as new, non-toxic technology for cryopreservation protocols used for clinical applications. PMID:25740431

  4. Genetic organization and mRNA expression of enolase genes of Candida albicans.

    Science.gov (United States)

    Postlethwait, P; Sundstrom, P

    1995-04-01

    In previous work, we cloned a Candida albicans cDNA for the glycolytic enzyme enolase and found a single, abundant enolase transcript on Northern (RNA) blots and a single protein on immunoblots, using antiserum raised against a recombinant enolase fusion protein. Because C. albicans enolase is abundantly produced during infection and elicits strong host immune responses, the mechanisms regulating enolase production are important for understanding the growth of C. albicans in vivo. To obtain more information on enolase gene expression by C. albicans, we used the enolase cDNA clone to investigate the genetic organization of enolase genes and the steady-state levels of enolase mRNA under several growth conditions. Gene disruption techniques in combination with Southern blot analyses of genomic DNA showed the presence of two enolase gene loci that could be distinguished by the locations of ClaI and Mn/I sites in their 3' flanking regions. Enolase steady-state mRNA levels were greatest during the middle phase of the logarithmic growth curve and were low during stationary phase. Minimal differences in enolase mRNA levels between yeast cells and hyphae were found. Propagation of C. albicans in glucose did not cause increased enolase mRNA levels compared with growth in a nonfermentable carbon source (pyruvate). It was concluded that two gene loci exist for C. albicans enolase and that enolase mRNA is constitutively produced at high levels during active metabolism. PMID:7896700

  5. EGCG decreases binding of calcium oxalate monohydrate crystals onto renal tubular cells via decreased surface expression of alpha-enolase.

    Science.gov (United States)

    Kanlaya, Rattiyaporn; Singhto, Nilubon; Thongboonkerd, Visith

    2016-06-01

    Crystal retention on tubular cell surface inside renal tubules is considered as the earliest and crucial step for kidney stone formation. Therapeutics targeting this step would cease the development of kidney stone. This study thus aimed to investigate the potential role of epigallocatechin-3-gallate (EGCG), a major antioxidant found in green tea leaves, in the reduction of calcium oxalate monohydrate (COM) crystal binding onto renal tubular cells. Pretreatment of the cells with EGCG for up to 6 h significantly diminished crystal-binding capability in a dose-dependent manner. Indirect immunofluorescence assay without and with cell permeabilization followed by laser-scanning confocal microscopy revealed that EGCG significantly reduced surface expression of alpha-enolase, whereas its intracellular level was increased. Western blot analysis confirmed such contradictory changes in membrane and cytosolic fractions of EGCG-treated cells, whereas the total level in whole cell lysate remained unchanged. Moreover, overexpression of surface alpha-enolase and enhancement of cell-crystal adhesion induced by 10 mM sodium oxalate were completely abolished by EGCG. Taken together, these data indicate that EGCG decreases binding of COM crystals onto renal tubular cells by decreasing the surface expression of alpha-enolase via re-localization or inhibition of alpha-enolase shuttling from the cytoplasm to the plasma membrane. These findings may also explain the effects of EGCG in reducing COM crystal deposition in previous animal models of kidney stone disease. Thus, EGCG may be useful for the prevention of new or recurrent stone formation. PMID:26898643

  6. Correlative study between neuron-specific enolase and blood sugar level in ischemic stroke patients

    OpenAIRE

    Aparna Pandey; Kiran Saxena; Meena Verma; Anuradha Bharosay

    2011-01-01

    Background: A study to investigate the level of the neurobiochemical marker, Neuron-Specific Enolase (NSE), at the time of admission and its correlation with the blood sugar level in ischemic stroke patients. Patients and Methods: We investigated 90 patients with complete stroke who were admitted to the Stroke Unit of the Department of Neurology at Sri Aurobindo Institute of Medical Sciences. NSE was measured with commercially available quantitative ‘sandwich’ enzyme-linked immunosorbent assa...

  7. Genetic and proteomic evidences support the localization of yeast enolase in the cell surface

    DEFF Research Database (Denmark)

    López-Villar, Elena; Monteoliva, Lucía; Larsen, Martin Røssel;

    2006-01-01

    Although enolase, other glycolytic enzymes, and a variety of cytoplasmic proteins lacking an N-terminal secretion signal have been widely described as located at the cell surface in yeast and in mammalian cells, their presence in this external location is still controversial. Here, we report that...

  8. Correlative study between neuron-specific enolase and blood sugar level in ischemic stroke patients

    Directory of Open Access Journals (Sweden)

    Aparna Pandey

    2011-01-01

    Full Text Available Background: A study to investigate the level of the neurobiochemical marker, Neuron-Specific Enolase (NSE, at the time of admission and its correlation with the blood sugar level in ischemic stroke patients. Patients and Methods: We investigated 90 patients with complete stroke who were admitted to the Stroke Unit of the Department of Neurology at Sri Aurobindo Institute of Medical Sciences. NSE was measured with commercially available quantitative ′sandwich′ enzyme-linked immunosorbent assay kits obtained from R and D Systems. Hyperglycemia was defined as blood glucose concentration ≥ 7 mmol / L, and measured using the glucose oxidase method immediately. Results: Significantly increased NSE and lipid profile levels were found in ischemic stroke patients as compared to the control. Hyperglycemic ischemic stroke patients had increased levels of NSE, lipid profile, and National Institute of Health stroke scale scores (NIHSS score compared to normoglycemic ischemic stroke patients. In addition the serum NSE level of hyperglycemic stroke patients was also positively correlated with the blood sugar level (r = 0.734 P < 0.001. Conclusions: Hyperglycemia predicts an increased risk of poor outcome after ischemic stroke and it is reflected by a significantly increased level of Neuron-Specific Enolase.

  9. Hypoxia induces differential translation of enolase/MBP-1

    International Nuclear Information System (INIS)

    Hypoxic microenvironments in tumors contribute to transformation, which may alter metabolism, growth, and therapeutic responsiveness. The α-enolase gene encodes both a glycolytic enzyme (α-enolase) and a DNA-binding tumor suppressor protein, c-myc binding protein (MBP-1). These divergent α-enolase gene products play central roles in glucose metabolism and growth regulation and their differential regulation may be critical for tumor adaptation to hypoxia. We have previously shown that MBP-1 and its binding to the c-myc P2 promoter regulates the metabolic and cellular growth changes that occur in response to altered exogenous glucose concentrations. To examine the regulation of α-enolase and MBP-1 by a hypoxic microenvironment in breast cancer, MCF-7 cells were grown in low, physiologic, or high glucose under 1% oxygen. Our results demonstrate that adaptation to hypoxia involves attenuation of MBP-1 translation and loss of MBP-1-mediated regulation of c-myc transcription, evidenced by decreased MBP-1 binding to the c-myc P2 promoter. This allows for a robust increase in c-myc expression, 'early c-myc response', which stimulates aerobic glycolysis resulting in tumor acclimation to oxidative stress. Increased α-enolase mRNA and preferential translation/post-translational modification may also allow for acclimatization to low oxygen, particularly under low glucose concentrations. These results demonstrate that malignant cells adapt to hypoxia by modulating α-enolase/MBP-1 levels and suggest a mechanism for tumor cell induction of the hyperglycolytic state. This important 'feedback' mechanism may help transformed cells to escape the apoptotic cascade, allowing for survival during limited glucose and oxygen availability

  10. Enolase is regulated by Liver X Receptors.

    Science.gov (United States)

    De Boussac, Hugues; Maqdasy, Salwan; Trousson, Amalia; Zelcer, Noam; Volle, David H; Lobaccaro, Jean-Marc A; Baron, Silvère

    2015-07-01

    Enolase is a glycolytic enzyme known to inhibit cholesteryl ester hydrolases (CEHs). Cholesteryl ester loading of macrophages, as occurs during atherosclerosis, is accompanied by increased Enolase protein and activity. Here, we describe that J774 macrophages treated with LXR agonists exhibit reduced Enolase transcript and protein abundance. Moreover, we show that this reduction is further potentiated by activation of the LXR/RXR heterodimer with the RXR ligand 9-cis retinoic acid. Enolase levels are also reduced in vivo following activation of LXRs in the intestine, but not in the liver. This effect is lost in Lxrαβ-/- mice. In aggregate, our study identified Enolase as a new target of LXRs in vivo, which may promote cholesterol mobilization for subsequent efflux. PMID:25708389

  11. Early detection of response in small cell bronchogenic carcinoma by changes in serum concentrations of creatine kinase, neuron specific enolase, calcitonin, ACTH, serotonin and gastrin releasing peptide

    DEFF Research Database (Denmark)

    Bork, E; Hansen, M; Urdal, P;

    1988-01-01

    Creatine kinase (CK-BB), neuron specific enolase (NSE), ACTH, calcitonin, serotonin and gastrin releasing peptide (GRP) were measured in serum or plasma before and immediately after initiation of treatment in patients with small cell lung cancer (SCC). Pretherapeutic elevated concentrations of CK...

  12. Research of the serum level of neuron-specific enolase in children with various types of seizure

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    WANG Chun

    2012-10-01

    Full Text Available Objective To explore the relevance between the level changes of serum neuron-specific enolase (NSE and neuronal damage in various seizure types of children with epilepsy. Methods According to the classification criteria of seizure types formulated by International League Against Epilepsy (ILAE in 1981, 190 children with epilepsy were enrolled including tonic-clonic seizure group (41 cases, tonic seizure group (34 cases, clonic seizure group (22 cases, myoclonic seizure group (12 cases, atonic seizure group (17 cases, absence seizure group (22 cases, simple partial seizure group (21 cases and complex partial seizure group (21 cases, and 64 healthy children were enrolled as control group. The long-range vedio-electroencephalogram (VEEG was operated and the blood samples were collected from these cases within 72 h after their seizures. Results The serum NSE levels of epileptic children were significantly higher than control group (P = 0.000. Among these seizure groups, serum NSE in myoclonic seizure group [(32.42 ± 6.62 ng/ml] was significantly higher than the other types, except for tonic-clonic seizure group (P = 0.062. There was no significant difference among the other types (P > 0.05, for all. According to rank correlation analysis, there was positive corrlation between serum NSE levels and VEEG abnormal intensity (rs = 0.613, P = 0.000. Conclusion The serum NSE were markedly increased in children with epilepsy after seizures, suggesting that a certain degree of neuronal damage may result from seizures; the higher NSE levels were, the more serious neuronal damage caused by epileptiform discharges was. The serum NSE levels in myoclonic seizure group and tonic-clonic seizure group were significantly higher than other seizure types, indicating the two kinds of seizures may result in greater neuronal damage.

  13. Enolase 1 (ENO1 and protein disulfide-isomerase associated 3 (PDIA3 regulate Wnt/β-catenin-driven trans-differentiation of murine alveolar epithelial cells

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    Kathrin Mutze

    2015-08-01

    Full Text Available The alveolar epithelium represents a major site of tissue destruction during lung injury. It consists of alveolar epithelial type I (ATI and type II (ATII cells. ATII cells are capable of self-renewal and exert progenitor function for ATI cells upon alveolar epithelial injury. Cell differentiation pathways enabling this plasticity and allowing for proper repair, however, are poorly understood. Here, we applied proteomics, expression analysis and functional studies in primary murine ATII cells to identify proteins and molecular mechanisms involved in alveolar epithelial plasticity. Mass spectrometry of cultured ATII cells revealed a reduction of carbonyl reductase 2 (CBR2 and an increase in enolase 1 (ENO1 and protein disulfide-isomerase associated 3 (PDIA3 protein expression during ATII-to-ATI cell trans-differentiation. This was accompanied by increased Wnt/β-catenin signaling, as analyzed by qRT-PCR and immunoblotting. Notably, ENO1 and PDIA3, along with T1α (podoplanin; an ATI cell marker, exhibited decreased protein expression upon pharmacological and molecular Wnt/β-catenin inhibition in cultured ATII cells, whereas CBR2 levels were stabilized. Moreover, we analyzed primary ATII cells from mice with bleomycin-induced lung injury, a model exhibiting activated Wnt/β-catenin signaling in vivo. We observed reduced CBR2 significantly correlating with surfactant protein C (SFTPC, whereas ENO1 and PDIA3 along with T1α were increased in injured ATII cells. Finally, siRNA-mediated knockdown of ENO1, as well as PDIA3, in primary ATII cells led to reduced T1α expression, indicating diminished cell trans-differentiation. Our data thus identified proteins involved in ATII-to-ATI cell trans-differentiation and suggest a Wnt/β-catenin-driven functional role of ENO1 and PDIA3 in alveolar epithelial cell plasticity in lung injury and repair.

  14. Immunoreactive neuron-specific enolase (NSE) is expressed in testicular carcinoma-in-situ

    DEFF Research Database (Denmark)

    Kang, J L; Rajpert-De Meyts, E; Skakkebaek, N E

    1996-01-01

    Neuron-specific enolase (NSE) is a well-known marker of tumours that have neuroendocrine origin. High levels of NSE have also been described in various types of testicular germ cell neoplasms, particularly in seminomas. To evaluate the presence of NSE in testicular carcinoma-in situ (CIS), a...

  15. Determination of serum neuron specific enolase and glutathion S transferases levels in patients with acute cerebral infarction and its clinical significance

    International Nuclear Information System (INIS)

    Objective: To evaluate the variation of serum neuron specific enolase (NSE) and glutathion S transferases (GST) levels in patients with cerebral infarction and its clinical significance. Methods: The serum levels of NSE in cerebral infarction patients were determined with immunoradiometric assay (IRMA), and the serum level of GST were determined by enzyme immuno sandwich assay (ELISA). Results: Serum NSE levels linked in patients were significantly higher (p<0.01) and GST serum levels were significantly lower (p < 0.01) within 3 days after onset of disease than those at two weeks and those in the controls. There was a positive correlation between serum NSE levels and neurological deficit scores (p < 0.001) and a negative correlation with serum GST levels (p < 0.05). There was also a close relationship between the serum NSE levels and the volume of infarction (p < 0.001). Conclusion: There was a close relationship between the Serum levels of NSE, GST and clinical features of Patients in the early stage of cerebral infarction

  16. Utility of squamous cell carcinoma antigen, carcinoembryonic antigen, Cyfra 21-1 and neuron specific enolase in lung cancer diagnosis: a prospective study from China

    Institute of Scientific and Technical Information of China (English)

    SONG Wei-an; LIU Xi; TIAN Xiao-dong; WANG Wei; LIANG Chao-yang; ZHANG Tao; GUO Jun-tang; PENG Yang-hong; ZHOU Nai-kang

    2011-01-01

    Background Early detection and diagnosis is urgent for the sake of effective treatment strategy for lung cancer.However,a convenient,economical and relatively precise method is not available.We here report a prospective study to find the possible value of the combined use of four popular tumor markers in the early diagnosis of lung cancer among patients with suspicious nodules in the lung.Methods Six hundred and sixty inpatients with suspicious nodules in the lung were divided into a lung cancer group and a benign pulmonary tumor group according to post-operative histological examinations.Serum levels of four tumor markers including squamous cell carcinoma antigen (SCC),carcinoembryonic antigen (CEA),Cyfra 21-1 and neuron specific enolase (NSE) were assayed for each patient.Receiver operating characteristic (ROC) curves were constructed for each tumor marker.The power of lung cancer diagnosis of each tumor marker,as well as a combination of them were analyzed and compared.Results The serum levels (median,range) of SCC,CEA,Cyfra 21-1 and NSE were 0.44 (0.01-35.70) ng/ml,2.49(0.30-26.78) ng/ml,2.30 (0.82-73.33) ng/ml and 10.54 (0.10-56.41) ng/ml respectively in lung cancer group,and were 0.32 (0.01-0.90) ng/ml,1.60 (0.20-8.93) ng/ml,1.41 (0.72-4.82) ng/ml and 9.36 (6.56-24.24) ng/ml respectively in the benign pulmonary tumor group.The difference in each tumor marker between the two groups was significant (P <0.05).The ROCs of SCC,CEA,Cyfra 21-1 and NSE were 0.702 (95% CI,0.654-0.751),0.611 (95% CI,0.563-0.659),0.650(95% CI,0.601-0.700) and 0.598 (95% CI,0.542-0.654) respectively,indicating very low power of these four tumor markers.When a combination of SCC,CEA,Cyfra 21-1 and NSE were employed,the diagnosis power was strengthened.Conclusion SCC,CEA,Cyfra 21-1 and NSE are valuable in the early diagnosis of lung cancer among suspicious nodules in the lung,especially when they were assayed together for one patient.

  17. Early detection of response in small cell bronchogenic carcinoma by changes in serum concentrations of creatine kinase, neuron specific enolase, calcitonin, ACTH, serotonin and gastrin releasing peptide

    DEFF Research Database (Denmark)

    Bork, E; Hansen, M; Urdal, P;

    1988-01-01

    Creatine kinase (CK-BB), neuron specific enolase (NSE), ACTH, calcitonin, serotonin and gastrin releasing peptide (GRP) were measured in serum or plasma before and immediately after initiation of treatment in patients with small cell lung cancer (SCC). Pretherapeutic elevated concentrations of CK...... extensive and local disease patients. Serotonin was generally overall elevated in 10% and GRP in 7% but elevations were seen only in patients with extensive disease. Out of the four most frequently elevated substances at least one marker was elevated in 80% of all the patients, including 91% in extensive...... determined within 4-8 weeks. The results indicate that serum CK-BB and NSE are potential markers for SCC at the time of diagnosis and that changes in the concentrations during the first course of cytostatic therapy are promising as biochemical tests for early detection of response to chemotherapy....

  18. Mycoplasma synoviae enolase is a plasminogen/fibronectin binding protein

    OpenAIRE

    Bao, Shijun; Guo, Xiaoqin; Yu, Shengqing; Ding, Jiabo; Tan, Lei; Zhang, Fanqin; Sun, Yingjie; Qiu, Xusheng; Chen, Guanghua; Ding, Chan

    2014-01-01

    Background Mycoplasma synoviae is an avian pathogen that can lead to respiratory tract infections and arthritis in chickens and turkeys, resulting in serious economic losses to the poultry industry. Enolase reportedly plays important roles in several bacterial pathogens, but its role in M. synoviae has not been established. Therefore, in this study, the enolase encoding gene (eno) of M. synoviae was amplified from strain WVU1853 and expressed in E. coli BL21 cells. Then the enzymatic activity...

  19. Cloning and expression of human neuron-specific enolase cDNA in Escherichia coli.

    Science.gov (United States)

    Pavlov, K A; Gurina, O I; Antonova, O M; Semenova, A V; Chekhonin, V P

    2011-12-01

    cDNA fragment encoding neuron-specific enolase was amplified from the cDNA library of human brain. Then the fragment was cloned for expression in E. coli using the vector pET28-a. High level of neuron-specific enolase expression was confirmed by SDS-PAAG electrophoresis and immunochemical identity by immunoblot analysis. The constructed producer strain is the cheapest source of neuron-specific enolase suitable for the use in diagnostic applications. PMID:22808461

  20. Co-purification of arrestin like proteins with alpha-enolase from bovine myocardial tissues and the possible role in heart diseases as an autoantigen

    Energy Technology Data Exchange (ETDEWEB)

    Mirshahi, M., E-mail: massoud.mirshahi@inserm.fr; Le Marchand, S.

    2015-05-08

    Aim: Previously, we reported that visual arrestin co-purified with glycolytic enzymes. The aim of this study was to analyze the co-purification of arrestin like proteins (ALP) in bovine cardiac tissues with enolases. Methods: The soluble extract of bovine myocardial tissues from different regions such as left and right atriums and ventricles of the bovine heart (n = 3) was analyzed by ACA-34 gel filtration, immuno-affinity column, SDS-PAGE, ELISA, western blot and a sandwich immune assay for quantification of ALP and sequence analysis. Results: We observed that; 1) The cardiac muscle contained a 50 kDa ALP at a concentration of 751 pg/mg of soluble protein extract, 2) ALP purified, by immunoaffinity, contained alpha-enolase of 48 kDa confirmed by protein sequence analysis; 3) Cardiomyocyte cells exposed to anti arrestin and anti enolase monoclonal antibodies showed decreased proliferation in vitro, 4) High level of autoantibodies were detected by ELISA (3.57% for arrestin and 9.12% for α-enolase) in serum of patients with infarcted heart disease. Conclusion: We suggest a possible interaction between ALP and alpha-enolases yielding a complex that may be involved in the induction of cardiac autoimmune diseases. - Highlights: • We examine a possible interaction between arrestin like protein and alpha-enolases in cardiomyocyte. • We demonstrated the effect of antibodies against arrestin and enolase on cardiomyocyte cell proliferation. • We suggest that this proteins complex may be involved in the induction of cardiac autoimmune diseases.

  1. Co-purification of arrestin like proteins with alpha-enolase from bovine myocardial tissues and the possible role in heart diseases as an autoantigen

    International Nuclear Information System (INIS)

    Aim: Previously, we reported that visual arrestin co-purified with glycolytic enzymes. The aim of this study was to analyze the co-purification of arrestin like proteins (ALP) in bovine cardiac tissues with enolases. Methods: The soluble extract of bovine myocardial tissues from different regions such as left and right atriums and ventricles of the bovine heart (n = 3) was analyzed by ACA-34 gel filtration, immuno-affinity column, SDS-PAGE, ELISA, western blot and a sandwich immune assay for quantification of ALP and sequence analysis. Results: We observed that; 1) The cardiac muscle contained a 50 kDa ALP at a concentration of 751 pg/mg of soluble protein extract, 2) ALP purified, by immunoaffinity, contained alpha-enolase of 48 kDa confirmed by protein sequence analysis; 3) Cardiomyocyte cells exposed to anti arrestin and anti enolase monoclonal antibodies showed decreased proliferation in vitro, 4) High level of autoantibodies were detected by ELISA (3.57% for arrestin and 9.12% for α-enolase) in serum of patients with infarcted heart disease. Conclusion: We suggest a possible interaction between ALP and alpha-enolases yielding a complex that may be involved in the induction of cardiac autoimmune diseases. - Highlights: • We examine a possible interaction between arrestin like protein and alpha-enolases in cardiomyocyte. • We demonstrated the effect of antibodies against arrestin and enolase on cardiomyocyte cell proliferation. • We suggest that this proteins complex may be involved in the induction of cardiac autoimmune diseases

  2. Food vacuole associated enolase in plasmodium undergoes multiple post-translational modifications: evidence for atypical ubiquitination.

    Directory of Open Access Journals (Sweden)

    Saudamini Shevade

    Full Text Available Plasmodium enolase localizes to several sub-cellular compartments viz. cytosol, nucleus, cell membrane, food vacuole (FV and cytoskeleton, without having any organelle targeting signal sequences. This enzyme has been shown to undergo multiple post-translational modifications (PTMs giving rise to several variants that show organelle specific localization. It is likely that these PTMs may be responsible for its diverse distribution and moonlighting functions. While most variants have a MW of ~50 kDa and are likely to arise due to changes in pI, food vacuole (FV associated enolase showed three forms with MW~50, 65 and 75 kDa. Evidence from immuno-precipitation and western analysis indicates that the 65 and 75 kDa forms of FV associated enolase are ubiquitinated. Using mass spectrometry (MS, definitive evidence is obtained for the nature of PTMs in FV associated variants of enolase. Results showed several modifications, viz. ubiquitination at K147, phosphorylation at Y148 and acetylation at K142 and K384. MS data also revealed the conjugation of three ubiquitin (Ub molecules to enolase through K147. Trimeric ubiquitin has a linear peptide linkage between the NH2-terminal methionine of the first ubiquitin (Ub1 and the C-terminal G76 of the second (Ub2. Ub2 and third ubiquitin (Ub3 were linked through an atypical isopeptide linkage between K6 of Ub2 and G76 of Ub3, respectively. Further, the tri-ubiquitinated form was found to be largely associated with hemozoin while the 50 and 65 kDa forms were present in the NP-40 soluble fraction of FV. Mass spectrometry results also showed phosphorylation of S42 in the cytosolic enolase from P. falciparum and T337 in the cytoskeleton associated enolase from P. yoelii. The composition of food vacuolar proteome and likely interactors of enolase are also being reported.

  3. Alpha-enolase immunohistochemical study of stem cels in limbal autograft tissue

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    Kozák I.

    2003-01-01

    Full Text Available The purpose was to detect the presence of stem cells in the limbus and cornea following limbal autotransplantation for chemical eye injury. Fifteen New Zealand white rabbits were used in the study. The first group (n=5 served as healthy controls, in group 2 (n=5 one eye of each rabbit was burnt by 1 N NaOH and in the last group (n=5, the same chemical injury was followed by limbal autotransplantation from the contralateral eye. The eyes were examined clinically and studied by immunohistochemistry after the enucleation. A monoclonal antibody against alpha-enolase - a biochemical marker for stem cells, was used for immunohistochemistry. The density of cells in both limbus and the cornea was calculated per mm2. The Tukey-Kramer parametric test was used for statistical evaluation. Healthy limbus and basal corneal layers showed numerous a-enolase positive cells. The injured group showed marked depletion of these cells (p<0.0001. Compared to this group the treated group, exibited a statistically significant (p<0.0001 increase in a-enolase positive cells. This study provides evidence that limbal autograft transplantation transfers stem cells to the chemically injured area with depletion of these cells. To demonstrate this we used a monoclonal antibody against alpha-enolase for immunohistochemical analysis of the presence of stem cells in transplant tissue and its surrounding milieu. We showed that the transplanted limbal graft was the source of corneal epithelial stem cells.

  4. Cloning and Characterization of Surface-Localized α-Enolase of Streptococcus iniae, an Effective Protective Antigen in Mice

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    Jun Wang

    2015-06-01

    Full Text Available Streptococcus iniae is a major fish pathogen that can also cause human bacteremia, cellulitis and meningitis. Screening for and identification of protective antigens plays an important role in developing therapies against S. iniae infections. In this study, we indicated that the α-enolase of S. iniae was not only distributed in the cytoplasm and associated to cell walls, but was also secreted to the bacterial cell surface. The functional identity of the purified recombinant α-enolase protein was verified by its ability to catalyze the conversion of 2-phosphoglycerate (2-PGE to phosphoenolpyruvate (PEP, and both the recombinant and native proteins interacted with human plasminogen. The rabbit anti-rENO serum blockade assay shows that α-enolase participates in S. iniae adhesion to and invasion of BHK-21 cells. In addition, the recombinant α-enolase can confer effective protection against S. iniae infection in mice, which suggests that α-enolase has potential as a vaccine candidate in mammals. We conclude that S. iniae α-enolase is a moonlighting protein that also associates with the bacterial outer surface and functions as a protective antigen in mice.

  5. Crystallization and preliminary X-ray analysis of 2,3-diketo-5-methylthiopentyl-1-phosphate enolase from Bacillus subtilis

    International Nuclear Information System (INIS)

    Crystals of the 45.1 kDa functional form of 2,3-diketo-5-methylthiopentyl-1-phosphate enolase from B. subtilis diffracted to 2.30 Å resolution. 2,3-Diketo-5-methylthiopentyl-1-phosphate enolase (DK-MTP-1P enolase) from Bacillus subtilis was crystallized using the hanging-drop vapour-diffusion method. Crystals grew using PEG 3350 as the precipitant at 293 K. The crystals diffracted to 2.3 Å resolution at 100 K using synchrotron radiation and were found to belong to the monoclinic space group P21, with unit-cell parameters a = 79.3, b = 91.5, c = 107.0 Å, β = 90.8°. The asymmetric unit contained four molecules of DK-MTP-1P enolase, with a VM value of 2.2 Å3 Da−1 and a solvent content of 43%

  6. Role of Caveolin 1, E-Cadherin, Enolase 2 and PKCalpha on resistance to methotrexate in human HT29 colon cancer cells

    DEFF Research Database (Denmark)

    Selga, Elisabet; Morales Torres, Christina; Noé, Véronique;

    2008-01-01

    ABSTRACT: BACKGROUND: Methotrexate is one of the earliest cytotoxic drugs used in cancer therapy, and despite the isolation of multiple other folate antagonists, methotrexate maintains its significant role as a treatment for different types of cancer and other disorders. The usefulness of treatment...... with methotrexate is limited by the development of drug resistance, which may be acquired through different ways. To get insights into the mechanisms associated with drug resistance and sensitization we performed a functional analysis of genes deregulated in methotrexate resistant cells, either due to...

  7. EWGWS insert in Plasmodium falciparum ookinete surface enolase is involved in binding of PWWP containing peptides: Implications to mosquito midgut invasion by the parasite.

    Science.gov (United States)

    Mukherjee, Debanjan; Mishra, Pushpa; Joshi, Mamata; Thakur, Prasoon Kumar; Hosur, R V; Jarori, Gotam K

    2016-01-01

    There are multiple stages in the life cycle of Plasmodium that invade host cells. Molecular machinery involved is such host-pathogen interactions constitute excellent drug targets and/or vaccine candidates. A screen using a phage display library has previously demonstrated presence of enolase on the surface of the Plasmodium ookinete. Phage-displayed peptides that bound to the ookinete contained a conserved motif (PWWP) in their sequence. Here, direct binding of these peptides with recombinant Plasmodium falciparum enolase (rPfeno) was investigated. These peptides showed specific binding to rPfeno, but failed to bind to other enolases. Plasmodium spp enolases are distinct in having an insert of five amino acids ((104)EWGWS(108)) that is not found in host enolases. The possibility of this insert being the recognition motif for the PWWP containing peptides was examined, (i) by comparing the binding of the peptides with rPfeno and a deletion variant Δ-rPfeno lacking (104)EWGWS(108), (ii) by measuring the changes in proton chemical shifts of PWWP peptides on binding to different enolases and (iii) by inter-molecular docking experiment to locate the peptide binding site. Results from these studies showed that the pentapeptide insert of Pfeno indeed constitutes the binding site for the PWWP domain containing peptide ligands. Search for sequences homologous to phage displayed peptides among peritrophic matrix proteins resulted in identification of perlecan, laminin, peritrophin and spacran. The possibility of these PWWP domain-containing proteins in the peritrophic matrix of insect gut to interact with ookinete cell surface enolase and facilitate the invasion of mosquito midgut epithelium is discussed. PMID:26592350

  8. Systematic Analysis Reveals Elongation Factor 2 and α-Enolase as Novel Interaction Partners of AKT2.

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    Katharina Bottermann

    Full Text Available AKT2 is one of the three isoforms of the protein kinase AKT being involved in the modulation of cellular metabolism. Since protein-protein interactions are one possibility to convey specificity in signal transduction, we performed AKT2-protein interaction analysis to elucidate their relevance for AKT2-dependent cellular functions. We identified heat shock protein 90 kDa (HSP90, Cdc37, heat shock protein 70 kDa (HSP70, 78 kDa glucose regulated protein (GRP78, tubulin, GAPDH, α-enolase and elongation factor 2 (EF2 as AKT2-interacting proteins by a combination of tandem affinity purification and mass spectrometry in HEK293T cells. Quantitative MS-analysis using stable isotope labeling by amino acids in cell culture (SILAC revealed that only HSP90 and Cdc37 interact stably with AKT2, whereas the other proteins interact with low affinity with AKT2. The interactions of AKT2 with α-enolase and EF2 were further analyzed in order to uncover the functional relevance of these newly discovered binding partners. Despite the interaction of AKT2 and α-enolase, which was additionally validated by proximity ligation assay (PLA, no significant impact of AKT on α-enolase activity was detected in activity measurements. AKT stimulation via insulin and/or inhibition with the ATP-competitive inhibitor CCT128930 did not alter enzymatic activity of α-enolase. Interestingly, the direct interaction of AKT2 and EF2 was found to be dynamically regulated in embryonic rat cardiomyocytes. Treatment with the PI3-kinase inhibitor LY294002 before stimulation with several hormones stabilized the complex, whereas stimulation alone led to complex dissociation which was analyzed in situ with PLA. Taken together, these findings point to new aspects of AKT2-mediated signal transduction in protein synthesis and glucose metabolism.

  9. Nanostructured materials detect epidermal growth factor receptor, neuron specific enolase and carcinoembryonic antigen

    Science.gov (United States)

    Stefan-van Staden, Raluca-Ioana; Comnea-Stancu, Ionela Raluca; Surdu-Bob, Carmen Cristina; Badulescu, Marius

    2015-09-01

    New nanostructured materials based on thin films of Cu and Ni deposited on textile material (veil), as well as gold nanostructured microspheres were used for the design of new stochastic sensors. The stochastic sensors were able to detect simultaneously a panel of biomarkers comprising epidermal growth factor receptor, neuron specific enolase, and carcinoembryonic antigen from whole blood samples with high reliabilities - recovery tests higher than 97.00%, with a RSD (%) lower than 0.1%. The stochastic sensors had shown high sensitivities and low determination levels for the detection of the proposed panel of biomarkers making early detection of lung cancer possible by fast screening of whole blood.

  10. Nuclear glycolytic enzyme enolase of Toxoplasma gondii functions as a transcriptional regulator.

    Directory of Open Access Journals (Sweden)

    Thomas Mouveaux

    Full Text Available Apicomplexan parasites including Toxoplasma gondii have complex life cycles within different hosts and their infectivity relies on their capacity to regulate gene expression. However, little is known about the nuclear factors that regulate gene expression in these pathogens. Here, we report that T. gondii enolase TgENO2 is targeted to the nucleus of actively replicating parasites, where it specifically binds to nuclear chromatin in vivo. Using a ChIP-Seq technique, we provide evidence for TgENO2 enrichment at the 5' untranslated gene regions containing the putative promoters of 241 nuclear genes. Ectopic expression of HA-tagged TgENO1 or TgENO2 led to changes in transcript levels of numerous gene targets. Targeted disruption of TgENO1 gene results in a decrease in brain cyst burden of chronically infected mice and in changes in transcript levels of several nuclear genes. Complementation of this knockout mutant with ectopic TgENO1-HA fully restored normal transcript levels. Our findings reveal that enolase functions extend beyond glycolytic activity and include a direct role in coordinating gene regulation in T. gondii.

  11. Nuclear glycolytic enzyme enolase of Toxoplasma gondii functions as a transcriptional regulator.

    Science.gov (United States)

    Mouveaux, Thomas; Oria, Gabrielle; Werkmeister, Elisabeth; Slomianny, Christian; Fox, Barbara A; Bzik, David J; Tomavo, Stanislas

    2014-01-01

    Apicomplexan parasites including Toxoplasma gondii have complex life cycles within different hosts and their infectivity relies on their capacity to regulate gene expression. However, little is known about the nuclear factors that regulate gene expression in these pathogens. Here, we report that T. gondii enolase TgENO2 is targeted to the nucleus of actively replicating parasites, where it specifically binds to nuclear chromatin in vivo. Using a ChIP-Seq technique, we provide evidence for TgENO2 enrichment at the 5' untranslated gene regions containing the putative promoters of 241 nuclear genes. Ectopic expression of HA-tagged TgENO1 or TgENO2 led to changes in transcript levels of numerous gene targets. Targeted disruption of TgENO1 gene results in a decrease in brain cyst burden of chronically infected mice and in changes in transcript levels of several nuclear genes. Complementation of this knockout mutant with ectopic TgENO1-HA fully restored normal transcript levels. Our findings reveal that enolase functions extend beyond glycolytic activity and include a direct role in coordinating gene regulation in T. gondii. PMID:25153525

  12. Stability of the octameric structure affects plasminogen-binding capacity of streptococcal enolase.

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    Amanda J Cork

    Full Text Available Group A Streptococcus (GAS is a human pathogen that has the potential to cause invasive disease by binding and activating human plasmin(ogen. Streptococcal surface enolase (SEN is an octameric α-enolase that is localized at the GAS cell surface. In addition to its glycolytic role inside the cell, SEN functions as a receptor for plasmin(ogen on the bacterial surface, but the understanding of the molecular basis of plasmin(ogen binding is limited. In this study, we determined the crystal and solution structures of GAS SEN and characterized the increased plasminogen binding by two SEN mutants. The plasminogen binding ability of SENK312A and SENK362A is ~2- and ~3.4-fold greater than for the wild-type protein. A combination of thermal stability assays, native mass spectrometry and X-ray crystallography approaches shows that increased plasminogen binding ability correlates with decreased stability of the octamer. We propose that decreased stability of the octameric structure facilitates the access of plasmin(ogen to its binding sites, leading to more efficient plasmin(ogen binding and activation.

  13. Stability of the Octameric Structure Affects Plasminogen-Binding Capacity of Streptococcal Enolase

    Science.gov (United States)

    Law, Ruby H. P.; Casey, Lachlan W.; Valkov, Eugene; Bertozzi, Carlo; Stamp, Anna; Jovcevski, Blagojce; Aquilina, J. Andrew; Whisstock, James C.; Walker, Mark J.; Kobe, Bostjan

    2015-01-01

    Group A Streptococcus (GAS) is a human pathogen that has the potential to cause invasive disease by binding and activating human plasmin(ogen). Streptococcal surface enolase (SEN) is an octameric α-enolase that is localized at the GAS cell surface. In addition to its glycolytic role inside the cell, SEN functions as a receptor for plasmin(ogen) on the bacterial surface, but the understanding of the molecular basis of plasmin(ogen) binding is limited. In this study, we determined the crystal and solution structures of GAS SEN and characterized the increased plasminogen binding by two SEN mutants. The plasminogen binding ability of SENK312A and SENK362A is ~2- and ~3.4-fold greater than for the wild-type protein. A combination of thermal stability assays, native mass spectrometry and X-ray crystallography approaches shows that increased plasminogen binding ability correlates with decreased stability of the octamer. We propose that decreased stability of the octameric structure facilitates the access of plasmin(ogen) to its binding sites, leading to more efficient plasmin(ogen) binding and activation. PMID:25807546

  14. Molecular Cloning and Characterization of Enolase from Oilseed Rape (Brassica napus)

    Institute of Scientific and Technical Information of China (English)

    ZHAOJing-Ya; ZUOKai-Jing; QINJie; TANGKe-Xuan

    2004-01-01

    An enolase-encoding cDNA clone in oilseed rape (Brassica napus L.) was isolated. This gene (accession number: AY307449) had a total length of 1 624 bp with an open reading frame of 1 335 bp, and encoded a predicted polypeptide of 444 amino acids with a molecular weight of 47.38 kD. The deduced amino acid sequence shared identity with a number of enolases ranging from Bacillus subtilis to human beings and had much higher identity with other plant enolases than with enolases from Bacillus, yeast and human beings. Comparison of its primary structure with those of other enolases revealed the presence of an insertion of five amino acids in enolase of B. napus. Southern blotting analysis of genomic DNA indicated that enolase was likely to be a low-copy gene in the oilseed rape genome. Expression of the cloned enolase gene increased under salt stress, but decreased in response to low temperature. Our studies suggested that the cloned gene was a new member of plant enolase gene family, which contributed to the energy supply in stress-treated tissues.

  15. Enolase of Streptococcus Suis Serotype 2 Enhances Blood-Brain Barrier Permeability by Inducing IL-8 Release.

    Science.gov (United States)

    Sun, Yingying; Li, Na; Zhang, Jing; Liu, Hongtao; Liu, Jianfang; Xia, Xiaojing; Sun, Changjiang; Feng, Xin; Gu, Jingmin; Du, Chongtao; Han, Wenyu; Lei, Liancheng

    2016-04-01

    Streptococcus suis serotype 2 (SS2) is an emerging zoonosis, and meningitis is the most frequent clinical manifestation, but mechanism of its virulent factor, enolase (Eno), is unknown in meningitis. In this study, Eno was inducibly expressed and added to an in vitro Transwell co-culture model of the blood-brain barrier (BBB) consisted of porcine brain microvascular endothelial cells (PBMECs) and astrocytes (ACs), the results showed that Eno induces a significant increase in BBB permeability and promotes the release of IL-8 et al. cytokines. Furthermore, IL-8 could significantly destroy the integrity of the BBB model in vitro. In mice models administered Eno for 24 h, Eno could significantly promote Evans blue (EB) moving from the blood to the brain and significantly increased the serum and brain levels of IL-8, as detected by ELISA. While G31P (IL-8 receptor antagonist) significantly decreased the concentration of EB in the brains of mice injected with Eno. The present study demonstrated that SS2 Eno may play an important role in disrupting BBB integrity by prompting IL-8 release. PMID:26732390

  16. Plasma autoantibodies against heat shock protein 70, enolase 1 and ribonuclease/angiogenin inhibitor 1 as potential biomarkers for cholangiocarcinoma.

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    Rucksak Rucksaken

    Full Text Available The diagnosis of cholangiocarcinoma (CCA is often challenging, leading to poor prognosis. CCA arises via chronic inflammation which may be associated with autoantibodies production. This study aims to identify IgG antibodies directed at self-proteins and tumor-associated antigens. Proteins derived from immortalized cholangiocyte cell line (MMNK1 and CCA cell lines (M055, M214 and M139 were separated using 2-dimensional electrophoresis and incubated with pooled plasma of patients with CCA and non-neoplastic controls by immunoblotting. Twenty five immunoreactive spots against all cell lines-derived proteins were observed on stained gels and studied by LC-MS/MS. Among these, heat shock protein 70 (HSP70, enolase 1 (ENO1 and ribonuclease/angiogenin inhibitor 1 (RNH1 obtained the highest matching scores and were thus selected for further validation. Western blot revealed immunoreactivity against HSP70 and RNH1 in the majority of CCA cases and weakly in healthy individuals. Further, ELISA showed that plasma HSP70 autoantibody level in CCA was significantly capable to discriminate CCA from healthy individuals with an area under the receiver operating characteristic curve of 0.9158 (cut-off 0.2630, 93.55% sensitivity and 73.91% specificity. Plasma levels of IgG autoantibodies against HSP70 were correlated with progression from healthy individuals to cholangitis to CCA (r = 0.679, P<0.001. In addition, circulating ENO1 and RNH1 autoantibodies levels were also significantly higher in cholangitis and CCA compared to healthy controls (P<0.05. Moreover, the combinations of HSP70, ENO1 or RNH1 autoantibodies positivity rates improved specificity to over 78%. In conclusion, plasma IgG autoantibodies against HSP70, ENO1 and RNH1 may represent new diagnostic markers for CCA.

  17. Serum levels of ferritin and neuron-specific enolase in children with hand-foot-mouth disease complicated by acute viral encephalitis%手足口病合并病毒性脑炎患儿血清铁蛋白及神经元特异性烯醇化酶水平的变化及意义

    Institute of Scientific and Technical Information of China (English)

    丰炳峰; 朱孔荣

    2012-01-01

    目的 探讨手足口病(HFMD)合并病毒性脑炎患儿血清铁蛋白和神经元特异性烯醇化酶(NSE)水平的变化及意义.方法 采用酶联免疫吸附(ELISA)与电化学发光法对20例HFMD合并病毒性脑炎(脑炎组)和20例单纯HFMD患儿(单纯HFMD组)进行血清铁蛋白和NSE水平测定,并与20例正常健康儿(对照组)进行比较.结果 脑炎组血清铁蛋白含量为212+71μg/L,明显高于单纯HFMD组(85±18 μg/L)及对照组(70±15μgL)(均P<0.01);脑炎组血清NSE含量(8.6±2.6μg/L)亦明显高于单纯HFMD组(6.0±1.3 μg/L)及对照组(5.6±1.8 μg/L),(均P <0.01).治疗后脑炎组血清铁蛋白及NSE分别下降至126±37 μg/L、6.8±1.9μg/L,较治疗前差异有统计学意义(P<0.01).结论 HFMD合并病毒性脑炎患儿血清铁蛋白和NSE含量显著升高,对血清铁蛋白和NSE含量的检测有利于HFMD合并病毒性脑炎的早期诊断.%Objective To study serum feiritin and neuron-specific enolase (NSE) levels in children with hand-foot-mouth disease (HFMD) complicated by acute viral encephalitis and their clinical significance. Methods Serum levels of ferritin and NSE were measured using ELISA and electrochemical luminescence in 20 children with HFMD complicated by viral encephalitis (encephalitis group), 20 children with HFMD only (simple HFMD group) and 20 healthy children (control group). Results Serum levels of ferritin in the encephalitis group (212 ±71 μg/L) were significantly higher than in the simple HFMD group (85 ±18 μg/L) and control group (70 ± 15 μg/L) (P<0.01). Serum levels of NSE in the encephalitis group (8.6 ±2.6 μg/L) were also significantly higher than in the simple HFMD group (6.0± 1.3 μg/L) and control group (5. 6 ± 1. 8 μg/L) {P < 0. 01 ). Significantly decreased serum ferritin ( 126 ± 37 μg/L) and NSE levels! (6.8 ±1.9 μg/L) were found in the encephalitis group (P < 0. 01) after treatment. Conclusions Serum levels of ferritin and NSE in children with

  18. The interaction of streptococcal enolase with canine plasminogen: the role of surfaces in complex formation.

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    Vinod Balhara

    Full Text Available The enolase from Streptococcus pyogenes (Str enolase F137L/E363G is a homo-octamer shaped like a donut. Plasminogen (Pgn is a monomeric protein composed of seven discrete separated domains organized into a lock washer. The enolase is known to bind Pgn. In past work we searched for conditions in which the two proteins would bind to one another. The two native proteins in solution would not bind under any of the tried conditions. We found that if the structures were perturbed binding would occur. We stated that only the non-native Str enolase or Pgn would interact such that we could detect binding. We report here the results of a series of dual polarization interferometry (DPI experiments coupled with atomic force microscopy (AFM, isothermal titration calorimetry (ITC, dynamic light scattering (DLS, and fluorescence. We show that the critical condition for forming stable complexes of the two native proteins involves Str enolase binding to a surface. Surfaces that attract Str enolase are a sufficient condition for binding Pgn. Under certain conditions, Pgn adsorbed to a surface will bind Str enolase.

  19. Different Effects of Phenobarbital and Gingko Leaf on Level of Serum Neuron-Specific Enolase after Simple Febrile Seizure in Children%苯巴比妥和银杏叶对单纯性热性惊厥患儿血清神经元特异性烯醇化酶的不同影响

    Institute of Scientific and Technical Information of China (English)

    陈曦; 白珺; 丁守梅; 李星萍; 古丽鲜·阿布都热依木; 朱炎

    2011-01-01

    目的 对照观察苯巴比妥(PB)和银杏叶对单纯性惊厥(SFC)患儿血清神经元特异性烯醇化酶(NSE)的不同影响.方法 将符合纳入标准的188例SFC患儿随机分为3组.惊厥未用药组:不使用PB及其他止惊药; PB止惊组:予PB(10 mg·kg-1,但每次不超过150 mg)肌肉注射, 每天1次,连用2 d; PB+银杏叶组:在PB止惊组基础上当日予银杏叶,≤2岁每次15 mg、>2岁每次20 mg,每天2次,连用14 d.健康对照组为同期体检抽血的健康儿童.各组SFC患儿分别在惊厥发作后2 d、7 d及21 d时取静脉血2 mL,ELISA法测定血清NSE水平.结果 1.>2岁组:各SFC组在惊厥发作后2 d时NSE水平均较健康对照组明显增高(Pa<0.01).7 d时NSE水平均有回落,但仍较健康对照组高(Pa<0.05,0.01).2.≤2岁组:PB止惊组在2 d、7 d和21 d时NSE均高于惊厥未用药组(Pa<0.01).而PB+银杏叶组在各时点NSE水平的增高与惊厥未用药组比较差异无统计学意义.惊厥未用药组NSE水平在2 d、7 d、21 d各时点呈逐渐下降过程.而PB组在各时点的NSE水平递降却不明显.PB+银杏叶组NSE水平自7 d 起至21 d 时明显递减.结论 SFC发作引起儿童血清NSE水平一过性轻度增高,提示惊厥性脑损伤的存在; PB会进一步加重SFC患儿血清NSE水平的增高,银杏叶则有降低血清NSE水平和减轻PB脑损伤的可能性.%Objective To observe the different effects of phenobarbital ( PB ) and Ginkgo Leaf pills on the serum neuron - specific enolase (NSE) level after seizures of simple febrile convulsion (SFC) in children.Methods One hundred and eighty -eight children were included,all of them were hospitalized due to their seizures of SFC, who randomly divided into 3 groups.Seizures without any drug group: seizures without any anticonvulsive drugs or PB;PB treatment group:PB with a dosage of 10 mg · kg-1 (the highest dose less than 150 mg ·kg-1) was given through intramuscular injection,once a day,for 2 days.PB plus Gingko Leaf

  20. Neurone-specific enolase and S-100: new markers for delineating the innervation of the respiratory tract in man and other mammals.

    OpenAIRE

    Sheppard, M N; Kurian, S S; Henzen-Logmans, S C; Michetti, F; Cocchia, D; Cole , P; Rush, R A; Marangos, P. J.; Bloom, S. R.; Polak, J M

    1983-01-01

    Lung innervation has been studied in the past by methylene blue staining and silver impregnation and more recently by histochemical methods. These techniques give only a partial picture of the total innervation. We have delineated the innervation of the lung in man and three other mammalian species by immunostaining with antibodies to two new markers of nervous tissue. These markers are neurone-specific enolase (NSE), an enzyme present in nerve cells in both the central and the peripheral ner...

  1. Research of the serum level of neuron-specific enolase in children with various types of seizure%不同发作类型癫(癎)患儿血清神经元特异性烯醇化酶水平研究

    Institute of Scientific and Technical Information of China (English)

    王春; 翟琼香; 汤志鸿; 卓木清

    2012-01-01

    目的 探讨不同发作类型癫(癎)患儿血清神经元特异性烯醇化酶水平变化与脑损害之间的关系.方法 按照1981年国际抗癫(癎)联盟制定的癫(癎)发作类型分类标准,共明确诊断190例癫(癎)患儿(强直-阵挛发作41例、强直性发作34例、阵挛性发作22例、肌阵挛发作12例、无张力性发作17例、失神发作22例、单纯部分性发作21例及复杂部分性发作21例),于癫(癎)发作72 h内施行长程视频脑电图观察和血清神经元特异性烯醇化酶检测.结果 不同发作类型癫(癎)患儿血清神经元特异性烯醇化酶水平均高于正常对照组(P=0.000),其中以肌阵挛发作组[(32.42±6.62)ng/ml]水平最高,除与强直-阵挛发作组(P=0.062)外,与其他各发作类型之间差异均有统计学意义(P=0.000);而其他各类型之间差异无统计学意义(均P>0.05).秩相关分析显示,癫(癎)患儿血清神经元特异性烯醇化酶水平与长程视频脑电图异常程度呈正相关(rs=O.613,P=0.000).结论 癫(癎)发作后血清神经元特异性烯醇化酶水平即升高,提示癫(癎)发作对患儿脑组织有一定损害;而且癫(癎)放电对神经元损害越严重、血清神经元特异性烯醇化酶水平升高越明显,不同发作类型中以肌阵挛发作、强直-阵挛发作患儿血清神经元特异性烯醇化酶水平最高,提示这两种发作类型对脑组织的损害高于其他类型.%Objective To explore the relevance between the level changes of serum neuron-specific enolase (NSE) and neuronal damage in various seizure types of children with epilepsy. Methods According to the classification criteria of seizure types formulated by International League Against Epilepsy (ILAE) in 1981, 190 children with epilepsy were enrolled including tonic-clonic seizure group (41 cases), tonic seizure group (34 cases), clonic seizure group (22 cases), myoclonic seizure group (12 cases), atonic seizure group (17 cases), absence

  2. Changes in plasma calcitonin gene-related peptide and serum neuron specific enolase in rats with acute cerebral ischemia after low-frequency electrical stimulation with different waveforms and intensities

    Institute of Scientific and Technical Information of China (English)

    Qiang Gao; Yonghong Yang; Shasha Li; Jing He; Chengqi He

    2011-01-01

    Following acute cerebral ischemia in rats, plasma calcitonin gene-related peptide decreased and the level of serum neuron specific enolase and the volume of the infarction increased. Square-wave and triangular-wave electrical stimulation with low or high intensities could increase the plasma calcitonin gene-related peptide, decrease the serum neuron specific enolase and reduce the infarction volume in the brain in rats with cerebral ischemia. There was no significant difference between different wave forms and intensities. The experimental findings indicate that low-frequency electrical stimulation with varying waveforms and intensities can treat acute cerebral ischemia in rats.

  3. Fasciola gigantica enolase is a major component of worm tegumental fraction protective against sheep fasciolosis.

    Science.gov (United States)

    Mahana, N; Abd-Allah, H A-S; Salah, M; Tallima, H; El Ridi, R

    2016-06-01

    Infection of cattle and sheep with the parasite Fasciola gigantica is a cause of important economic losses throughout Asia and Africa. Many of the available anthelmintics have undesirable side effects, and the parasite may acquire drug resistance as a result of mass and repeated treatments of livestock. Accordingly, the need for developing a vaccine is evident. Triton-soluble surface membrane and tegumental proteins (TSMTP) of 60, 32, and 28kDa previously shown to elicit protective immunity in mice against challenge F. gigantica infection were found to be strongly immunogenic in sheep eliciting vigorous specific antibody responses to a titer>1:16,000 as assessed by enzyme-linked immunosorbent assay. Furthermore, the 60kDa fraction induced production of antibodies able to bind to the surface membrane of newly excysted juvenile flukes and mediate their attrition in antibody-dependent complement- and cell-mediated cytotoxicity assays, and significant (PFasciola hepatica enolase, suggesting that a fasciolosis vaccine might be effective against both tropical and temperate liver flukes. PMID:26970372

  4. The Cloning and Characterization of the Enolase2 Gene of Gekko japonicus and Its Polyclonal Antibody Preparation

    Directory of Open Access Journals (Sweden)

    Mei Liu

    2013-04-01

    Full Text Available The enolase2 gene is usually expressed in mature neurons and also named neuron specific enolase (NSE. In the present study, we first obtained the NSE gene cDNA sequence by using the RACE method based on the expressed sequence tag (EST fragment from the cDNA library of Gekko japonicus and identified one transcript of about 2.2 kb in central nervous system of Gekko japonicus by Northern blotting. The open reading frame of NSE is 1305 bp, which encodes a 435 amino-acid protein. We further investigated the multi-tissue expression pattern of NSE by RT-PCR and found that the expression of NSE mRNA was very high in brain, spinal cord and low in heart, while it was not detectable in other tissues. The real-time quantitative PCR was used to investigate the time-dependent change in the expression of the NSE mRNA level after gecko spinal cord transection and found it significantly increased at one day, reaching its highest level three days post-injury and then decreasing at the seventh day of the experiment. The recombinant plasmid of pET-32a-NSE was constructed and induced to express His fused NSE protein. The purified NSE protein was used to immunize rabbits to generate polyclonal antisera. The titer of the antiserum was more than 1:65536 determined by ELISA. Western blotting showed that the prepared antibody could specifically recognize the recombinant and endogenous NSE protein. The result of immunohistochemistry revealed that positive signals were present in neurons of the brain and the spinal cord. This study provided the tools of cDNA and polyclonal antibody for studying NSE function in Gekko japonicus.

  5. Neuron-specific enolase in cerebrospinal fluid and plasma of patients with acute ischemic brain disease

    Directory of Open Access Journals (Sweden)

    Selaković Vesna M.

    2003-01-01

    Full Text Available The objective of this research was to determine the dynamics of change of neuron-specific enolase concentration in patients with acute ischemic brain disease in cerebrospinal fluid and plasma. The study included 103 patients, their mean age 58-66 years. The control group consisted of 16 patients, of matching age and sex, with radicular lesions of discal origin, subjected to diagnostic radiculography. Concentration of neuron-specific enolase was measured by a flouroimmunometric method. The results showed that the concentration of neuron-specific enolase in cerebrospinal fluid and plasma of patients with brain ischemic disease within first seven days significantly increased compared to the control. The highest increase of concentration was established in brain infarction, somewhat lower in reversible ischemic attack, and the lowest in transient ischemic attack. Maximal concentration was established on the 3rd-4th day upon the brain infarction. Neuron-specific enolase concentration in cerebrospinal fluid and plasma may be an indicator of pathophysiological processes in the acute phase of brain ischemia and is significant in early diagnostics and therapy of the disease.

  6. The structure of bradyzoite-specific enolase from Toxoplasma gondii reveals insights into its dual cytoplasmic and nuclear functions

    Energy Technology Data Exchange (ETDEWEB)

    Ruan, Jiapeng [Northwestern University, 320 E. Superior Street, Morton 7-601, Chicago, IL 60611 (United States); Mouveaux, Thomas [Université Lille Nord de France, (France); Light, Samuel H.; Minasov, George; Anderson, Wayne F. [Northwestern University, 320 E. Superior Street, Morton 7-601, Chicago, IL 60611 (United States); Tomavo, Stanislas [Université Lille Nord de France, (France); Ngô, Huân M., E-mail: h-ngo@northwestern.edu [Northwestern University, 320 E. Superior Street, Morton 7-601, Chicago, IL 60611 (United States); BrainMicro LLC, 21 Pendleton Street, New Haven, CT 06511 (United States)

    2015-03-01

    The second crystal structure of a parasite protein preferentially enriched in the brain cyst of T. gondii has been solved at 2.75 Å resolution. Bradyzoite enolase 1 is reported to have differential functions as a glycolytic enzyme and a transcriptional regulator in bradyzoites. In addition to catalyzing a central step in glycolysis, enolase assumes a remarkably diverse set of secondary functions in different organisms, including transcription regulation as documented for the oncogene c-Myc promoter-binding protein 1. The apicomplexan parasite Toxoplasma gondii differentially expresses two nuclear-localized, plant-like enolases: enolase 1 (TgENO1) in the latent bradyzoite cyst stage and enolase 2 (TgENO2) in the rapidly replicative tachyzoite stage. A 2.75 Å resolution crystal structure of bradyzoite enolase 1, the second structure to be reported of a bradyzoite-specific protein in Toxoplasma, captures an open conformational state and reveals that distinctive plant-like insertions are located on surface loops. The enolase 1 structure reveals that a unique residue, Glu164, in catalytic loop 2 may account for the lower activity of this cyst-stage isozyme. Recombinant TgENO1 specifically binds to a TTTTCT DNA motif present in the cyst matrix antigen 1 (TgMAG1) gene promoter as demonstrated by gel retardation. Furthermore, direct physical interactions of both nuclear TgENO1 and TgENO2 with the TgMAG1 gene promoter are demonstrated in vivo using chromatin immunoprecipitation (ChIP) assays. Structural and biochemical studies reveal that T. gondii enolase functions are multifaceted, including the coordination of gene regulation in parasitic stage development. Enolase 1 provides a potential lead in the design of drugs against Toxoplasma brain cysts.

  7. The structure of bradyzoite-specific enolase from Toxoplasma gondii reveals insights into its dual cytoplasmic and nuclear functions

    International Nuclear Information System (INIS)

    The second crystal structure of a parasite protein preferentially enriched in the brain cyst of T. gondii has been solved at 2.75 Å resolution. Bradyzoite enolase 1 is reported to have differential functions as a glycolytic enzyme and a transcriptional regulator in bradyzoites. In addition to catalyzing a central step in glycolysis, enolase assumes a remarkably diverse set of secondary functions in different organisms, including transcription regulation as documented for the oncogene c-Myc promoter-binding protein 1. The apicomplexan parasite Toxoplasma gondii differentially expresses two nuclear-localized, plant-like enolases: enolase 1 (TgENO1) in the latent bradyzoite cyst stage and enolase 2 (TgENO2) in the rapidly replicative tachyzoite stage. A 2.75 Å resolution crystal structure of bradyzoite enolase 1, the second structure to be reported of a bradyzoite-specific protein in Toxoplasma, captures an open conformational state and reveals that distinctive plant-like insertions are located on surface loops. The enolase 1 structure reveals that a unique residue, Glu164, in catalytic loop 2 may account for the lower activity of this cyst-stage isozyme. Recombinant TgENO1 specifically binds to a TTTTCT DNA motif present in the cyst matrix antigen 1 (TgMAG1) gene promoter as demonstrated by gel retardation. Furthermore, direct physical interactions of both nuclear TgENO1 and TgENO2 with the TgMAG1 gene promoter are demonstrated in vivo using chromatin immunoprecipitation (ChIP) assays. Structural and biochemical studies reveal that T. gondii enolase functions are multifaceted, including the coordination of gene regulation in parasitic stage development. Enolase 1 provides a potential lead in the design of drugs against Toxoplasma brain cysts

  8. Determining Concentration of Neurotrophic Factors and Neuron Specific Enolase in the Blood of Newborns with Central Nervous System Damages as a New Approach in Clinical Diagnostics

    Directory of Open Access Journals (Sweden)

    M.V. Vedunova

    2015-06-01

    Full Text Available The aim of the investigation is to assess the quantity of brain-derived neurotrophic factor (BDNF, glial cell line-derived neurotrophic factor (GDNF and neuron specific enolase (NSE in plasma of newborns with perinatal hypoxic damage of CNS. Materials and Methods. Neurotrophic factors and NSE enzyme concentrations in plasma of newborns (gestation age 31–42 weeks was studied. The main groups consisted of newborns with the symptoms of perinatal CNS damage (group 1 — with convulsive states, group 2 — with the signs of severe perinatal CNS damage, diagnosed according to physical examination, evaluation of the neurological status dynamics and neurosonographic studies. Control group included healthy neonates. Concentration of BDNF, GDNF (R&D Systems, USA and NSE enzyme (Vector Best, Russia was determined by ELISA kit during hospitalization and on day 10–14 after the rehabilitation therapy. Results. Carried out experiments revealed the significant increase of NSE concentration in plasma of newborns with convulsive states. The higher levels of this enzyme were detected in infants with severe perinatal CNS damage. Moreover, BDNF concentration significantly increases in plasma of patients with the symptoms of severe CNS damage in the period following rehabilitation therapy. These experiments also demonstrate the inverse correlation between BDNF and GDNF levels. It was shown the important prognostic value of BDNF and NSE determination in plasma of newborns with CNS injury. Conclusion. The most diagnostic value for assessing the severity of brain damage in early neonatal period is associated with measurements of NSE and BDNF concentrations in plasma, which allows to use these markers immediately after birth and before the development of neurological symptoms.

  9. Nuclear Glycolytic Enzyme Enolase of Toxoplasma gondii Functions as a Transcriptional Regulator

    OpenAIRE

    Thomas Mouveaux; Gabrielle Oria; Elisabeth Werkmeister; Christian Slomianny; Fox, Barbara A.; Bzik, David J.; Stanislas Tomavo

    2014-01-01

    Apicomplexan parasites including Toxoplasma gondii have complex life cycles within different hosts and their infectivity relies on their capacity to regulate gene expression. However, little is known about the nuclear factors that regulate gene expression in these pathogens. Here, we report that T. gondii enolase TgENO2 is targeted to the nucleus of actively replicating parasites, where it specifically binds to nuclear chromatin in vivo. Using a ChIP-Seq technique, we provide evidence for TgE...

  10. Serial measurement of neuron specific enolase improves prognostication in cardiac arrest patients treated with hypothermia: A prospective study

    Directory of Open Access Journals (Sweden)

    Storm Christian

    2012-01-01

    Full Text Available Abstract Background Neuron specific enolase (NSE has repeatedly been evaluated for neurological prognostication in patients after cardiac arrest. However, it is unclear whether current guidelines for NSE cutoff levels also apply to cardiac arrest patients treated with hypothermia. Thus, we investigated the prognostic significance of absolute NSE levels and NSE kinetics in cardiac arrest patients treated with hypothermia. Methods In a prospective study of 35 patients resuscitated from cardiac arrest, NSE was measured daily for four days following admission. Outcome was assessed at ICU discharge using the CPC score. All patients received hypothermia treatment for 24 hours at 33°C with a surface cooling device according to current guidelines. Results The cutoff for absolute NSE levels in patients with unfavourable outcome (CPC 3-5 72 hours after cardiac arrest was 57 μg/l with an area under the curve (AUC of 0.82 (sensitivity 47%, specificity 100%. The cutoff level for NSE kinetics in patients with unfavourable outcome (CPC 3-5 was an absolute increase of 7.9 μg/l (AUC 0.78, sensitivity 63%, specificity 100% and a relative increase of 33.1% (AUC 0.803, sensitivity 67%, specificity 100% at 48 hours compared to admission. Conclusion In cardiac arrest patients treated with hypothermia, prognostication of unfavourable outcome by NSE kinetics between admission and 48 hours after resuscitation may be superior to prognostication by absolute NSE levels.

  11. Significance of serum neuron-specific enolase in patients with acute traumatic brain injury

    Institute of Scientific and Technical Information of China (English)

    官卫; 杨伊林; 夏为民; 李璐; 龚德生

    2003-01-01

    Objective: To study the association between serum neuron-specific enolase (NSE) and the extent of brain damage and the outcome after acute traumatic brain injury (TBI). Methods: The release patterns of serum NSE in 78 patients after acute TBI were analyzed by using the enzyme linked immunosobent assay. The levels of NSE were compared with Glasgow coma scale, the category of brain injury and the outcome after 6 months of injury. Results: There were different NSE values in patients with minor (12.96 μg/L±2.39 μg/L), moderate (23.44 μg/L±5.33 μg/L) and severe brain injury (42.68 μg/L±4.57 μg/L). After severe TBI, the concentration of NSE in patients with epidural hematomas was 13.38 μg/L±4.01 μg/L, 24.03 μg/L±2.85 μg/L in brain contusion without surgical intervention group, 55.20 μg/L±6.35 μg/L in brain contusion with surgical intervention group, and 83.85 μg/L±15.82 μg/L in diffuse brain swelling group. There were close correlations between NSE values and Glasgow coma scale (r=-0.608, P<0.01) and the extent of brain injury (r=0.75, P<0.01). Patients with poor outcome had significantly higher initial and peak NSE values than those with good outcome (66.40 μg/L±9.46 μg/L, 94.24 μg/L±13.75 μg/L vs 32.16 μg/L±4.21 μg/L, 34.08 μg/L±4.40 μg/L, P<0.01, respectively). Initial NSE values were negatively related to the outcome (r=-0.501, P<0.01). Most patients with poor outcomes had persisting or secondary elevated NSE values. Conclusions: Serum NSE is one of the valuable neurobiochemical markers for assessment of the severity of brain injury and outcome prediction.

  12. A new nuclear function of the Entamoeba histolytica glycolytic enzyme enolase: the metabolic regulation of cytosine-5 methyltransferase 2 (Dnmt2 activity.

    Directory of Open Access Journals (Sweden)

    Ayala Tovy

    2010-02-01

    Full Text Available Cytosine-5 methyltransferases of the Dnmt2 family function as DNA and tRNA methyltransferases. Insight into the role and biological significance of Dnmt2 is greatly hampered by a lack of knowledge about its protein interactions. In this report, we address the subject of protein interaction by identifying enolase through a yeast two-hybrid screen as a Dnmt2-binding protein. Enolase, which is known to catalyze the conversion of 2-phosphoglycerate (2-PG to phosphoenolpyruvate (PEP, was shown to have both a cytoplasmatic and a nuclear localization in the parasite Entamoeba histolytica. We discovered that enolase acts as a Dnmt2 inhibitor. This unexpected inhibitory activity was antagonized by 2-PG, which suggests that glucose metabolism controls the non-glycolytic function of enolase. Interestingly, glucose starvation drives enolase to accumulate within the nucleus, which in turn leads to the formation of additional enolase-E.histolytica DNMT2 homolog (Ehmeth complex, and to a significant reduction of the tRNA(Asp methylation in the parasite. The crucial role of enolase as a Dnmt2 inhibitor was also demonstrated in E.histolytica expressing a nuclear localization signal (NLS-fused-enolase. These results establish enolase as the first Dnmt2 interacting protein, and highlight an unexpected role of a glycolytic enzyme in the modulation of Dnmt2 activity.

  13. RNase G-dependent degradation of the eno mRNA encoding a glycolysis enzyme enolase in Escherichia coli.

    Science.gov (United States)

    Kaga, Naoko; Umitsuki, Genryou; Nagai, Kazuo; Wachi, Masaaki

    2002-10-01

    Escherichia coli RNase G, encoded by the rng gene, is involved in the processing of 16S rRNA and degradation of the adhE mRNA encoding a fermentative alcohol dehydrogenase. In a search for the intracellular target RNAs of RNase G other than the 16S rRNA precursor and adhE mRNA, total cellular proteins from rng+ and rng::cat cells were compared by two-dimensional gel electrophoresis. The amount of enolase encoded by the eno gene reproducibly increased two- to three-fold in the rng::cat mutant strain compared with the rng+ parent strain. Rifampicin chase experiments showed that the half-life of the eno mRNA was some 3 times longer in the rng::cat mutant than in the wild type. These results indicate that the eno mRNA was a substrate of RNase G in vivo, in addition to 16S rRNA precursor and adhE mRNA. PMID:12450135

  14. Curcumin attenuates the middle cerebral artery occlusion-induced reduction in γ-enolase expression in an animal model.

    Science.gov (United States)

    Gim, Sang-Ah; Lee, So-Ra; Shah, Fawad-Ali; Koh, Phil-Ok

    2015-12-01

    Curcumin exerts a protective effect in cerebral ischemia through its anti-oxidant and anti-inflammatory activities. γ-enolase is a glycolytic enzyme expressed in neurons that is known to exerts a neuroprotective effect. We investigated whether curcumin regulates γ-enolase expression in focal cerebral ischemic injury in rats. Middle cerebral artery occlusion (MCAO) was performed to induce focal cerebral ischemia. Adult male rats were injected intraperitoneally with either vehicle or curcumin (50 mg/kg) 1 h after MCAO and cerebral cortex tissues were isolated 24 h after MCAO. We found that MCAO-induced injury resulted in a reduction in γ-enolase expression in vehicle-treated animals using a proteomics approach. However, this reduction was attenuated in animals with MCAO treated with curcumin. Reverse-transcription PCR and Western blot analyses also showed that curcumin treatment prevented the MCAO injury-induced reduction in γ-enolase expression. The results of this study suggest that curcumin exerts its neuroprotective function in focal cerebral ischemia by regulating the expression of γ-enolase. PMID:26755923

  15. Decreased levels of serum cytokeratin 19 fragment CYFRA 21-1 predict objective response to chemotherapy in patients with non-small cell lung cancer

    OpenAIRE

    Pang, Li; Jing WANG; Jiang, Yanwen; Chen, Liangan

    2013-01-01

    Diagnostic tools capable of predicting early responses to chemotherapy are required to improve the individual management of cancer patients. The present study aimed to evaluate the prognostic significance of the serum tumor markers CYFRA 21-1, carcinoembryonic antigen (CEA), neuron-specific enolase (NSE), carbohydrate antigen (CA) 125, and CA 19-9 for predicting responses to different chemotherapy regimens in patients with non-small cell lung cancer (NSCLC). A total of 276 patients with posto...

  16. Antibodies to soluble liver antigen and alpha-enolase in patients with autoimmune hepatitis.

    OpenAIRE

    Bogdanos, Dimitrios-Petrou; Gilbert, Daniele; Bianchi, Ilaria; Leoni, Simona; Mitry, Ragai; Ma, Yun; Mieli-vergani, Giorgina; Vergani, Diego

    2004-01-01

    BACKGROUND: Antibodies to a cytosolic soluble liver antigen (SLA) are specifically detected in patients with autoimmune hepatitis (AIH). The target of anti-SLA has been identified as a ~50 kDa UGA serine tRNA-associated protein complex (tRNP(Ser)Sec), through the screening of cDNA libraries. A recent report questioned the identity of tRNP(Ser)Sec as the real SLA antigen. The latter study identified alpha-enolase as a major anti-SLA target, through proteomic analysis. METHODS: In an attempt to...

  17. Release of erythropoietin and neuron-specific enolase after breath holding in competing free divers

    DEFF Research Database (Denmark)

    Kjeld, Thomas; Jattu, T; Nielsen, Henrik;

    2015-01-01

    Free diving is associated with extreme hypoxia. This study evaluated the combined effect of maximal static breath holding and underwater swimming on plasma biomarkers of tissue hypoxemia: erythropoietin, neuron-specific enolase and S100B, C-reactive protein, pro-atrial natriuretic peptide, and...... troponin T. Venous blood samples were obtained from 17 competing free divers before and 3 h after sessions of static apnea and underwater swimming. The heart was evaluated by echocardiography. Static apnea for 293 ± 78 s (mean ± SD) and subsequent 88 ± 21 m underwater swimming increased plasma...... = 0.549), and troponin T (P = 0.125) remained unchanged and, as assessed by echocardiography, the heart was not affected. In competitive free divers, bouts of static and dynamic apnea increase plasma erythropoietin and neuron-specific enolase, suggesting that renal and neural tissue, rather than the...

  18. Elevated serum neuron-specific enolase in patients with temporal lobe epilepsy: a video-EEG study.

    Science.gov (United States)

    Palmio, Johanna; Keränen, Tapani; Alapirtti, Tiina; Hulkkonen, Janne; Mäkinen, Riikka; Holm, Päivi; Suhonen, Jaana; Peltola, Jukka

    2008-10-01

    Established markers of brain damage, neuron-specific enolase (NSE) and S-100b protein (S-100), may increase after status epilepticus, but whether a single tonic-clonic or complex partial seizure induces elevation of these markers is not known. Furthermore, it is unclear whether the risk of seizure-related neuronal damage in temporal lobe epilepsy (TLE) differs from that in extratemporal lobe epilepsies (XTLE). The aim of this study was to analyze NSE and S-100 in patients with TLE and XTLE after acute seizures. The levels of NSE and S-100 were measured in serum before (0h) and at 3, 6, 12, and 24h after acute seizures in 31 patients during inpatient video-EEG monitoring. The patients were categorized into the TLE and the XTLE group based on video-EEG recordings and MRI findings. Fifteen patients had TLE and 16 XTLE. Index seizures were mainly complex partial seizures (n=21). In TLE mean+/-S.D. values for NSE levels (mug/L) were 8.36+/-2.64 (0h), 11.35+/-3.84 (3h), 13.48+/-4.49 (6h), 12.95+/-5.46 (12h) and 10.33+/-3.13 (24h) (p=0.006, ANOVA). In XTLE the changes were not significant (p=0.3). There was less increase in the levels of S-100 in TLE (p=0.05) and no significant change in XTLE (p=0.4). The levels of markers of neuronal damage were increased in patients with TLE, not only after tonic-clonic but also after complex partial seizures. These data suggest that TLE may be associated with brain damage. PMID:18595663

  19. Cardiac, renal, and neurological benefits of preoperative levosimendan administration in patients with right ventricular dysfunction and pulmonary hypertension undergoing cardiac surgery: evaluation with two biomarkers neutrophil gelatinase-associated lipocalin and neuronal enolase

    Science.gov (United States)

    Guerrero-Orriach, José Luis; Ariza-Villanueva, Daniel; Florez-Vela, Ana; Garrido-Sánchez, Lourdes; Moreno-Cortés, María Isabel; Galán-Ortega, Manuel; Ramírez-Fernández, Alicia; Alcaide Torres, Juan; Fernandez, Concepción Santiago; Navarro Arce, Isabel; Melero-Tejedor, José María; Rubio-Navarro, Manuel; Cruz-Mañas, José

    2016-01-01

    Purpose To evaluate if the preoperative administration of levosimendan in patients with right ventricular (RV) dysfunction, pulmonary hypertension, and high perioperative risk would improve cardiac function and would also have a protective effect on renal and neurological functions, assessed using two biomarkers neutrophil gelatinase-associated lipocalin (N-GAL) and neuronal enolase. Methods This is an observational study. Twenty-seven high-risk cardiac patients with RV dysfunction and pulmonary hypertension, scheduled for cardiac valve surgery, were prospectively followed after preoperative administration of levosimendan. Levosimendan was administered preoperatively on the day before surgery. All patients were considered high risk of cardiac and perioperative renal complications. Cardiac function was assessed by echocardiography, renal function by urinary N-GAL levels, and the acute kidney injury scale. Neuronal damage was assessed by neuron-specific enolase levels. Results After surgery, no significant variations were found in mean and SE levels of N-GAL (14.31 [28.34] ng/mL vs 13.41 [38.24] ng/mL), neuron-specific enolase (5.40 [0.41] ng/mL vs 4.32 [0.61] ng/mL), or mean ± SD creatinine (1.06±0.24 mg/dL vs 1.25±0.37 mg/dL at 48 hours). RV dilatation decreased from 4.23±0.7 mm to 3.45±0.6 mm and pulmonary artery pressure from 58±18 mmHg to 42±19 mmHg at 48 hours. Conclusion Preoperative administration of levosimendan has shown a protective role against cardiac, renal, and neurological damage in patients with a high risk of multiple organ dysfunctions undergoing cardiac surgery. PMID:27143905

  20. Probing bacterial adhesion at the single-cell level

    DEFF Research Database (Denmark)

    Zeng, Guanghong; Müller, Torsten; Meyer, Rikke Louise

    cantilever coated with the commercial cell adhesive CellTakTM. We applied the method to study adhesion of living cells to abiotic surfaces at the single-cell level. Immobilisation of single bacterial cells to the cantilever was stable for several hours, and viability was confirmed by Live/Dead staining and......Bacteria initiate attachment to surfaces with the aid of different extracellular proteins and polymeric adhesins. To quantitatively analyse the cell-cell and cell-surface interactions provided by bacterial adhesins, it is essential to go down to single cell level where cell-to-cell variation can be...... considered. We have developed a simple and versatile method to make single-cell bacterial probes for measuring single cell adhesion by force spectroscopy using atomic force microscopy (AFM). A single-cell probe was readily made by picking up a bacterial cell from a glass surface by approaching a tipless AFM...

  1. Molecular cloning of cDNA and analysis of protein secondary structure of Candida albicans enolase, an abundant, immunodominant glycolytic enzyme.

    OpenAIRE

    Sundstrom, P; Aliaga, G R

    1992-01-01

    We isolated and sequenced a clone for Candida albicans enolase from a C. albicans cDNA library by using molecular genetic techniques. The 1.4-kbp cDNA encoded one long open reading frame of 440 amino acids which was 87 and 75% similar to predicted enolases of Saccharomyces cerevisiae and enolases from other organisms, respectively. The cDNA included the entire coding region and predicted a protein of molecular weight 47,178. The codon usage was highly biased and similar to that found for the ...

  2. Mechanistic Diversity in the RuBisCO Superfamily: The Enolase in the Methionine Salvage Pathway in Geobacillus kaustophilus

    International Nuclear Information System (INIS)

    D-Ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO), the most abundant enzyme, is the paradigm member of the recently recognized mechanistically diverse RuBisCO superfamily. The RuBisCO reaction is initiated by abstraction of the proton from C3 of the D-ribulose 1,5-bisphosphate substrate by a carbamate oxygen of carboxylated Lys 201 (spinach enzyme). Heterofunctional homologues of RuBisCO found in species of Bacilli catalyze the tautomerization ('enolization') of 2,3-diketo-5-methylthiopentane 1-phosphate (DK-MTP 1-P) in the methionine salvage pathway in which 5-methylthio-D-ribose (MTR) derived from 5'-methylthioadenosine is converted to methionine (Ashida, H., Saito, Y., Kojima, C., Kobayashi, K., Ogasawara, N., and Yokota, A. (2003) A functional link between RuBisCO-like protein of Bacillus and photosynthetic RuBisCO, Science 302, 286-290]. The reaction catalyzed by this 'enolase' is accomplished by abstraction of a proton from C1 of the DK-MTP 1-P substrate to form the tautomerized product, a conjugated enol. Because the RuBisCO- and 'enolase'-catalyzed reactions differ in the regiochemistry of proton abstraction but are expected to share stabilization of an enolate anion intermediate by coordination to an active site Mg2+, we sought to establish structure-function relationships for the 'enolase' reaction so that the structural basis for the functional diversity could be established. We determined the stereochemical course of the reaction catalyzed by the 'enolases' from Bacillus subtilis and Geobacillus kaustophilus. Using stereospecifically deuterated samples of an alternate substrate derived from D-ribose (5-OH group instead of the 5-methylthio group in MTR) as well as of the natural DK-MTP 1-P substrate, we determined that the 'enolase'-catalyzed reaction involves abstraction of the 1-proS proton. We also determined the structure of the activated 'enolase' from G. kaustophilus (carboxylated on Lys 173) liganded with Mg2+ and 2,3-diketohexane 1

  3. Serum neuronal specific enolase as a biomarker in differentiating the side of brain lesion in acute hemorrhagic stroke: a hospital based study

    OpenAIRE

    Omkar Prasad Baidya; Sunita Tiwari; Kauser Usman

    2016-01-01

    Background: Neuronal specific Enolase (NSE) is the neuronal form of the glycolytic enzyme enolase. This study has been conducted to see the role of serum NSE in differentiating the side of brain lesion within 24 hours of acute hemorrhagic stroke onset. Methods: The study was conducted in collaboration with the Department of Physiology and Medicine after Ethical clearance from December 2013 to April 2015. Our study group consists of 35 acute hemorrhagic stroke patients (clinically and radio...

  4. Evolution of Enzymatic Activities in the Enolase Superfamily: L-Fuconate Dehydratase from Xanthomonas campestris

    Energy Technology Data Exchange (ETDEWEB)

    Yew,W.; Fedorov, A.; Fedorov, E.; Rakus, J.; Pierce, R.; Almo, S.; Gerlt, J.

    2006-01-01

    Many members of the mechanistically diverse enolase superfamily have unknown functions. In this report the authors use both genome (operon) context and screening of a library of acid sugars to assign the L-fuconate dehydratase (FucD) function to a member of the mandelate racemase (MR) subgroup of the superfamily encoded by the Xanthomonas campestris pv. campestris str. ATCC 33913 genome (GI: 21233491). Orthologues of FucD are found in both bacteria and eukaryotes, the latter including the rTS beta protein in Homo sapiens that has been implicated in regulating thymidylate synthase activity. As suggested by sequence alignments and confirmed by high-resolution structures in the presence of active site ligands, FucD and MR share the same active site motif of functional groups: three carboxylate ligands for the essential Mg2+ located at the ends of th third, fourth, and fifth-strands in the (/)7-barrel domain (Asp 248, Glu 274, and Glu 301, respectively), a Lys-x-Lys motif at the end of the second-strand (Lys 218 and Lys 220), a His-Asp dyad at the end of the seventh and sixth-strands (His 351 and Asp 324, respectively), and a Glue at the end of the eighth-strand (Glu 382). The mechanism of the FucD reaction involves initial abstraction of the 2-proton by Lys 220, acid catalysis of the vinylogous-elimination of the 3-OH group by His 351, and stereospecific ketonization of the resulting 2-keto-3-deoxy-L-fuconate product. Screening of the library of acid sugars revealed substrate and functional promiscuity: In addition to L-fuconate, FucD also catalyzes the dehydration of L-galactonate, D-arabinonate, D-altronate, L-talonate, and D-ribonate. The dehydrations of L-fuconate, L-galactonate, and D-arabinonate are initiated by abstraction of the 2-protons by Lys 220. The dehydrations of L-talonate and D-ribonate are initiated by abstraction of the 2-protons by His 351; however, protonation of the enediolate intermediates by the conjugate acid of Lys 220 yields L

  5. Glycation of the muscle-specific enolase by reactive carbonyls: effect of temperature and the protection role of carnosine, pyridoxamine and phosphatidylserine.

    Science.gov (United States)

    Pietkiewicz, Jadwiga; Bronowicka-Szydełko, Agnieszka; Dzierzba, Katarzyna; Danielewicz, Regina; Gamian, Andrzej

    2011-03-01

    Reactive carbonyls such as 4-hydroxy-2-nonenal (4-HNE), trans-2-nonenal (T2 N), acrolein (ACR) can react readily with nucleophilic protein sites forming of advanced glycation end-products (AGE). In this study, the human and pig muscle-specific enolase was used as a protein model for in vitro modification by 4-HNE, T2 N and ACR. While the human enolase interaction with reactive α-oxoaldehyde methylglyoxal (MOG) was demonstrated previously, the effect of 4-HNE, T2N and ACR has not been identified yet. Altering in catalytic function were observed after the enzyme incubation with these active compounds for 1-24 h at 25, 37 and 45 °C. The inhibition degree of enolase activity occurred in following order: 4-HNE > ACR > MOG > T2N and inactivation of pig muscle-specific enolase was more effective relatively to human enzyme. The efficiency of AGE formation depends on time and incubation temperature with glycating agent. More amounts of insoluble AGE were formed at 45 °C. We found that pyridoxamine and natural dipeptide carnosine counteracted AGE formation and protected enolase against the total loss of catalytic activity. Moreover, we demonstrated for the first time that phosphatidylserine may significantly protect enolase against decrease of catalytic activity in spite of AGE production. PMID:21347838

  6. Salivary neuron specific enolase: an indicator for neuronal damage in patients with ischemic stroke and stroke-prone patients

    OpenAIRE

    Al-Rawi, Natheer H.; Atiyah, Karim M.

    2009-01-01

    Background: The blood-brain barrier is compromised in patients with stroke. The release of neuro-biochemical protein markers, such as neuron specific enolase (NSE) into the circulation may allow the pathophysiology and prognosis of patients with cerebrovascular diseases to be evaluated further. The present study was designed to measure the marker of neuronal damage, NSE, in saliva and serum of patients with acute ischemic stroke and patients with stroke related diseases as a diagnostic and/or...

  7. Anti-α-enolase is a prognostic marker in postoperative lung cancer patients

    OpenAIRE

    Hsiao, Kuan-Chung; Shih, Neng-Yao; Chu, Pei-Yi; Hung, Yi-Mei; Liao, Jia-Yi; Chou, Shao-Wen; Yang, Yi-Yuan; Chang, Gee-Chen; Liu, Ko-Jiunn

    2015-01-01

    Our previous studies suggest that antibodies against ENO1 (anti-ENO1 Ab) have a protective role in patients with non-small cell lung carcinoma. In this study, we evaluated the prognostic value of anti-ENO1 Ab levels in non-small cell lung carcinoma patients undergoing surgery. Circulating levels of anti-ENO1 Ab were assessed in 85 non-small cell lung carcinoma patients before and after surgery, and were correlated with clinical outcome. After surgery, patients with a higher increase of anti-E...

  8. System-level design of bacterial cell cycle control

    OpenAIRE

    McAdams, Harley H.; Shapiro, Lucy

    2009-01-01

    Understanding of the cell cycle control logic in Caulobacter has progressed to the point where we now have an integrated view of the operation of an entire bacterial cell cycle system functioning as a state machine. Oscillating levels of a few temporally-controlled master regulator proteins in a cyclical circuit drive cell cycle progression. To a striking degree, the cell cycle regulation is a whole cell phenomenon. Phospho-signaling proteins and proteases dynamically deployed to specific loc...

  9. A new level set model for cell image segmentation

    Institute of Scientific and Technical Information of China (English)

    Ma Jing-Feng; Hou Kai; Bao Shang-Lian; Chen Chun

    2011-01-01

    In this paper we first determine three phases of cell images: background, cytoplasm and nucleolus according to the general physical characteristics of cell images, and then develop a variational model, based on these characteristics, to segment nucleolus and cytoplasm from their relatively complicated backgrounds. In the meantime, the preprocessing obtained information of cell images using the OTSU algorithm is used to initialize the level set function in the model, which can speed up the segmentation and present satisfactory results in cell image processing.

  10. Validation of an algorithm able to differentiate small-cell lung cancer (SCLC) from non-small-cell lung cancer (NSCLC) patients by means of a tumour marker panel: analysis of the errors.

    OpenAIRE

    Paone, G; De Angelis, G.; Portalone, L.; Greco, S.; Giosué, S.; Taglienti, A.; Bisetti, A; Ameglio, F.

    1997-01-01

    By means of a mathematical score previously generated by discriminant analysis on 90 lung cancer patients, a new and larger group of 261 subjects [209 with non-small-cell lung cancer (NSCLC) and 52 with small-cell lung cancer (SCLC)] was analysed to confirm the ability of the method to distinguish between these two types of cancers. The score, which included the serum neuron-specific enolase (NSE) and CYFRA-21.1 levels, permitted correct classification of 93% of the patients. When the misclas...

  11. Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

    Directory of Open Access Journals (Sweden)

    Facincani Agda Paula

    2003-01-01

    Full Text Available The objective of this work was to assess the functionality of the glycolytic pathways in the bacterium Xylella fastidiosa. To this effect, the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase of the glycolytic pathway, and glucose 6-phosphate dehydrogenase of the Entner-Doudoroff pathway were studied, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates, which explains the increased duplication time of this phytopatogen. Recombinant enolase was expressed as inclusion bodies and solubilized with urea (most efficient extractor, Triton X-100, and TCA. Enolase extracted from X. fastidiosa and from chicken muscle and liver is irreversibly inactivated by urea. The purification of enolase was partial and resulted in a low yield. No enzymatic activity was detected for either recombinant and native enolases, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, suggesting that X. fastidiosa uses the Entner-Doudoroff pathway to produce pyruvate. Evidence is presented supporting the idea that the regulation of genes and the presence of isoforms with regulation patterns might make it difficult to understand the metabolism of carbohydrates in X. fastidiosa.

  12. System-level modeling and simulation of the cell culture microfluidic biochip ProCell

    DEFF Research Database (Denmark)

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan;

    2010-01-01

    -defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chip. ProCell contains a cell culture chamber, which is envisioned to run 256 simultaneous experiments (viewed as a...

  13. Alpha contamination levels in SMF south cell and compartments

    International Nuclear Information System (INIS)

    This document describes the detailed contamination survey performed in the Shielded Materials Facility (SMF) South Cell and the four compartments used during the CsCl activities. Smears were obtained at each operating station in South Cell and analyzed at the 325 Building. The smear results indicate that the highest contamination levels are in Compartment 1 and South Cell proper, with significantly lower contamination levels measured in the other three compartments. Although some of the smears indicated the presence of alpha contamination, it will be shown that the source of the alpha contamination was cross-contamination during processing in the 325 Building hot cells and that the SMF is free of alpha contamination. The alpha-free status of South Cell is consistent with process knowledge of previous South Cell activities

  14. Enolase: A Key Player in the Metabolism and a Probable Virulence Factor of Trypanosomatid Parasites—Perspectives for Its Use as a Therapeutic Target

    Directory of Open Access Journals (Sweden)

    Luisana Avilán

    2011-01-01

    Full Text Available Glycolysis and glyconeogenesis play crucial roles in the ATP supply and synthesis of glycoconjugates, important for the viability and virulence, respectively, of the human-pathogenic stages of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania spp. These pathways are, therefore, candidate targets for antiparasite drugs. The glycolytic/gluconeogenic enzyme enolase is generally highly conserved, with similar overall fold and identical catalytic residues in all organisms. Nonetheless, potentially important differences exist between the trypanosomatid and host enzymes, with three unique, reactive residues close to the active site of the former that might be exploited for the development of new drugs. In addition, enolase is found both in the secretome and in association with the surface of Leishmania spp. where it probably functions as plasminogen receptor, playing a role in the parasite’s invasiveness and virulence, a function possibly also present in the other trypanosomatids. This location and possible function of enolase offer additional perspectives for both drug discovery and vaccination.

  15. Flunarizine and lamotngine propnyiaxis effects on neuron-specific enolase,S-100,and brain-specific creatine kinase in a fetal rat model of hypoxic-ischemic brain damage

    Institute of Scientific and Technical Information of China (English)

    Li He; Jingyi Deng; Wendan He

    2008-01-01

    BACKGROUND:Calcium antagonists may act as neuroprotectants,diminishing the influx of calcium ions through voltage-sensitive calcium channels. When administered prophylactically,they display neuroprotective effects against hypoxic-ischemic brain damage in newborn rats.OBJECTIVE:To investigate the neuroprotective effects of flunarizine(FNZ),lamotrigine (LTG)and the combination of both drugs,on hypoxic-ischemic brain damage in fetal rats.DESIGN AND SETTING:This randomized,complete block design was performed at the Department of Pediatrics.Shenzhen Fourth People's Hospital,Guangdong Medical College.MATERIALS:Forty pregnant Wistar rats,at gestational day 20,were selected for the experiment and were randomly divided into FNZ,LTG,FNZ+LTG,and model groups,with 10 rats in each group.METHODS:Rats in the FNZ.LTG,and FNZ+LTG groups received intragastric injections of FNZ (0.5 mg/kg/d),LTG(10 mg/kg/d),and FNZ(0.5 mg/kg/d)+LTG(10 mg/kg/d),respectively.Drugs were administered once a day for 3 days prior to induction of hypoxia-ischemia.Rats in the modeJ group were not administered any drugs.Three hours after the final administration,eight pregnant rats from each group underwent model establishment hypoxia-ischemia brain damage to the fetal rats.Cesareans were performed at 6,12,24,and 48 hours later;and 5 fetal rats were removed from each mother and kept warm.Twe fetuses without model establishment were removed by planned cesarean at the same time and served as controls.A total of 0.3 mL serum was collected from fetal rats at 6,12,24,and 48 hours,respectively,following birth.MAIN OUTCOME MEASURES:Serum protein concentrations of neuron-specific enolase and S-100 were measured by ELISA.Serum concentrations of brain-specific creatine kinase were measured using an electrogenerated chemiluminescence method.RESULTS:Serum concentrations of neuron-specific enolase,S-100,and brain-specific creatine kinase were significantly higher in the hypoxic-ischemic fetal rats.compared with the non

  16. Evaluating the prognosis and degree of brain injury by combined S-100 protein and neuron specific enolase determination

    Institute of Scientific and Technical Information of China (English)

    Xihua Wang; Xinding Zhang

    2006-01-01

    Background:S-100 and neuron specific enolase(NSE)possess the characteristics of specific distribution in brain and relative stable content.Some studies suggest that combined detection of the both is of very importance for evaluating the degree of brain injury.OBJECTIVE: To observe the changes of S-100 protein and NSE levels at different time points after acute brain injury,and evaluate the values of combined detection detection of the both for different injury degrees,pathological changes and prognosis.DESIGN: Case-control observation SETTING: Department of Neurosurgery,Second Affiliated Hospital,Lanzhou University.PARTICIPANTS:Thirty-four inpatients with brain injury,19 males and 15 females,aged 15 to 73 years.who received treatment between September 2005 and May 2006 in the Department of Neurosurgery. Second Affiliated Hospital,Lanzhou University,were recruited.The patients were admitted to hospital at 24 hours after brain injury.After admission,skull CT confirmed that they suffered from brain injury.Following Glasgow coma score(GCS)on admission,the patients were assigned into 3 groups:severe group(GCS 3 to 8 points,n=15).moderate group(GCS 9 to 12 points,n=8)and mild group(GCS 13 to 15 points,n=11).Following Glasgow outcome scale(GOS)at 3 months after brain injury,the patients were assigned into good outcome group (GOS 4 to 5 points,good recovery and moderate disability included,n=19)and poor outcome group(GOS 1 to 3 points,severe disability,vegetative state and death,n=15).Ten subjects who received health examination concurrently were chosen as normal control group,including 6 males and 4 females,aged(45.4±14.3)years.In our laboratory,the normal level of NSE was≤15.2 ng/L,and that of S100 was≤0.105 μg/L.METHODS:①Blood samples of control group were collected when the subjects received health examination Blood samples of patients with brain injury were collected at 24 hours,3,7 and 14 days after injury.According to the instructions of NSE and S-100 kits

  17. A new level set model for cell image segmentation

    International Nuclear Information System (INIS)

    In this paper we first determine three phases of cell images: background, cytoplasm and nucleolus according to the general physical characteristics of cell images, and then develop a variational model, based on these characteristics, to segment nucleolus and cytoplasm from their relatively complicated backgrounds. In the meantime, the preprocessing obtained information of cell images using the OTSU algorithm is used to initialize the level set function in the model, which can speed up the segmentation and present satisfactory results in cell image processing. (cross-disciplinary physics and related areas of science and technology)

  18. Systems Level Modeling of the Cell Cycle Using Budding Yeast

    Directory of Open Access Journals (Sweden)

    D.R. Kim

    2007-01-01

    Full Text Available Proteins involved in the regulation of the cell cycle are highly conserved across all eukaryotes, and so a relatively simple eukaryote such as yeast can provide insight into a variety of cell cycle perturbations including those that occur in human cancer. To date, the budding yeast Saccharomyces cerevisiae has provided the largest amount of experimental and modeling data on the progression of the cell cycle, making it a logical choice for in-depth studies of this process. Moreover, the advent of methods for collection of high-throughput genome, transcriptome, and proteome data has provided a means to collect and precisely quantify simultaneous cell cycle gene transcript and protein levels, permitting modeling of the cell cycle on the systems level. With the appropriate mathematical framework and suffi cient and accurate data on cell cycle components, it should be possible to create a model of the cell cycle that not only effectively describes its operation, but can also predict responses to perturbations such as variation in protein levels and responses to external stimuli including targeted inhibition by drugs. In this review, we summarize existing data on the yeast cell cycle, proteomics technologies for quantifying cell cycle proteins, and the mathematical frameworks that can integrate this data into representative and effective models. Systems level modeling of the cell cycle will require the integration of high-quality data with the appropriate mathematical framework, which can currently be attained through the combination of dynamic modeling based on proteomics data and using yeast as a model organism.

  19. High epitope expression levels increase competition between T cells.

    Directory of Open Access Journals (Sweden)

    Almut Scherer

    2006-08-01

    Full Text Available Both theoretical predictions and experimental findings suggest that T cell populations can compete with each other. There is some debate on whether T cells compete for aspecific stimuli, such as access to the surface on antigen-presenting cells (APCs or for specific stimuli, such as their cognate epitope ligand. We have developed an individual-based computer simulation model to study T cell competition. Our model shows that the expression level of foreign epitopes per APC determines whether T cell competition is mainly for specific or aspecific stimuli. Under low epitope expression, competition is mainly for the specific epitope stimuli, and, hence, different epitope-specific T cell populations coexist readily. However, if epitope expression levels are high, aspecific competition becomes more important. Such between-specificity competition can lead to competitive exclusion between different epitope-specific T cell populations. Our model allows us to delineate the circumstances that facilitate coexistence of T cells of different epitope specificity. Understanding mechanisms of T cell coexistence has important practical implications for immune therapies that require a broad immune response.

  20. Serum Neuron-Specific Enolase, Biogenic Amino-Acids and Neurobehavioral Function in Lead-Exposed Workers from Lead-Acid Battery Manufacturing Process

    Directory of Open Access Journals (Sweden)

    K Ravibabu

    2015-01-01

    Full Text Available Background: The interaction between serum neuron-specific enolase (NSE, biogenic amino-acids and neurobehavioral function with blood lead levels in workers exposed to lead form lead-acid battery manufacturing process was not studied.Objective: To evaluate serum NSE and biogenic amino-acids (dopamine and serotonin levels, and neurobehavioral performance among workers exposed to lead from lead-acid storage battery plant, and its relation with blood lead levels (BLLs.Methods: In a cross-sectional study, we performed biochemical and neurobehavioral function tests on 146 workers exposed to lead from lead-acid battery manufacturing process. BLLs were assessed by an atomic absorption spectrophotometer. Serum NSE, dopamine and serotonin were measured by ELISA. Neurobehavioral functions were assessed by CDC-recommended tests—simple reaction time (SRT, symbol digit substitution test (SDST, and serial digit learning test (SDLT.Results: There was a significant correlation (r 0.199, p<0.05 between SDST and BLL. SDLT and SRT had also a significant positive correlation (r 0.238, p<0.01. NSE had a negative correlation (r –0.194, p<0.05 with serotonin level. Multiple linear regression analysis revealed that both SRT and SDST had positive significant associations with BLL. SRT also had a positive significant association with age.Conclusion: Serum NSE cannot be used as a marker for BLL. The only domain of neurobehavioral function tests that is affected by increased BLL in workers of lead-acid battery manufacturing process is that of the “attention and perception” (SDST.

  1. [Benign and malignant granular cell tumors. An immunohistochemical classification of tumor cells].

    Science.gov (United States)

    Kuhn, A; Mahrle, G; Steigleder, G K

    1987-06-15

    Eight benign and three malignant granular cell tumors were characterized by means of antibodies and antisera against keratin, desmin, epithelial membrane antigen, factor VIII-related protein, lysozyme, myelin basic protein, myoglobin, neurone-specific enolase, S 100 protein, myelin-associated protein (Leu 7), glial fibrillary acidic protein, vimentin, and neurofilament. All benign granular cell tumours showed positive staining of the tumor cells to antibodies against vimentin, S 100 protein, and neurone-specific enolase; myelin-associated protein (Leu 7), in contrast, was only detectable in a few tumor sections. Histogenetically the granular cells may be classified as Schwann's cells which lost their expression of laminin. The three malignant granular cell tumors showed a staining pattern significantly different from that of the benign tumours. Thus, only neurone-specific enolase was detectable in all the tumors, whereas S 100 protein and vimentin could not be demonstrated but in one and two, resp., out of three tumors. PMID:3303714

  2. NSE与视频脑电图在儿童热性惊厥的相关研究%THE CORRELATION BETWEEN SERUM NEURON SPECIFIC ENOLASE AND VIDEO EEG IN FEBRILE CONVULSION OF CHILDREN

    Institute of Scientific and Technical Information of China (English)

    刘秀琴; 于树红; 孙若鹏

    2002-01-01

    Objective:To evaluate the diagnostic value of serum neuron specific enolase and Video EEG in Febrile Convulsion of childen. Method:Serum NSE was detected by RIA on the first day and the seventh day after seizure in 40 children with simple febrile convulsion and 18 with complex febrile convulsion. Video EEG was performed at 1st, 7th and 30th day in all the patients. Results: There were significant differences between NSE levels at 24th hour and on 7th day after convulsion (P<0.01). NSE concentrations in patients with SFC and CFC were also different (P < 0.01). The more slowly NSE restored, the higher the abnormal ratio of EGG was . Conclusion: NSE increased in patients within 24hours after seizure, especially in CFC patients. The abnormal ratio of VideoEEG was related to the change of NSE concentration.

  3. Chitosan-collagen porous scaffold and bone marrow mesenchymal stem cell transplantation for ischemic stroke

    Institute of Scientific and Technical Information of China (English)

    Feng Yan; Wei Yue; Yue-lin Zhang; Guo-chao Mao; Ke Gao; Zhen-xing Zuo; Ya-jing Zhang; Hui Lu

    2015-01-01

    In this study, we successfully constructed a composite of bone marrow mesenchymal stem cells and a chitosan-collagen scaffoldin vitro, transplanted either the composite or bone marrow mesenchymal stem cells alone into the ischemic area in animal models, and compared their effects. At 14 days after co-transplantation of bone marrow mesenchymal stem cells and the hi-tosan-collagen scaffold, neurological function recovered noticeably. Vascular endothelial growth factor expression and nestin-labeled neural precursor cells were detected in the ischemic area, surrounding tissue, hippocampal dentate gyrus and subventricular zone. Simultaneously, a high level of expression of glial ifbrillary acidic protein and a low level of expression of neuron-spe-ciifc enolase were visible in BrdU-labeled bone marrow mesenchymal stem cells. These ifndings suggest that transplantation of a composite of bone marrow mesenchymal stem cells and a chi-tosan-collagen scaffold has a neuroprotective effect following ischemic stroke.

  4. Chitosan-collagen porous scaffold and bone marrow mesenchymal stem cell transplantation for ischemic stroke

    Directory of Open Access Journals (Sweden)

    Feng Yan

    2015-01-01

    Full Text Available In this study, we successfully constructed a composite of bone marrow mesenchymal stem cells and a chitosan-collagen scaffold in vitro, transplanted either the composite or bone marrow mesenchymal stem cells alone into the ischemic area in animal models, and compared their effects. At 14 days after co-transplantation of bone marrow mesenchymal stem cells and the hitosan-collagen scaffold, neurological function recovered noticeably. Vascular endothelial growth factor expression and nestin-labeled neural precursor cells were detected in the ischemic area, surrounding tissue, hippocampal dentate gyrus and subventricular zone. Simultaneously, a high level of expression of glial fibrillary acidic protein and a low level of expression of neuron-specific enolase were visible in BrdU-labeled bone marrow mesenchymal stem cells. These findings suggest that transplantation of a composite of bone marrow mesenchymal stem cells and a chitosan-collagen scaffold has a neuroprotective effect following ischemic stroke.

  5. Comparison of Serum MicroRNA21 and Tumor Markers in Diagnosis of Early Non-Small Cell Lung Cancer

    OpenAIRE

    Mingzhong Sun; Jiangxiang Song; Zhongwei Zhou; Rong Zhu; Hao Jin; Yuqiao Ji; Qiang Lu; Huixiang Ju

    2016-01-01

    Objective. To compare the clinical value of serum microRNA21 (miR21) and other tumor markers in early diagnosis of non-small cell lung cancer (NSCLC). Methods. Serums carcinoembryonic antigen (CEA), cytokeratin 19 fragment (CYFRA21-1), neuron-specific enolase (NSE), and miR21 were detected in 50 NSCLC cases and 60 healthy control individuals. Results. Average serums miR21, CEA, NSE, and CYFRA21-1 levels were significantly higher in the case group than in control group (P < 0.01). Analysis of ...

  6. Human mast cells decrease SLPI levels in type II – like alveolar cell model, in vitro

    Directory of Open Access Journals (Sweden)

    Nyström Max

    2003-08-01

    Full Text Available Abstract Background Mast cells are known to accumulate at sites of inflammation and upon activation to release their granule content, e.g. histamine, cytokines and proteases. The secretory leukocyte protease inhibitor (SLPI is produced in the respiratory mucous and plays a role in regulating the activity of the proteases. Result We have used the HMC-1 cell line as a model for human mast cells to investigate their effect on SLPI expression and its levels in cell co-culture experiments, in vitro. In comparison with controls, we found a significant reduction in SLPI levels (by 2.35-fold, p Conclusion These results indicate that SLPI-producing cells may assist mast cell migration and that the regulation of SLPI release and/or consumption by mast cells requires interaction between these cell types. Therefore, a "local relationship" between mast cells and airway epithelial cells might be an important step in the inflammatory response.

  7. MicroRNA-9 promotes the neuronal differentiation of rat bone marrow mesenchymal stem cells by activating autophagy

    Directory of Open Access Journals (Sweden)

    Guang-yu Zhang

    2015-01-01

    Full Text Available MicroRNA-9 (miR-9 has been shown to promote the differentiation of bone marrow mesenchymal stem cells into neuronal cells, but the precise mechanism is unclear. Our previous study confirmed that increased autophagic activity improved the efficiency of neuronal differentiation in bone marrow mesenchymal stem cells. Accumulating evidence reveals that miRNAs adjust the autophagic pathways. This study used miR-9-1 lentiviral vector and miR-9-1 inhibitor to modulate the expression level of miR-9. Autophagic activity and neuronal differentiation were measured by the number of light chain-3 (LC3-positive dots, the ratio of LC3-II/LC3, and the expression levels of the neuronal markers enolase and microtubule-associated protein 2. Results showed that LC3-positive dots, the ratio of LC3-II/LC3, and expression of neuron specific enolase and microtubule-associated protein 2 increased in the miR-9 + group. The above results suggest that autophagic activity increased and bone marrow mesenchymal stem cells were prone to differentiate into neuronal cells when miR-9 was overexpressed, demonstrating that miR-9 can promote neuronal differentiation by increasing autophagic activity.

  8. MicroRNA-9 promotes the neuronal differentiation of rat bone marrow mesenchymal stem cells by activating autophagy

    Institute of Scientific and Technical Information of China (English)

    Guang-yu Zhang; Jun Wang; Yan-jie Jia; Rui Han; Ping Li; Deng-na Zhu

    2015-01-01

    MicroRNA-9 (miR-9) has been shown to promote the differentiation of bone marrow mesen-chymal stem cells into neuronal cells, but the precise mechanism is unclear. Our previous study conifrmed that increased autophagic activity improved the efifciency of neuronal differentiation in bone marrow mesenchymal stem cells. Accumulating evidence reveals that miRNAs adjust the autophagic pathways. This study used miR-9-1 lentiviral vector and miR-9-1 inhibitor to modulate the expression level of miR-9. Autophagic activity and neuronal differentiation were measured by the number of light chain-3 (LC3)-positive dots, the ratio of LC3-II/LC3, and the expression levels of the neuronal markers enolase and microtubule-associated protein 2. Re-sults showed that LC3-positive dots, the ratio of LC3-II/LC3, and expression of neuron speciifc enolase and microtubule-associated protein 2 increased in the miR-9+ group. The above results suggest that autophagic activity increased and bone marrow mesenchymal stem cells were prone to differentiate into neuronal cells when miR-9 was overexpressed, demonstrating that miR-9 can promote neuronal differentiation by increasing autophagic activity.

  9. Soluble CD163 levels in children with sickle cell disease

    DEFF Research Database (Denmark)

    Moller, Holger Jon; Nielsen, Marianne Jensby; Bartram, Jack;

    2011-01-01

    Sickle cell disease (SCD) is characterized by vasculopathy, which has been causally linked to intravascular haemolysis and high levels of free plasma haemoglobin. Soluble CD163 (sCD163) is implicated in the clearance of free plasma haemoglobin and high plasma concentrations have been linked to...... arterial disease. We therefore investigated the value of sCD163 as a biomarker in children with SCD, and also measured haptoglobin levels in this population. We measured sCD163 in 25 control children with no haemoglobinopathy, 41 with sickle cell anaemia (HbSS) in the steady state, 27 with HbSS taking...

  10. Cancer Cell Analyses at the Single Cell-Level Using Electroactive Microwell Array Device.

    Directory of Open Access Journals (Sweden)

    Marina Kobayashi

    Full Text Available Circulating tumor cells (CTCs, shed from primary tumors and disseminated into peripheral blood, are playing a major role in metastasis. Even after isolation of CTCs from blood, the target cells are mixed with a population of other cell types. Here, we propose a new method for analyses of cell mixture at the single-cell level using a microfluidic device that contains arrayed electroactive microwells. Dielectrophoretic (DEP force, induced by the electrodes patterned on the bottom surface of the microwells, allows efficient trapping and stable positioning of single cells for high-throughput biochemical analyses. We demonstrated that various on-chip analyses including immunostaining, viability/apoptosis assay and fluorescent in situ hybridization (FISH at the single-cell level could be conducted just by applying specific reagents for each assay. Our simple method should greatly help discrimination and analysis of rare cancer cells among a population of blood cells.

  11. Phosphatidylserine increases IKBKAP levels in familial dysautonomia cells.

    Directory of Open Access Journals (Sweden)

    Hadas Keren

    Full Text Available Familial Dysautonomia (FD is an autosomal recessive congenital neuropathy that results from abnormal development and progressive degeneration of the sensory and autonomic nervous system. The mutation observed in almost all FD patients is a point mutation at position 6 of intron 20 of the IKBKAP gene; this gene encodes the IκB kinase complex-associated protein (IKAP. The mutation results in a tissue-specific splicing defect: Exon 20 is skipped, leading to reduced IKAP protein expression. Here we show that phosphatidylserine (PS, an FDA-approved food supplement, increased IKAP mRNA levels in cells derived from FD patients. Long-term treatment with PS led to a significant increase in IKAP protein levels in these cells. A conjugate of PS and an omega-3 fatty acid also increased IKAP mRNA levels. Furthermore, PS treatment released FD cells from cell cycle arrest and up-regulated a significant number of genes involved in cell cycle regulation. Our results suggest that PS has potential for use as a therapeutic agent for FD. Understanding its mechanism of action may reveal the mechanism underlying the FD disease.

  12. Levels of immune cells in transcendental meditation practitioners

    Directory of Open Access Journals (Sweden)

    Jose R Infante

    2014-01-01

    Conclusions: The technique of meditation studied seems to have a significant effect on immune cells, manifesting in the different circulating levels of lymphocyte subsets analyzed. The significant effect of TM on the neuroendocrine axis and its relationship with the immune system may partly explain our results.

  13. Quantification of Protein Levels in Single Living Cells

    Directory of Open Access Journals (Sweden)

    Chiu-An Lo

    2015-12-01

    Full Text Available Accurate measurement of the amount of specific protein a cell produces is important for investigating basic molecular processes. We have developed a technique that allows for quantitation of protein levels in single cells in vivo. This protein quantitation ratioing (PQR technique uses a genetic tag that produces a stoichiometric ratio of a fluorescent protein reporter and the protein of interest during protein translation. The fluorescence intensity is proportional to the number of molecules produced of the protein of interest and is used to determine the relative amount of protein within the cell. We use PQR to quantify protein expression of different genes using quantitative imaging, electrophysiology, and phenotype. We use genome editing to insert Protein Quantitation Reporters into endogenous genomic loci in three different genomes for quantitation of endogenous protein levels. The PQR technique will allow for a wide range of quantitative experiments examining gene-to-phenotype relationships with greater accuracy.

  14. Intraepidermal neuron-specific enolase (NSE)-immunoreactive nerve fibres: evidence for sprouting in uremic patients on maintenance hemodialysis.

    Science.gov (United States)

    Johansson, O; Hilliges, M; Ståhle-Bäckdahl, M

    1989-05-01

    The use of indirect immunohistochemistry in 12 patients on maintenance hemodialysis has shown weak or moderately strong neuron-specific enolase (NSE)-immunoreactive nerve terminals and fibres sprouting throughout the layers of the epidermis. No such terminals or fibres were found in any of 15 controls. There was no difference between uremic patients with pruritus and those without. Furthermore, NSE-positive nerve fibres with a normal appearance were seen in the dermis, at the epidermal-dermal junctional zone and sometimes entering the stratum basale in both patients and controls. The immunoreactive nerves were thin, smooth and, at their terminal fields, varicose. The immunoreactivity seemed to be associated chiefly with sensory nerves. Thus, our results suggest that uremic patients undergoing maintenance hemodialysis develop an abnormal pattern of cutaneous innervation. PMID:2657508

  15. Inactive enzymatic mutant proteins (phosphoglycerate mutase and enolase as sugar binders for ribulose-1,5-bisphosphate regeneration reactors

    Directory of Open Access Journals (Sweden)

    Giri Ashok

    2005-02-01

    Full Text Available Abstract Background Carbon dioxide fixation bioprocess in reactors necessitates recycling of D-ribulose1,5-bisphosphate (RuBP for continuous operation. A radically new close loop of RuBP regenerating reactor design has been proposed that will harbor enzyme-complexes instead of purified enzymes. These reactors will need binders enabling selective capture and release of sugar and intermediate metabolites enabling specific conversions during regeneration. In the current manuscript we describe properties of proteins that will act as potential binders in RuBP regeneration reactors. Results We demonstrate specific binding of 3-phosphoglycerate (3PGA and 3-phosphoglyceraldehyde (3PGAL from sugar mixtures by inactive mutant of yeast enzymes phosphoglycerate mutase and enolase. The reversibility in binding with respect to pH and EDTA has also been shown. No chemical conversion of incubated sugars or sugar intermediate metabolites were found by the inactive enzymatic proteins. The dissociation constants for sugar metabolites are in the micromolar range, both proteins showed lower dissociation constant (Kd for 3-phosphoglycerate (655–796 μM compared to 3-phosphoglyceraldehyde (822–966 μM indicating higher affinity for 3PGA. The proteins did not show binding to glucose, sucrose or fructose within the sensitivity limits of detection. Phosphoglycerate mutase showed slightly lower stability on repeated use than enolase mutants. Conclusions The sugar and their intermediate metabolite binders may have a useful role in RuBP regeneration reactors. The reversibility of binding with respect to changes in physicochemical factors and stability when subjected to repeated changes in these conditions are expected to make the mutant proteins candidates for in-situ removal of sugar intermediate metabolites for forward driving of specific reactions in enzyme-complex reactors.

  16. Cell Pluripotency Levels Associated with Imprinted Genes in Human

    Directory of Open Access Journals (Sweden)

    Liyun Yuan

    2015-01-01

    Full Text Available Pluripotent stem cells are exhibited similarly in the morphology, gene expression, growth properties, and epigenetic modification with embryonic stem cells (ESCs. However, it is still controversial that the pluripotency of induced pluripotent stem cell (iPSC is much inferior to ESC, and the differentiation capacity of iPSC and ESC can also be separated by transcriptome and epigenetics. miRNAs, which act in posttranscriptional regulation of gene expression and are involved in many basic cellular processes, may reveal the answer. In this paper, we focused on identifying the hidden relationship between miRNAs and imprinted genes in cell pluripotency. Total miRNA expression patterns in iPSC and ES cells were comprehensively analysed and linked with human imprinted genes, which show a global picture of their potential function in pluripotent level. A new CPA4-KLF14 region which locates in chromosomal homologous segments (CHSs within mammals and include both imprinted genes and significantly expressed miRNAs was first identified. Molecular network analysis showed genes interacted with imprinted genes closely and enriched in modules such as cancer, cell death and survival, and tumor morphology. This imprinted region may provide a new look for those who are interested in cell pluripotency of hiPSCs and hESCs.

  17. Cell Pluripotency Levels Associated with Imprinted Genes in Human.

    Science.gov (United States)

    Yuan, Liyun; Tang, Xiaoyan; Zhang, Binyan; Ding, Guohui

    2015-01-01

    Pluripotent stem cells are exhibited similarly in the morphology, gene expression, growth properties, and epigenetic modification with embryonic stem cells (ESCs). However, it is still controversial that the pluripotency of induced pluripotent stem cell (iPSC) is much inferior to ESC, and the differentiation capacity of iPSC and ESC can also be separated by transcriptome and epigenetics. miRNAs, which act in posttranscriptional regulation of gene expression and are involved in many basic cellular processes, may reveal the answer. In this paper, we focused on identifying the hidden relationship between miRNAs and imprinted genes in cell pluripotency. Total miRNA expression patterns in iPSC and ES cells were comprehensively analysed and linked with human imprinted genes, which show a global picture of their potential function in pluripotent level. A new CPA4-KLF14 region which locates in chromosomal homologous segments (CHSs) within mammals and include both imprinted genes and significantly expressed miRNAs was first identified. Molecular network analysis showed genes interacted with imprinted genes closely and enriched in modules such as cancer, cell death and survival, and tumor morphology. This imprinted region may provide a new look for those who are interested in cell pluripotency of hiPSCs and hESCs. PMID:26504487

  18. Multiple cell and population-level interactions with mouse embryonic stem cell heterogeneity.

    Science.gov (United States)

    Cannon, Danielle; Corrigan, Adam M; Miermont, Agnes; McDonel, Patrick; Chubb, Jonathan R

    2015-08-15

    Much of development and disease concerns the generation of gene expression differences between related cells sharing similar niches. However, most analyses of gene expression only assess population and time-averaged levels of steady-state transcription. The mechanisms driving differentiation are buried within snapshots of the average cell, lacking dynamic information and the diverse regulatory history experienced by individual cells. Here, we use a quantitative imaging platform with large time series data sets to determine the regulation of developmental gene expression by cell cycle, lineage, motility and environment. We apply this technology to the regulation of the pluripotency gene Nanog in mouse embryonic stem cells. Our data reveal the diversity of cell and population-level interactions with Nanog dynamics and heterogeneity, and how this regulation responds to triggers of pluripotency. Cell cycles are highly heterogeneous and cycle time increases with Nanog reporter expression, with longer, more variable cycle times as cells approach ground-state pluripotency. Nanog reporter expression is highly stable over multiple cell generations, with fluctuations within cycles confined by an attractor state. Modelling reveals an environmental component to expression stability, in addition to any cell-autonomous behaviour, and we identify interactions of cell density with both cycle behaviour and Nanog. Rex1 expression dynamics showed shared and distinct regulatory effects. Overall, our observations of multiple partially overlapping dynamic heterogeneities imply complex cell and environmental regulation of pluripotent cell behaviour, and suggest simple deterministic views of stem cell states are inappropriate. PMID:26209649

  19. Sorting of cells of the same size, shape, and cell cycle stage for a single cell level assay without staining

    Directory of Open Access Journals (Sweden)

    Yomo Tetsuya

    2006-06-01

    Full Text Available Abstract Background Single-cell level studies are being used increasingly to measure cell properties not directly observable in a cell population. High-performance data acquisition systems for such studies have, by necessity, developed in synchrony. However, improvements in sample purification techniques are also required to reveal new phenomena. Here we assessed a cell sorter as a sample-pretreatment tool for a single-cell level assay. A cell sorter is routinely used for selecting one type of cells from a heterogeneous mixture of cells using specific fluorescence labels. In this case, we wanted to select cells of exactly the same size, shape, and cell-cycle stage from a population, without using a specific fluorescence label. Results We used four light scatter parameters: the peak height and area of the forward scatter (FSheight and FSarea and side scatter (SSheight and SSarea. The rat pheochromocytoma PC12 cell line, a neuronal cell line, was used for all experiments. The living cells concentrated in the high FSarea and middle SSheight/SSarea fractions. Single cells without cell clumps were concentrated in the low SS and middle FS fractions, and in the higher FSheight/FSarea and SSheight/SSarea fractions. The cell populations from these viable, single-cell-rich fractions were divided into twelve subfractions based on their FSarea-SSarea profiles, for more detailed analysis. We found that SSarea was proportional to the cell volume and the FSarea correlated with cell roundness and elongation, as well as with the level of DNA in the cell. To test the method and to characterize the basic properties of the isolated single cells, sorted cells were cultured in separate wells. The cells in all subfractions survived, proliferated and differentiated normally, suggesting that there was no serious damage. The smallest, roundest, and smoothest cells had the highest viability. There was no correlation between proliferation and differentiation. NGF increases

  20. Decreased serum cell-free DNA levels in rheumatoid arthritis

    OpenAIRE

    Dunaeva, Marina; Buddingh’, Bastiaan C.; René E M Toes; Luime, Jolanda J.; Lubberts, Erik; Pruijn, Ger J. M.

    2015-01-01

    Purpose Recent studies have demonstrated that serum/plasma DNA and RNA molecules in addition to proteins can serve as biomarkers. Elevated levels of these nucleic acids have been found not only in acute, but also in chronic conditions, including autoimmune diseases. The aim of this study was to assess cell-free DNA (cfDNA) levels in sera of rheumatoid arthritis (RA) patients compared to controls. Methods cfDNA was extracted from sera of patients with early and established RA, relapsing-remitt...

  1. Low Reactive Level Laser Therapy for Mesenchymal Stromal Cells Therapies

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    Toshihiro Kushibiki

    2015-01-01

    Full Text Available Low reactive level laser therapy (LLLT is mainly focused on the activation of intracellular or extracellular chromophore and the initiation of cellular signaling by using low power lasers. Over the past forty years, it was realized that the laser therapy had the potential to improve wound healing and reduce pain and inflammation. In recent years, the term LLLT has become widely recognized in the field of regenerative medicine. In this review, we will describe the mechanisms of action of LLLT at a cellular level and introduce the application to mesenchymal stem cells and mesenchymal stromal cells (MSCs therapies. Finally, our recent research results that LLLT enhanced the MSCs differentiation to osteoblast will also be described.

  2. Protein expression analyses at the single cell level.

    Science.gov (United States)

    Ohno, Masae; Karagiannis, Peter; Taniguchi, Yuichi

    2014-01-01

    The central dogma of molecular biology explains how genetic information is converted into its end product, proteins, which are responsible for the phenotypic state of the cell. Along with the protein type, the phenotypic state depends on the protein copy number. Therefore, quantification of the protein expression in a single cell is critical for quantitative characterization of the phenotypic states. Protein expression is typically a dynamic and stochastic phenomenon that cannot be well described by standard experimental methods. As an alternative, fluorescence imaging is being explored for the study of protein expression, because of its high sensitivity and high throughput. Here we review key recent progresses in fluorescence imaging-based methods and discuss their application to proteome analysis at the single cell level. PMID:25197931

  3. Protein Expression Analyses at the Single Cell Level

    Directory of Open Access Journals (Sweden)

    Masae Ohno

    2014-09-01

    Full Text Available The central dogma of molecular biology explains how genetic information is converted into its end product, proteins, which are responsible for the phenotypic state of the cell. Along with the protein type, the phenotypic state depends on the protein copy number. Therefore, quantification of the protein expression in a single cell is critical for quantitative characterization of the phenotypic states. Protein expression is typically a dynamic and stochastic phenomenon that cannot be well described by standard experimental methods. As an alternative, fluorescence imaging is being explored for the study of protein expression, because of its high sensitivity and high throughput. Here we review key recent progresses in fluorescence imaging-based methods and discuss their application to proteome analysis at the single cell level.

  4. System-level modeling and simulation of the cell culture microfluidic biochip ProCell

    OpenAIRE

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan; Hemmingsen, Mette; Dufva, Martin

    2010-01-01

    Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory. There are two technologies for the microfluidic biochips: droplet-based and flow-based. In this paper we are interested in flow-based microfluidic biochips, where the liquid flows continuously through pre-defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chi...

  5. Rising cyclin-CDK levels order cell cycle events.

    Directory of Open Access Journals (Sweden)

    Catherine Oikonomou

    Full Text Available BACKGROUND: Diverse mitotic events can be triggered in the correct order and time by a single cyclin-CDK. A single regulator could confer order and timing on multiple events if later events require higher cyclin-CDK than earlier events, so that gradually rising cyclin-CDK levels can sequentially trigger responsive events: the "quantitative model" of ordering. METHODOLOGY/PRINCIPAL FINDINGS: This 'quantitative model' makes predictions for the effect of locking cyclin at fixed levels for a protracted period: at low cyclin levels, early events should occur rapidly, while late events should be slow, defective, or highly variable (depending on threshold mechanism. We titrated the budding yeast mitotic cyclin Clb2 within its endogenous expression range to a stable, fixed level and measured time to occurrence of three mitotic events: growth depolarization, spindle formation, and spindle elongation, as a function of fixed Clb2 level. These events require increasingly more Clb2 according to their normal order of occurrence. Events occur efficiently and with low variability at fixed Clb2 levels similar to those observed when the events normally occur. A second prediction of the model is that increasing the rate of cyclin accumulation should globally advance timing of all events. Moderate (<2-fold overexpression of Clb2 accelerates all events of mitosis, resulting in consistently rapid sequential cell cycles. However, this moderate overexpression also causes a significant frequency of premature mitoses leading to inviability, suggesting that Clb2 expression level is optimized to balance the fitness costs of variability and catastrophe. CONCLUSIONS/SIGNIFICANCE: We conclude that mitotic events are regulated by discrete cyclin-CDK thresholds. These thresholds are sequentially triggered as cyclin increases, yielding reliable order and timing. In many biological processes a graded input must be translated into discrete outputs. In such systems, expression of

  6. Genetic variants regulating immune cell levels in health and disease.

    Science.gov (United States)

    Orrù, Valeria; Steri, Maristella; Sole, Gabriella; Sidore, Carlo; Virdis, Francesca; Dei, Mariano; Lai, Sandra; Zoledziewska, Magdalena; Busonero, Fabio; Mulas, Antonella; Floris, Matteo; Mentzen, Wieslawa I; Urru, Silvana A M; Olla, Stefania; Marongiu, Michele; Piras, Maria G; Lobina, Monia; Maschio, Andrea; Pitzalis, Maristella; Urru, Maria F; Marcelli, Marco; Cusano, Roberto; Deidda, Francesca; Serra, Valentina; Oppo, Manuela; Pilu, Rosella; Reinier, Frederic; Berutti, Riccardo; Pireddu, Luca; Zara, Ilenia; Porcu, Eleonora; Kwong, Alan; Brennan, Christine; Tarrier, Brendan; Lyons, Robert; Kang, Hyun M; Uzzau, Sergio; Atzeni, Rossano; Valentini, Maria; Firinu, Davide; Leoni, Lidia; Rotta, Gianluca; Naitza, Silvia; Angius, Andrea; Congia, Mauro; Whalen, Michael B; Jones, Chris M; Schlessinger, David; Abecasis, Gonçalo R; Fiorillo, Edoardo; Sanna, Serena; Cucca, Francesco

    2013-09-26

    The complex network of specialized cells and molecules in the immune system has evolved to defend against pathogens, but inadvertent immune system attacks on "self" result in autoimmune disease. Both genetic regulation of immune cell levels and their relationships with autoimmunity are largely undetermined. Here, we report genetic contributions to quantitative levels of 95 cell types encompassing 272 immune traits, in a cohort of 1,629 individuals from four clustered Sardinian villages. We first estimated trait heritability, showing that it can be substantial, accounting for up to 87% of the variance (mean 41%). Next, by assessing ∼8.2 million variants that we identified and confirmed in an extended set of 2,870 individuals, 23 independent variants at 13 loci associated with at least one trait. Notably, variants at three loci (HLA, IL2RA, and SH2B3/ATXN2) overlap with known autoimmune disease associations. These results connect specific cellular phenotypes to specific genetic variants, helping to explicate their involvement in disease. PMID:24074872

  7. Nutritional stress enhances cell viability of odontoblast-like cells subjected to low level laser irradiation

    International Nuclear Information System (INIS)

    In spite of knowing that cells under stress are biostimulated by low level laser (LLL) irradiation, the ideal condition of stress to different cell lines has not yet been established. Consequently, the aim of the present in vitro study was to evaluate the effects of a defined parameter of LLL irradiation applied on stressed odontoblast-like pulp cells (MDPC-23). The cells were seeded (12500 cells/cm2) in wells of 24-well plates using complete culture medium (DMEM) and incubated for 24 hours. Then, the DMEM was replaced by a new medium with low concentrations (nutritional stress condition) of fetal bovine serum (FBS) giving rise to the following experimental groups: G1: 2% FBS; G2: 5% FBS; and G3: 10% FBS. The cells were irradiated three times with LLL in specific parameters (808±3 nm, 100 mW, 1.5 J/cm2) every 24 hours. No irradiation was carried out in groups G4 (2% FBS-Control), G5 (5% FBS-Control), and G6 (10% FBS-Control). For all groups, the cell metabolism (MTT assay) and morphology (SEM) was evaluated. The experimental groups showed enhanced cell metabolism and normal cell morphology regardless of FBS concentration. A slight increase in the cell metabolism was observed only in group G2. It was concluded that cell nutritional stress caused by reducing the concentration of FBS to 5% is the most suitable method to assess the biostimulation of LLL irradiated MDPC-23 cells

  8. Effects of fluctuating glucose levels on neuronal cells in vitro.

    Science.gov (United States)

    Russo, Vincenzo C; Higgins, Sandra; Werther, George A; Cameron, Fergus J

    2012-08-01

    There is increasing evidence for glucose fluctuation playing a role in the damaging effects of diabetes on various organs, including the brain. We aimed to study the effects of glycaemic variation (GV) upon mitochondrial activity using an in vitro human neuronal model. The metabolic disturbance of GV in neuronal cells, was mimicked via exposure of neuroblastoma cells SH-SY5Y to constant glucose or fluctuating (i.e. 6 h cycles) for 24 and 48 h. Mitochondrial dehydrogenase activity was determined via MTT assay. Cell mitochondrial activity (MTT) was moderately decreased in constant high glucose, but markedly decreased following 24 and 48 h of cyclical glucose fluctuations. Glucose transport determined via 2-deoxy-D-[1-(14)C] glucose uptake was regulated in an exaggerated manner in response to glucose variance, accompanied by modest changes in GLUT 1 mRNA abundance. Osmotic components of these glucose effects were investigated in the presence of the osmotic-mimics mannitol and L: -glucose. Both treatments showed that fluctuating osmolality did not result in a significant change in mitochondrial activity and had no effects on (14)Cglucose uptake, suggesting that adverse effects on mitochondrial function were specifically related to metabolically active glucose fluctuations. Apoptosis gene expression showed that both intrinsic and extrinsic apoptotic pathways were modulated by glucose variance, with two major response clusters corresponding to (i) glucose stress-modulated genes, (ii) glucose mediated osmotic stress-modulated genes. Gene clustering analysis by STRING showed that most of the glucose stress-modulated genes were components of the intrinsic/mitochondrial apoptotic pathway including Bcl-2, Caspases and apoptosis executors. On the other hand the glucose mediated osmotic stress-modulated genes were mostly within the extrinsic apoptotic pathway, including TNF receptor and their ligands and adaptors/activators/initiators of apoptosis. Fluctuating glucose levels

  9. Serum leptin, neuron specific enolase and S-100B in relation to post-stroke depression in a prospective nested case-control study

    Directory of Open Access Journals (Sweden)

    Mei-ying ZHAO

    2015-04-01

    Full Text Available Objective To investigate the relationship between the serum levels of leptin, neuron-specific enolase (NSE and S-100B in patients of stroke and the incidence of post-stroke depression (PSD. Methods The clinical data of 121 cases of acute ischemic stroke, admitted to Zhengzhou Central Hospital affiliated to Zhengzhou University from Jun. 2010 to Dec. 2012, were retrospectively analyzed. After six months of follow-up 42 patients were diagnosed as suffering from PSD (Hamilton Depression Scale score ≥8. Another 42 participants with available matching data on onset time, age, gender and lesions of brain were selected. The serum samples were collected from all patients at time of discharge, and the concentrations of serum leptin, NSE and S-100B were analyzed by enzyme-linked immunosorbent assay (ELISA kit. Correlation and efficiency of diagnosing PSD among them was validated by receptor operator curve (ROC. Results The concentration of serum leptin, NSE, and S-100B in PSD group (25.84±13.80, 2.59±1.48 and 25.03±8.24μg/L, respectively was higher than that in the control group (8.67±6.17, 2.27±1.84 and 22.40±6.84μg/L, respectively. No obvious correlation was found between serum leptin and the NSE and S-100B in PSD patients. Based on the ROC curve, the area under the curve of serum leptin in PSD patients was 0.935 (95%CI 0.885-0.984, and the optimal cutoff value of serum leptin level was 16.17μg/L, which was an indicator for predicting of PSD with 81.0% sensitivity and 90.1% specificity. Conclusion Elevation of serum leptin level at admission was found to be associated with PSD, and it may act as a new marker for predicting the occurrence of PSD. DOI: 10.11855/j.issn.0577-7402.2015.03.11

  10. Valor da enolase específica do neurônio como indicador de prognóstico pós-parada cardiorrespiratória Serum neuron-specific enolase as a prognostic marker after a cardiac arrest

    Directory of Open Access Journals (Sweden)

    Tatiana H. Rech

    2006-12-01

    Full Text Available JUSTIFICATIVA E OBJETIVOS: A parada cardíaca é um estado de grave hipoperfusão cerebral. Os pacientes que sobrevivem a uma reanimação cardiorrespiratória estão sob grande risco de vir a morrer ou desenvolver lesão cerebral incapacitante, inclusive estado vegetativo persistente. Uma definição precoce do prognóstico desses pacientes tem implicações éticas e econômicas. O objetivo desse estudo foi revisar o valor prognóstico da Enolase Específica do Neurônio (NSE em predizer precocemente os desfechos de pacientes após uma parada cardíaca. CONTEÚDO: A lesão cerebral permanente é a complicação mais temida de uma reanimação cardíaca prolongada. Muitos estudos têm tentado isolar fatores prognósticos que possam estar associados com desfechos clínicos em pacientes sobreviventes de parada cardíaca. Indicadores bioquímicos de morte neuronal parecem promissores nesse cenário. Nesse contexto, a NSE vem sendo estudada em pacientes reanimados de paradas cardíacas e níveis elevados dessa enzima sugerem lesão encefálica mais extensa e estão associados a desfechos clínicos desfavoráveis. CONCLUSÕES: Os desfechos depois de uma parada cardíaca são determinados principalmente pelo grau de lesão cerebral isquêmica e medidas precoces de NSE sérica podem ser um método adjunto de grande valor na avaliação prognóstica desses pacientes.BACKGROUND AND OBJECTIVES: Cardiac arrest is a state of severe cerebral perfusion deficit. Patients recovering from a cardiopulmonary resuscitation are at great risk of subsequent death or incapacitating neurologic injury, including persistent vegetative state. The early definition of prognosis for these patients has ethical and economic implications. The main purpose of this manuscript was to review the prognostic value of serum Neuron-Specific Enolase (NSE in predicting outcomes in patients early after a cardiac arrest. CONTENTS: Severe neurologic disability is the most feared

  11. Dose levels in the hot cells area ININ

    International Nuclear Information System (INIS)

    The Laboratory of Hot Cells (LCC) located in the National Institute of Nuclear Research (ININ) is an institution, it is an area where radioactive material is managed with different activity values, in function of its original design for 10,000 curies of Co-60. Managing this materials in the installation, it implies to measure and to analyze the dose levels that the POE will receive as well as the implementation of appropriate measures of radiological protection and radiological safety, so that that is completed settled down by the concept ALARA. In this work they are carried out mensurations of the levels of the dose to receive for the POE when managing radionuclides with maximum activities that can be allowed in function of the current conditions of the cells and an evaluation of the obtained results is made comparing them with the effective international norms as well as the application of the program of surveillance and radiological protection implemented for the development of the works that are carry out in the installation. (Author)

  12. Amyloid β levels in human red blood cells.

    Directory of Open Access Journals (Sweden)

    Takehiro Kiko

    Full Text Available UNLABELLED: Amyloid β-peptide (Aβ is hypothesized to play a key role by oxidatively impairing the capacity of red blood cells (RBCs to deliver oxygen to the brain. These processes are implicated in the pathogenesis of Alzheimer's disease (AD. Although plasma Aβ has been investigated thoroughly, the presence and distribution of Aβ in human RBCs are still unclear. In this study, we quantitated Aβ40 and Aβ42 in human RBCs with ELISA assays, and provided evidence that significant amounts of Aβ could be detected in RBCs and that the RBC Aβ levels increased with aging. The RBC Aβ levels increased with aging. On the other hand, providing an antioxidant supplement (astaxanthin, a polar carotenoid to humans was found to decrease RBC Aβ as well as oxidative stress marker levels. These results suggest that plasma Aβ40 and Aβ42 bind to RBCs (possibly with aging, implying a pathogenic role of RBC Aβ. Moreover, the data indicate that RBC Aβ40 and Aβ42 may constitute biomarkers of AD. As a preventive strategy, therapeutic application of astaxanthin as an Aβ-lowering agent in RBCs could be considered as a possible anti-dementia agent. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN42483402.

  13. Autophagy Alleviates Melamine-Induced Cell Death in PC12 Cells Via Decreasing ROS Level.

    Science.gov (United States)

    Wang, Hui; Gao, Na; Li, Zhigui; Yang, Zhuo; Zhang, Tao

    2016-04-01

    Since melamine was illegally added to raw milk for increasing the apparent protein content, such a scandal has not been quite blown out. Previous studies showed that melamine induced apoptosis and oxidative damage in both in vivo and in vitro experiments. It is well known that autophagy is closely related to oxidative stress. In the present study, we examined whether autophagy played an important role in protecting PC12 cells, which were damaged by melamine. Immunofluorescence assay showed that melamine enhanced the number of punctuate dot, indicating the increase of autophagosomes. Western blot assay presented that melamine significantly elevated the expression level of autophagy markers including LC3-II/LC3-I ratio, beclin-1, and Atg 7. Rapamycin further enhanced the effect, whereas 3-methyadenine (3-MA) inhibited it. MTT assay exhibited that rapamycin significantly enhanced the cell viability (P < 0.01), while 3-MA considerably reduced it in melamine-treated PC12 cells (P < 0.01). Furthermore, flow cytometry assay showed that rapamycin considerably reduced the reactive oxygen species (ROS) level of the cells (P < 0.01), but 3-MA increased the generation of ROS (P < 0.01). Additionally, the superoxide dismutase (SOD) activity was notably increased by rapamycin in melamine-treated PC12 cells (P < 0.01), while the activity of which was prominently decreased by 3-MA (P < 0.01). Malondialdehyde (MDA) assay showed that rapamycin remarkably decreased the MDA level of the cells (P < 0.05), while 3-MA increased it (P < 0.01). Consequently, this study demonstrated that autophagy protected PC12 cells from melamine-induced cell death via inhibiting the excessive generation of ROS. Regulating autophagy may become a new targeted therapy to relieve the damage induced by melamine. PMID:25724280

  14. Insect cell transformation vectors that support high level expression and promoter assessment in insect cell culture

    Science.gov (United States)

    A somatic transformation vector, pDP9, was constructed that provides a simplified means of producing permanently transformed cultured insect cells that support high levels of protein expression of foreign genes. The pDP9 plasmid vector incorporates DNA sequences from the Junonia coenia densovirus th...

  15. Macro Level Modeling of a Tubular Solid Oxide Fuel Cell

    Directory of Open Access Journals (Sweden)

    Farshid Zabihian

    2010-11-01

    Full Text Available This paper presents a macro-level model of a solid oxide fuel cell (SOFC stack implemented in Aspen Plus® for the simulation of SOFC system. The model is 0-dimensional and accepts hydrocarbon fuels such as reformed natural gas, with user inputs of current density, fuel and air composition, flow rates, temperature, pressure, and fuel utilization factor. The model outputs the composition of the exhaust, work produced, heat available for the fuel reformer, and electrochemical properties of SOFC for model validation. It was developed considering the activation, concentration, and ohmic losses to be the main over-potentials within the SOFC, and mathematical expressions for these were chosen based on available studies in the literature. The model also considered the water shift reaction of CO and the methane reforming reaction. The model results were validated using experimental data from Siemens Westinghouse. The results showed that the model could capture the operating pressure and temperature dependency of the SOFC performance successfully in an operating range of 1–15 atm for pressure and 900 °C–1,000 °C for temperature. Furthermore, a sensitivity analysis was performed to identify the model constants and input parameters that impacted the over-potentials.

  16. Intracellular levels of calmodulin are increased in transformed cells

    Institute of Scientific and Technical Information of China (English)

    WANG; HONGQINGZHANG; 等

    1992-01-01

    By using Hoechst 33342,rabbit anti calmodulin antibody,FITC-labeled goat anti rabbit IgG and SR101(sulfo rhodamine 101)simultaneously to stain individual normal and transformed cells,the microspectrophotometric analysis demonstrated that 3 markers which represented the nucleus,calmodulin and total protein respectively,could be recognized in individualj cells without interference,The phase of the cell cycle was determined by DNA content(Hoechst 33342),We found that in transformed cells(NIH3T3) tsRSV-LA90,cultured at 33℃ and transformed C3H10T1/2 Cells),the ration of calmodulin to total protein (based on the phases of cell cycle)was higher than that in normal cells (NIH3T3 tsRSV-LA90 cells,cultured at 39℃ and C3H10T1/2 cells)in every cell cycle phase,This ration increased obviously only from G1 to S phase in either normal or transformed cells.The results showed that calmodulinreally increased during the transformation,and its increase was specific.In the meantime when cells proceeded from G1 to S.the intraceollular calmodulin content also increased specifically.

  17. Mesenchymal stem cells-derived vascular smooth muscle cells release abundant levels of osteoprotegerin

    Directory of Open Access Journals (Sweden)

    M Vaccarezza

    2009-03-01

    Full Text Available Although several studies have shown that the serum levels of osteoprotegerin (OPG are significantly elevated in patients affected with atherosclerotic lesions in coronary and peripheral arteries, the cellular source and the role of OPG in the physiopathology of atherosclerosis are not completely defined. Therefore, we aimed to investigate the potential contribution of mesenchymal stem cells in the production/release of OPG. OPG was detectable by immunohistochemistry in aortic and coronary atherosclerotic plaques, within or in proximity of intimal vascular smooth muscle cells (SMC. In addition, bone marrow mesenchymal stem cell (MSC-derived vascular SMC as well as primary aortic SMC released in the culture supernatant significantly higher levels of OPG with respect to MSCderived endothelial cells (EC or primary aortic EC. On the other hand, in vitro exposure to full-length human recombinant OPG significantly increased the proliferation rate of aortic SMC cultures, as monitored by bromodeoxyuridine incorporation. Taken together, these data suggest that OPG acts as an autocrine/paracrine growth factor for vascular SMC, which might contribute to the progression of atherosclerotic lesions.

  18. Influences of lamin A levels on induction of pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Bingfeng Zuo

    2012-09-01

    Lamin A is an inner nuclear membrane protein that maintains nuclear structure integrity, is involved in transcription, DNA damage response and genomic stability, and also links to cell differentiation, senescence, premature aging and associated diseases. Induced pluripotent stem (iPS cells have been successfully generated from various types of cells and used to model human diseases. It remains unclear whether levels of lamin A influence reprogramming of somatic cells to pluripotent states during iPS induction. Consistently, lamin A is expressed more in differentiated than in relatively undifferentiated somatic cells, and increases in expression levels with age. Somatic cells with various expression levels of lamin A differ in their dynamics and efficiency during iPS cell induction. Cells with higher levels of lamin A show slower reprogramming and decreased efficiency to iPS cells. Furthermore, depletion of lamin A by transient shRNA accelerates iPS cell induction from fibroblasts. Reduced levels of lamin A are associated with increased expression of pluripotent genes Oct4 and Nanog, and telomerase genes Tert and Terc. On the contrary, overexpression of lamin A retards somatic cell reprogramming to iPS-like colony formation. Our data suggest that levels of lamin A influence reprogramming of somatic cells to pluripotent stem cells and that artificial silencing of lamin A facilitates iPS cell induction. These findings may have implications in enhancing rejuvenation of senescent or older cells by iPS technology and manipulating lamin A levels.

  19. Cost and Performance Tradeoff Analysis of Cell Planning Levels

    OpenAIRE

    Gao, Jun

    2013-01-01

    In wireless communication systems, optimal placement of base station locations, i.e.,cell planning, has been considered one of major tools for performance improvement.However, the cell planning requires signicant eort for acquiring suitable sites aswell as expensive computer software for nding out optimal locations. While theprice of equipment has dropped rapidly, the cost of cell planing remains similar orbecomes even more costly with increasing complexity of wireless systems.Therefore,the c...

  20. Cell surface N-glycans influence the level of functional E-cadherin at the cell–cell border

    OpenAIRE

    M Kristen Hall; Douglas A Weidner; Sahil Dayal; Ruth A. Schwalbe

    2014-01-01

    E-cadherin is crucial for adhesion of cells to each other and thereby development and maintenance of tissue. While it is has been established that N-glycans inside the cell impact the level of E-cadherin at the cell surface of epithelial-derived cells, it is unclear whether N-glycans outside the cell control the clustering of E-cadherin at the cell–cell border. Here, we demonstrate reduction of N-glycans at the cell surface weakened the recruitment and retention of E-cadherin at the cell–cell...

  1. Effects of high doping levels silicon solar cell performance

    Science.gov (United States)

    Godlewski, M. P.; Brandhorst, H. W., Jr.; Baraona, C. R.

    1975-01-01

    The significance of the heavy doping effects (HDE) on the open-circuit voltage of silicon solar cells is assessed. Voltage calculations based on diffusion theory are modified to include the first order features of the HDE. Comparisions of the open-circuit voltage measured for cells of various base resistivities are made with those calculated using the diffusion model with and without the HDE. Results indicate that the observed variation of voltage with base resistivity is predicted by these effects. A maximum efficiency of 19% (AM0) and a voltage of 0.7 volts are calculated for 0.1 omega-cm cells assuming an optimum diffused layer impurity profile.

  2. Relationship of circulating cell-free DNA levels to cell-free fetal DNA levels, clinical characteristics and laboratory parameters in preeclampsia

    Directory of Open Access Journals (Sweden)

    Mézes Miklós

    2009-01-01

    Full Text Available Abstract Background The aim of our study was to examine whether increased circulating total cell-free DNA levels are related to the clinical characteristics and standard laboratory parameters of preeclamptic patients, to markers of inflammation, endothelial activation or injury, oxidative stress and to cell-free fetal DNA levels. Methods Circulating total cell-free DNA was measured by real-time quantitative PCR in plasma samples obtained from 67 preeclamptic and 70 normotensive pregnant women. Standard laboratory parameters, C-reactive protein, plasma von Willebrand factor antigen, plasma fibronectin, plasma malondialdehyde and cell-free fetal DNA levels were also determined. Results and Conclusion Circulating total cell-free and fetal deoxyribonucleic acid levels were significantly elevated in pregnancies complicated by preeclampsia (median: 11.395 vs. 32.460 and 0.001 vs. 0.086 pg/μl; P < .001. The quantity of plasma total cell-free DNA did not correlate with most of the laboratory parameters, except for serum aspartate aminotransferase and alanine aminotransferase activities (correlation coefficient: 0.31; P = 0.012 and 0.46; P < .001. There was no correlation with clinical characteristics, including body mass index. The releases of both free fetal and total cell-free deoxyribonucleic acid were found to be affected in preeclampsia. Hepatocellular necrosis seems to be responsible - at least partly - for increased circulating total DNA levels in preeclampsia, as suggested by the significant correlation with liver enzyme activities.

  3. Differing levels of excision repair in human fetal dermis and brain cells

    Energy Technology Data Exchange (ETDEWEB)

    Gibson, R.E. (Ohio State Univ., Columbus (USA). Dept. of Radiology); D' Ambrosio, S.M. (Ohio State Univ., Columbus (USA). Dept. of Radiology; Ohio State Univ., Columbus (USA). Dept. of Pharmacology)

    1982-01-01

    The levels of DNA excision repair, as measured by unscheduled DNA synthesis (UDS) and the UV-endonuclease sensitive site assay, were compared in cells derived from human fetal brain and dermal tissues. The level of UDS induced following ultraviolet (UV) irradiation was found to be lower (approx. 60%) in the fetal brain cells than in fetal dermal cells. It was determined, using the UV-endonuclease sensitive site assay to confirm the UDS observation, that 50% of the dimers induced by UV in fetal dermal cells were repaired in 8 h. while only 15% were removed in the fetal brain cells during the same period of time. Even after 24 h. only 44% of the dimers induced by UV in the fetal brain cells were repaired, while 65% were removed in the dermal cells. These data suggest that cultured human fetal brain cells exhibit lower levels of excision repair compared to cultured human fetal dermal cells.

  4. Production in a factory (the cell) requires high level of organisation : the cell: The plant’s smallest building block

    NARCIS (Netherlands)

    Heuvelink, E.

    2015-01-01

    The cell is the plant’s smallest building block. Many cultivation techniques and climate control measures have an effect at this level. Some knowledge about the functioning of the cell is therefore very useful. Many components of the cell have bizarre names so to understand it all better, for the pu

  5. Low p21(Waf1/Cip1) protein level sensitizes testicular germ cell tumor cells to Fas-mediated apoptosis

    NARCIS (Netherlands)

    Spierings, DCJ; de Vries, EGE; Stel, AJ; Rietstap, NT; Vellenga, E; de Jong, S

    2004-01-01

    In the present study, we investigated the relation between p21 expression and the sensitivity of testicular germ cell tumor (TGCT) cells to apoptotic stimuli. Despite similar cisplatin-induced wild-type p53 accumulation, the TGCT cell lines Tera and Scha expressed low p21 protein and mRNA levels in

  6. Modelling Cell Cycle using Different Levels of Representation

    CERN Document Server

    Basuki, Thomas Anung; Carvalho, Rafael V; 10.4204/EPTCS.11.4

    2009-01-01

    Understanding the behaviour of biological systems requires a complex setting of in vitro and in vivo experiments, which attracts high costs in terms of time and resources. The use of mathematical models allows researchers to perform computerised simulations of biological systems, which are called in silico experiments, to attain important insights and predictions about the system behaviour with a considerably lower cost. Computer visualisation is an important part of this approach, since it provides a realistic representation of the system behaviour. We define a formal methodology to model biological systems using different levels of representation: a purely formal representation, which we call molecular level, models the biochemical dynamics of the system; visualisation-oriented representations, which we call visual levels, provide views of the biological system at a higher level of organisation and are equipped with the necessary spatial information to generate the appropriate visualisation. We choose Spati...

  7. Thermal conductivity of biological cells at cellular level and correlation with disease state

    Science.gov (United States)

    Park, Byoung Kyoo; Woo, Yunho; Jeong, Dayeong; Park, Jaesung; Choi, Tae-Youl; Simmons, Denise Perry; Ha, Jeonghong; Kim, Dongsik

    2016-06-01

    This paper reports the thermal conductivity k of matched pair cell lines: two pairs of a normal and a cancer cell, one pair of a primary and metastatic cell. The 3ω method with a nanoscale thermal sensor was used to measure k at the single-cell level. To observe the difference in k between normal and cancer cells, the measurements were conducted for Hs 578Bst/Hs 578 T (human breast cells) and TE 353.Sk/TE 354.T (human skin cells). Then k of WM-115/WM-266-4, a primary and metastatic pair of human skin cell, was measured to find the effect of disease progression on k. The measured k data for normal and disease cell samples show statistically meaningful differences. In all cases, k decreased as the disease progressed. This work shows that thermal-analysis schemes, such as the 3ω method, have a potential to detect diseases at the cell level.

  8. Does Ploidy Level Directly Control Cell Size? Counterevidence from Arabidopsis Genetics

    OpenAIRE

    Tsukaya, Hirokazu

    2013-01-01

    Ploidy level affects cell size in many organisms, and ploidy-dependent cell enlargement has been used to breed many useful organisms. However, how polyploidy affects cell size remains unknown. Previous studies have explored changes in transcriptome data caused by polyploidy, but have not been successful. The most naïve theory explaining ploidy-dependent cell enlargement is that increases in gene copy number increase the amount of protein, which in turn increases the cell volume. This hypothes...

  9. Cell-Free Fetal DNA and Cell-Free Total DNA Levels in Spontaneous Abortion with Fetal Chromosomal Aneuploidy

    OpenAIRE

    Ji Hyae Lim; Min Hyoung Kim; You Jung Han; Da Eun Lee; So Yeon Park; Jung Yeol Han; Moon Young Kim; Hyun Mee Ryu

    2013-01-01

    BACKGROUND: Cell-free fetal DNA and cell-free total DNA in maternal circulation have been proposed as potential markers for noninvasive monitoring of the placental condition during the pregnancy. However, the correlation of and change in cell-free fetal DNA and cell-free total DNA in spontaneous abortion (SA) with fetal chromosomal aneuploidy have not yet been reported. Therefore, we investigated cell-free fetal DNA and cell-free total DNA levels in SA women with fetal chromosomal aneuploidy....

  10. Distinct gene expression signatures in human embryonic stem cells differentiated towards definitive endoderm at single-cell level

    DEFF Research Database (Denmark)

    Norrman, Karin; Strömbeck, Anna; Semb, Henrik; Ståhlberg, Anders

    2013-01-01

    of anterior definitive endoderm (DE). Here, we differentiated human embryonic stem cells towards DE using three different activin A based treatments. Differentiation efficiencies were evaluated by gene expression profiling over time at cell population level. A panel of key markers was used to study...... were useful to monitor the temporal expression of genes involved in primitive streak formation and endoderm formation, while single-cell analysis allowed us to study cell culture heterogeneity and fingerprint individual cells. In addition, single-cell analysis revealed distinct gene expression patterns...

  11. Effects of high doping levels on silicon solar cell performance

    Science.gov (United States)

    Godlewski, M. P.; Brandhorst, H. W., Jr.; Baraona, C. R.

    1975-01-01

    Open-circuit voltages measured in silicon solar cells made from 0.01 ohm-cm material are 150 mV lower than voltages calculated from simple diffusion theory and cannot be explained by poor diffusion lengths or surface leakage currents. An analytical study was made to determine whether high doping effects, which increase the intrinsic carrier concentration, could account for the low observed voltages and to determine the limits on voltage and efficiency imposed by high doping effects. The results indicate that the observed variation of voltage with base resistivity is predicted by these effects. A maximum efficiency of 19% (AMO) and a voltage of 0.7 volts were calculated for 0.1 ohm-cm cells assuming an optimum diffused layer impurity profile.

  12. Host Cell Sumoylation Level Influences Papillomavirus E2 Protein Stability

    OpenAIRE

    Wu, Yu-Chieh; Bian, Xue-Lin; Heaton, Phillip R.; G. Wilson

    2009-01-01

    The stability of papillomavirus E2 proteins is regulated by proteasomal degradation, and regulation of degradation could contribute to the higher expression levels E2 proteins observed in suprabasal layers of differentiated skin. We have recently shown that the E2 proteins are modified by sumoylation [Wu Y-C, Roark AA, Bian X-L, Wilson, VG (2008) Virol 378:329–338], and that sumoylation levels are up-regulated during keratinocyte differentiation [Deyrieux AF, Rosas-Acosta G, Ozbun MA, Wilson ...

  13. Neuron-Specific Enolase as a Predictor of Death or Poor Neurological Outcome After Out-of-Hospital Cardiac Arrest and Targeted Temperature Management at 33°C and 36°C

    DEFF Research Database (Denmark)

    Stammet, Pascal; Collignon, Olivier; Hassager, Christian; Wise, Matthew P; Hovdenes, Jan; Åneman, Anders; Horn, Janneke; Devaux, Yvan; Erlinge, David; Kjaergaard, Jesper; Gasche, Yvan; Wanscher, Michael; Cronberg, Tobias; Friberg, Hans; Wetterslev, Jørn; Pellis, Tommaso; Kuiper, Michael; Gilson, Georges; Nielsen, Niklas

    2015-01-01

    BACKGROUND: Neuron-specific enolase (NSE) is a widely-used biomarker for prognostication of neurological outcome after cardiac arrest, but the relevance of recommended cutoff values has been questioned due to the lack of a standardized methodology and uncertainties over the influence of temperatu...

  14. Effect of coffee extracts on intracellular calcium level in levels in glial cells

    OpenAIRE

    Akın, Demet; Görmüş, Uzay; Yapışlar, Hande; Farah, Adriana

    2012-01-01

    Widely used antidepressant drugs such as fluoxetine exert additional blocking effects on voltage gated Ca⁺² channels. Differences in intracellular calcium levels may be involved in the release of monoamines, which play important role in the pathogenesis of depression.

  15. Single-cell level based approach to investigate bacterial metabolism during batch industrial fermentation

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Larsen, Poul; Eriksen, Niels T.;

    Escherichia coli fermentations have been studied for decades, but most results are based on average measurements of the whole populations of cells, whilst averaged data can mask the distribution of activities at the sub-population or single-cell level. A population of genetically identical cells ...

  16. Low Reactive Level Laser Therapy for Mesenchymal Stromal Cells Therapies

    OpenAIRE

    Toshihiro Kushibiki; Takeshi Hirasawa; Shinpei Okawa; Miya Ishihara

    2015-01-01

    Low reactive level laser therapy (LLLT) is mainly focused on the activation of intracellular or extracellular chromophore and the initiation of cellular signaling by using low power lasers. Over the past forty years, it was realized that the laser therapy had the potential to improve wound healing and reduce pain and inflammation. In recent years, the term LLLT has become widely recognized in the field of regenerative medicine. In this review, we will describe the mechanisms of action of LLLT...

  17. Intracellular ATP Levels are a Pivotal Determinant of Chemoresistance in Colon Cancer Cells

    OpenAIRE

    Zhou, Yunfei; Tozzi, Federico; Chen, Jinyu; Fan, Fan; Xia, Ling; Wang, JinRong; Gao, Guang; Zhang, Aijun; Xia, Xuefeng; Brasher, Heather; Widger, William; Ellis, Lee M.; Weihua, Zhang

    2011-01-01

    Altered metabolism in cancer cells is suspected to contribute to chemoresistance but the precise mechanisms are unclear. Here we show that intracellular ATP levels are a core determinant in the development of acquired cross-drug resistance of human colon cancer cells that harbor different genetic backgrounds. Drug-resistant cells were characterized by defective mitochondrial ATP production, elevated aerobic glycolysis, higher absolute levels of intracellular ATP and enhanced HIF-1α-mediated s...

  18. Cell-free fetal DNA and cell-free total DNA levels in spontaneous abortion with fetal chromosomal aneuploidy.

    Directory of Open Access Journals (Sweden)

    Ji Hyae Lim

    Full Text Available BACKGROUND: Cell-free fetal DNA and cell-free total DNA in maternal circulation have been proposed as potential markers for noninvasive monitoring of the placental condition during the pregnancy. However, the correlation of and change in cell-free fetal DNA and cell-free total DNA in spontaneous abortion (SA with fetal chromosomal aneuploidy have not yet been reported. Therefore, we investigated cell-free fetal DNA and cell-free total DNA levels in SA women with fetal chromosomal aneuploidy. METHODOLOGY/PRINCIPAL FINDINGS: A nested case-control study was conducted with maternal plasma collected from 268 women in their first trimester of pregnancy. Subjects included 41 SA with normal fetal karyotype, 26 SA with fetal chromosomal aneuploidy, and 201 normal controls. The unmethylated PDE9A gene was used to measure the maternal plasma levels of cell-free fetal DNA. The GAPDH gene was used to measure the maternal plasma levels of cell-free total DNA. The diagnostic accuracy was measured using receiver-operating characteristic (ROC curves. Levels of cell-free fetal DNA and cell-free total DNA were significantly higher in both SA women with normal fetal karyotype and SA women with fetal chromosomal aneuploidy in comparison with the normal controls (P<0.001 in both. The correlation between cell-free fetal DNA and cell-free total DNA levels was stronger in the normal controls (r = 0.843, P<0.001 than in SA women with normal karyotype (r = 0.465, P = 0.002 and SA women with fetal chromosomal aneuploidy (r = 0.412, P = 0.037. The area under the ROC curve for cell-free fetal DNA and cell-free total DNA was 0.898 (95% CI, 0.852-0.945 and 0.939 (95% CI, 0.903-0.975, respectively. CONCLUSIONS: Significantly high levels of cell-free fetal DNA and cell-free total DNA were found in SA women with fetal chromosomal aneuploidy. Our findings suggest that cell-free fetal DNA and cell-free total DNA may be useful biomarkers for the prediction of SA

  19. Modular Approach for Continuous Cell-Level Balancing to Improve Performance of Large Battery Packs: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Muneed ur Rehman, M.; Evzelman, M.; Hathaway, K.; Zane, R.; Plett, G. L.; Smith, K.; Wood, E.; Maksimovic, D.

    2014-10-01

    Energy storage systems require battery cell balancing circuits to avoid divergence of cell state of charge (SOC). A modular approach based on distributed continuous cell-level control is presented that extends the balancing function to higher level pack performance objectives such as improving power capability and increasing pack lifetime. This is achieved by adding DC-DC converters in parallel with cells and using state estimation and control to autonomously bias individual cell SOC and SOC range, forcing healthier cells to be cycled deeper than weaker cells. The result is a pack with improved degradation characteristics and extended lifetime. The modular architecture and control concepts are developed and hardware results are demonstrated for a 91.2-Wh battery pack consisting of four series Li-ion battery cells and four dual active bridge (DAB) bypass DC-DC converters.

  20. Effects of physiological levels of the green tea extract Epigallocatechin gallate (EGCG on breast cancer cells

    Directory of Open Access Journals (Sweden)

    Li eZeng

    2014-05-01

    Full Text Available Physiological concentrations of the green tea extract EGCG caused growth inhibition in oestrogen receptor α (ERα-positive MCF7 cells, that was associated with down-regulation of the ERα and reduced insulin-like growth factor (IGF binding protein-2 (IGFBP-2 abundance and increased protein abundance of the tumour suppressor genes p53/p21. In contrast to MCF7 cells that have wt p53, EGCG alone did not change cell proliferation or death significantly in another ERα-positive cell line T47D that possesses mutant p53. EGCG increased ERα protein levels and as a consequence, the cells responded significantly better to an ERα antagonist Tamoxifen (TAM in the presence of EGCG. EGCG significantly increased cell death in an ERα-negative cell line, MDA-MB-231 that also possesses mutant p53. EGCG significantly increased the ERα and IGF-I receptor (IGF-IR levels and thereby enhanced the sensitivities of the cells to Tamoxifen and a blocking antibody targeting the IGF-1R (αIR3. In contrast to MCF7, T47D and MDA-MB-231 breast cancer cells that exhibited significant changes in key molecules involved in breast growth and survival upon treatment with physiological levels of EGCG, the growth, survival and levels of these proteins in non-malignant breast epithelial cells, MCF10A cells, were not affected.

  1. Stem cells modified by brain-derived neurotrophic fac-tor to promote stem cells differentiation into neurons and enhance neuromotor function after brain injury

    Institute of Scientific and Technical Information of China (English)

    ZHANG Sai; LIU Xiao-zhi; LIU Zhen-lin; WANG Yan-min; HU Qun-liang; MA Tie-zhu; SUN Shi-zhong

    2009-01-01

    Objective: To promote stem cells differentiation into neurons and enhance neuromotor function after brain in-jury through brain-derived neurotrophic factor (BDNF) induction.Methods: Recombinant adenovirus vector was ap-plied to the transfection of BDNF into human-derived um-bilical cord mesenchymal stem cells (UCMSCs). Enzyme linked immunosorbent assay (ELISA) was used to deter-mine the secretion phase of BDNF. The brain injury model of athymic mice induced by hydraulic pressure percussion was established for transplantation of stem cells into the edge of injury site. Nerve function scores were obtained, and the expression level of transfected and non-transfected BDNF, proportion of neuron specific enolase (NSE) andglial fibrillary acidic protein (GFAP), and the number of apoptosis cells were compared respectively. Results: The BDNF expression achieved its stabiliza-tion at a high level 72 hours after gene transfection. The mouse obtained a better score of nerve function, and the proportion of the NSE-positive cells increased significantly (P<0.05), but GFAP-positive cells decreased in BDNF-UCMSCs group compared with the other two groups (P<0.05). At the site of high expression of BDNF, the number of apoptosis cells decreased markedly.Conclusion: BDNF gene can promote the differentia-tion of the stem cells into neurons rather than gliai cells, and enhance neuromotor function after brain injury.

  2. Cell density related gene expression: SV40 large T antigen levels in immortalized astrocyte lines

    Directory of Open Access Journals (Sweden)

    Jacobberger James W

    2002-04-01

    Full Text Available Abstract Background Gene expression is affected by population density. Cell density is a potent negative regulator of cell cycle time during exponential growth. Here, we asked whether SV40 large T antigen (Tag levels, driven by two different promoters, changed in a predictable and regular manner during exponential growth in clonal astrocyte cell lines, immortalized and dependent on Tag. Results Expression and cell cycle phase fractions were measured and correlated using flow cytometry. T antigen levels did not change or increased during exponential growth as a function of the G1 fraction and increasing cell density when Tag was transcribed from the Moloney Murine Leukemia virus (MoMuLV long terminal repeat (LTR. When an Rb-binding mutant T antigen transcribed from the LTR was tested, levels decreased. When transcribed from the herpes thymidine kinase promoter, Tag levels decreased. The directions of change and the rates of change in Tag expression were unrelated to the average T antigen levels (i.e., the expression potential. Conclusions These data show that Tag expression potential in these lines varies depending on the vector and clonal variation, but that the observed level depends on cell density and cell cycle transit time. The hypothetical terms, expression at zero cell density and expression at minimum G1 phase fraction, were introduced to simplify measures of expression potential.

  3. High content analysis at single cell level identifies different cellular responses dependent on nanomaterial concentrations

    Science.gov (United States)

    Manshian, Bella B.; Munck, Sebastian; Agostinis, Patrizia; Himmelreich, Uwe; Soenen, Stefaan J.

    2015-09-01

    A mechanistic understanding of nanomaterial (NM) interaction with biological environments is pivotal for the safe transition from basic science to applied nanomedicine. NM exposure results in varying levels of internalized NM in different neighboring cells, due to variances in cell size, cell cycle phase and NM agglomeration. Using high-content analysis, we investigated the cytotoxic effects of fluorescent quantum dots on cultured cells, where all effects were correlated with the concentration of NMs at the single cell level. Upon binning the single cell data into different categories related to NM concentration, this study demonstrates, for the first time, that quantum dots activate both cytoprotective and cytotoxic mechanisms, resulting in a zero net result on the overall cell population, yet with significant effects in cells with higher cellular NM levels. Our results suggest that future NM cytotoxicity studies should correlate NM toxicity with cellular NM numbers on the single cell level, as conflicting mechanisms in particular cell subpopulations are commonly overlooked using classical toxicological methods.

  4. Intercellular Variability in Protein Levels from Stochastic Expression and Noisy Cell Cycle Processes

    Science.gov (United States)

    Soltani, Mohammad; Vargas-Garcia, Cesar A.; Antunes, Duarte; Singh, Abhyudai

    2016-01-01

    Inside individual cells, expression of genes is inherently stochastic and manifests as cell-to-cell variability or noise in protein copy numbers. Since proteins half-lives can be comparable to the cell-cycle length, randomness in cell-division times generates additional intercellular variability in protein levels. Moreover, as many mRNA/protein species are expressed at low-copy numbers, errors incurred in partitioning of molecules between two daughter cells are significant. We derive analytical formulas for the total noise in protein levels when the cell-cycle duration follows a general class of probability distributions. Using a novel hybrid approach the total noise is decomposed into components arising from i) stochastic expression; ii) partitioning errors at the time of cell division and iii) random cell-division events. These formulas reveal that random cell-division times not only generate additional extrinsic noise, but also critically affect the mean protein copy numbers and intrinsic noise components. Counter intuitively, in some parameter regimes, noise in protein levels can decrease as cell-division times become more stochastic. Computations are extended to consider genome duplication, where transcription rate is increased at a random point in the cell cycle. We systematically investigate how the timing of genome duplication influences different protein noise components. Intriguingly, results show that noise contribution from stochastic expression is minimized at an optimal genome-duplication time. Our theoretical results motivate new experimental methods for decomposing protein noise levels from synchronized and asynchronized single-cell expression data. Characterizing the contributions of individual noise mechanisms will lead to precise estimates of gene expression parameters and techniques for altering stochasticity to change phenotype of individual cells. PMID:27536771

  5. Liraglutide prevents high glucose level induced insulinoma cells apoptosis by targeting autophagy

    Institute of Scientific and Technical Information of China (English)

    CHEN Ze-fang; LI Yan-bo; HAN Jun-yong; YIN Jia-jing; WANG Yang; ZHU Li-bo; XIE Guang-ying

    2013-01-01

    Background The pathophysiology of type 2 diabetes is progressive pancreatic beta cell failure with consequential reduced insulin secretion.Glucotoxicity results in the reduction of beta cell mass in type 2 diabetes by inducing apoptosis.Autophagy is essential for the maintenance of normal islet architecture and plays a crucial role in maintaining the intracellular insulin content by accelerating the insulin degradation rate in beta cells.Recently more attention has been paid to the effect of autophagy in type 2 diabetes.The regulatory pathway of autophagy in controlling pancreatic beta cells is still not clear.The aim of our study was to evaluate whether liraglutide can inhibit apoptosis and modulate autophagy in vitro in insulinoma cells (INS-1 cells).Methods INS-1 cells were incubated for 24 hours in the presence or absence of high levels of glucose,liraglutide (a long-acting human glucagon-like peptide-1 analogue),or 3-methyadenine (3-MA).Cell viability was measured using the Cell Counting Kit-8 (CCK8) viability assay.Autophagy of INS-1 cells was tested by monodansylcadaverine (MDC)staining,an autophagy fluorescent compound used for the labeling of autophagic vacuoles,and by Western blotting of microtubule-associated protein I light chain 3 (LC3),a biochemical markers of autophagic initiation.Results The viability of INS-1 cells was reduced after treatment with high levels of glucose.The viability of INS-1 cells was reduced and apoptosis was increased when autophagy was inhibited.The viability of INS-1 cells was significantly increased by adding liraglutide to supplement high glucose level medium compared with the cells treated with high glucose levels alone.Conclusions Apoptosis and autophagy were increased in rat INS-1 cells when treated with high level of glucose,and the viability of INS-1 cells was significantly reduced by inhibiting autophagy.Liraglutide protected INS-1 cells from high glucose level-induced apoptosis that is accompanied by a significant

  6. A Comprehensive Review of Optical Stretcher for Cell Mechanical Characterization at Single-Cell Level

    Directory of Open Access Journals (Sweden)

    Tie Yang

    2016-05-01

    Full Text Available This paper presents a comprehensive review of the development of the optical stretcher, a powerful optofluidic device for single cell mechanical study by using optical force induced cell stretching. The different techniques and the different materials for the fabrication of the optical stretcher are first summarized. A short description of the optical-stretching mechanism is then given, highlighting the optical force calculation and the cell optical deformability characterization. Subsequently, the implementations of the optical stretcher in various cell-mechanics studies are shown on different types of cells. Afterwards, two new advancements on optical stretcher applications are also introduced: the active cell sorting based on cell mechanical characterization and the temperature effect on cell stretching measurement from laser-induced heating. Two examples of new functionalities developed with the optical stretcher are also included. Finally, the current major limitation and the future development possibilities are discussed.

  7. Licochalcone A induces T24 bladder cancer cell apoptosis by increasing intracellular calcium levels.

    Science.gov (United States)

    Yang, Xinhui; Jiang, Jiangtao; Yang, Xinyan; Han, Jichun; Zheng, Qiusheng

    2016-07-01

    Licochalcone A (LCA) has been reported to significantly inhibit cell proliferation, increase reactive oxygen species (ROS) levels, and induce apoptosis of T24 human bladder cancer cells via mitochondria and endoplasmic reticulum (ER) stress-triggered signaling pathways. Based on these findings, the present study aimed to investigate the mechanisms by which LCA induces apoptosis of T24 cells. Cultured T24 cells were treated with LCA, and cell viability was measured using the sulforhodamine B assay. Apoptosis was detected by flow cytometry with Annexin V/propidium iodide staining, and by fluorescent microscopy with Hoechst 33258 staining. The levels of intracellular free calcium ions were determined using Fluo-3 AM dye marker. Intracellular ROS levels were assessed using the 2',7'-dichlorodihydrofluorescein diacetate probe assay. The mitochondrial membrane potential was measured using 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl benzimidazole carbocyanine iodide. Furthermore, the mRNA expression levels of B‑cell lymphoma (Bcl)‑extra large, Bcl‑2‑associated X protein, Bcl‑2‑interacting mediator of cell death, apoptotic protease activating factor‑1 (Apaf‑1), calpain 2, cysteinyl aspartate specific proteinase (caspase)‑3, caspase‑4 and caspase‑9 were determined using reverse transcription semiquantitative and quantitative polymerase chain reaction analyses. Treatment with LCA inhibited proliferation and induced apoptosis of T24 cells, and increased intracellular Ca2+ levels and ROS production. Furthermore, LCA induced mitochondrial dysfunction, decreased mitochondrial membrane potential, and increased the mRNA expression levels of Apaf‑1, caspase‑9 and caspase‑3. Exposure of T24 cells to LCA also triggered calpain 2 and caspase‑4 activation, resulting in apoptosis. These findings indicated that LCA increased intracellular Ca2+ levels, which may be associated with mitochondrial dysfunction. In addition, the ER stress pathway may be

  8. Quantification of active caspases in stem cells: single cell analysis at femtogram level

    Czech Academy of Sciences Publication Activity Database

    Adamová, Eva; Lišková, Marcela; Klepárník, Karel; Hampl, Aleš; Matalová, Eva

    Leipzig: Fraunhofer Institute for Cell Therapy and Immunology, 2013. s. 358. [World Conference on Regenerative Medicine 2013 /WCRM 2013/. 23.10.2013-25.10.2013, Leipzig] R&D Projects: GA ČR GAP206/11/2377; GA ČR GAP304/11/1418 Institutional support: RVO:68081715 ; RVO:67985904 ; RVO:68378041 Keywords : caspases * cell analysis * cancer cell Subject RIV: CB - Analytical Chemistry, Separation; EA - Cell Biology (UZFG-Y); EB - Genetics ; Molecular Biology (UEM-P)

  9. Cell-to-cell diversity in protein levels of a gene driven by a tetracycline inducible promoter

    Directory of Open Access Journals (Sweden)

    Yli-Harja Olli

    2011-05-01

    Full Text Available Abstract Background Gene expression in Escherichia coli is regulated by several mechanisms. We measured in single cells the expression level of a single copy gene coding for green fluorescent protein (GFP, integrated into the genome and driven by a tetracycline inducible promoter, for varying induction strengths. Also, we measured the transcriptional activity of a tetracycline inducible promoter controlling the transcription of a RNA with 96 binding sites for MS2-GFP. Results The distribution of GFP levels in single cells is found to change significantly as induction reaches high levels, causing the Fano factor of the cells' protein levels to increase with mean level, beyond what would be expected from a Poisson-like process of RNA transcription. In agreement, the Fano factor of the cells' number of RNA molecules target for MS2-GFP follows a similar trend. The results provide evidence that the dynamics of the promoter complex formation, namely, the variability in its duration from one transcription event to the next, explains the change in the distribution of expression levels in the cell population with induction strength. Conclusions The results suggest that the open complex formation of the tetracycline inducible promoter, in the regime of strong induction, affects significantly the dynamics of RNA production due to the variability of its duration from one event to the next.

  10. Gene mutations and increased levels of p53 protein in human squamous cell carcinomas and their cell lines.

    OpenAIRE

    Burns, J E; Baird, M. C.; Clark, L. J.; Burns, P A; Edington, K.; Chapman, C; Mitchell, R; Robertson, G; Soutar, D; Parkinson, E. K.

    1993-01-01

    Using immunocytochemical and Western blotting techniques we have demonstrated the presence of abnormally high levels of p53 protein in 8/24 (33%) of human squamous cell carcinomas (SCC) and 9/18 (50%) of SCC cell lines. There was a correlation between the immunocytochemical results obtained with eight SCC samples and their corresponding cell lines. Direct sequencing of PCR-amplified, reverse transcribed, p53 mRNA confirmed the expression of point mutations in six of the positive cell lines an...

  11. Isolation of mammalian cell variants with enhanced endogenous thiol content at low survival levels following irradiation

    International Nuclear Information System (INIS)

    Approximately half of a group of Chinese hamster V79 cell clones isolated from radiation survivors at low surviving fractions had significantly higher endogenous levels of non-protein and protein thiols than unirradiated cells. A similar group of cell lines cloned from unirradiated cells had thiol levels in the same range as the original unirradiated population. In some cases, clones isolated following irradiation are also more resistant to misonidazole toxicity and to radiation. This phenotype can persist through many cell generations for weeks or months of continuous growth; however, in many clones with altered phenotypes isolated following irradiation, reversion to the same phenotype as that of unirradiated populations has been observed. Induction of elevated thiol levels in tumours by radiotherapy could reduce both efficacy of the radiation itself and of radiation-modifying or chemotherapeutic drugs given in combination with radiation. (author)

  12. Does ploidy level directly control cell size? Counterevidence from Arabidopsis genetics.

    Directory of Open Access Journals (Sweden)

    Hirokazu Tsukaya

    Full Text Available Ploidy level affects cell size in many organisms, and ploidy-dependent cell enlargement has been used to breed many useful organisms. However, how polyploidy affects cell size remains unknown. Previous studies have explored changes in transcriptome data caused by polyploidy, but have not been successful. The most naïve theory explaining ploidy-dependent cell enlargement is that increases in gene copy number increase the amount of protein, which in turn increases the cell volume. This hypothesis can be evaluated by examining whether any strains, mutants, or transgenics show the same cell size before and after a tetraploidization event. I performed this experiment by tetraploidizing various mutants and transgenics of Arabidopsis thaliana, which show a wide range in cell size, and found that the ploidy-dependent increase in cell volume is genetically regulated. This result is not in agreement with the theory described above.

  13. Rule-based multi-level modeling of cell biological systems

    Directory of Open Access Journals (Sweden)

    Maus Carsten

    2011-10-01

    Full Text Available Abstract Background Proteins, individual cells, and cell populations denote different levels of an organizational hierarchy, each of which with its own dynamics. Multi-level modeling is concerned with describing a system at these different levels and relating their dynamics. Rule-based modeling has increasingly attracted attention due to enabling a concise and compact description of biochemical systems. In addition, it allows different methods for model analysis, since more than one semantics can be defined for the same syntax. Results Multi-level modeling implies the hierarchical nesting of model entities and explicit support for downward and upward causation between different levels. Concepts to support multi-level modeling in a rule-based language are identified. To those belong rule schemata, hierarchical nesting of species, assigning attributes and solutions to species at each level and preserving content of nested species while applying rules. Further necessities are the ability to apply rules and flexibly define reaction rate kinetics and constraints on nested species as well as species that are nested within others. An example model is presented that analyses the interplay of an intracellular control circuit with states at cell level, its relation to cell division, and connections to intercellular communication within a population of cells. The example is described in ML-Rules - a rule-based multi-level approach that has been realized within the plug-in-based modeling and simulation framework JAMES II. Conclusions Rule-based languages are a suitable starting point for developing a concise and compact language for multi-level modeling of cell biological systems. The combination of nesting species, assigning attributes, and constraining reactions according to these attributes is crucial in achieving the desired expressiveness. Rule schemata allow a concise and compact description of complex models. As a result, the presented approach

  14. DNA level in guard cells nuclei of Ornithogalum umbellatum ovary is 2C-4C

    Directory of Open Access Journals (Sweden)

    Maria Kwiatkowska

    2011-04-01

    Full Text Available The DNA content after Feulgen reaction in the guard cells and epidermis of Omithogalum umbellatum ovary was cytophotometrically measured in different phases of flower development. Only in bud of flowers guard cells DNA content was 2C while in full blown flowers it was higher, between 2C-4C. This observation was supported by autoradiographic studies with 3H-thymidine which was incorporated into guard cell nuclei in the ovary epidermis of newly developed flowers. Thus DNA level in O. umbellatum guard cells was higher than those in other plants described in literature. On the other hand, DNA content in the epidermis cells increased gradually with ovary growth reaching the maximum level of 8C in some cells.

  15. Cell-cell channels, viruses, and evolution: via infection, parasitism, and symbiosis toward higher levels of biological complexity.

    Science.gov (United States)

    Baluska, Frantisek

    2009-10-01

    Between prokaryotic cells and eukaryotic cells there is dramatic difference in complexity which represents a problem for the current version of the cell theory, as well as for the current version of evolution theory. In the past few decades, the serial endosymbiotic theory of Lynn Margulis has been confirmed. This results in a radical departure from our understanding of living systems: the eukaryotic cell represents de facto"cells-within-cell." Higher order "cells-within-cell" situations are obvious at the eukaryotic cell level in the form of secondary and tertiary endosymbiosis, or in the male and female gametophytes of higher plants. The next challenge of the current version of the cell theory is represented by the fact that the multicellular fungi and plants are, in fact, supracellular assemblies as their cells are not physically separated from each other. Moreover, there are also examples of alliances and mergings between multicellular organisms. Infection, especially the viral one, but also bacterial and fungal infections, followed by symbiosis, is proposed to act as the major force that drives the biological evolution toward higher complexity. PMID:19845631

  16. Levels of polyunsaturated fatty acids correlate with growth rate in plant cell cultures

    Science.gov (United States)

    Meï, Coline; Michaud, Morgane; Cussac, Mathilde; Albrieux, Catherine; Gros, Valérie; Maréchal, Eric; Block, Maryse A.; Jouhet, Juliette; Rébeillé, Fabrice

    2015-01-01

    In higher plants, fatty acids (FAs) with 18 carbons (18C) represent about 70% of total FAs, the most abundant species being 18:2 and 18:3. These two polyunsaturated FAs (PUFAs) represent about 55% of total FAs in Arabidopsis cell suspension cultures, whereas 18:1 represents about 10%. The level of PUFAs may vary, depending on ill-defined factors. Here, we compared various sets of plant cell cultures and noticed a correlation between the growth rate of a cell population and the level of unsaturation of 18C FAs. These observations suggest that the final level of PUFAs might depend in part on the rate of cell division, and that FAD2 and FAD3 desaturases, which are respectively responsible for the formation of 18:2 and 18:3 on phospholipids, have limiting activities in fast-growing cultures. In plant cell culture, phosphate (Pi) deprivation is known to impair cell division and to trigger lipid remodeling. We observed that Pi starvation had no effect on the expression of FAD genes, and that the level of PUFAs in this situation was also correlated with the growth rate. Thus, the level of PUFAs appears as a hallmark in determining cell maturity and aging. PMID:26469123

  17. Multiweek Cell Culture Project for Use in Upper-Level Biology Laboratories

    Science.gov (United States)

    Marion, Rebecca E.; Gardner, Grant E.; Parks, Lisa D.

    2012-01-01

    This article describes a laboratory protocol for a multiweek project piloted in a new upper-level biology laboratory (BIO 426) using cell culture techniques. Human embryonic kidney-293 cells were used, and several culture media and supplements were identified for students to design their own experiments. Treatments included amino acids, EGF,…

  18. TRPV Channels in Mast Cells as a Target for Low-Level-Laser Therapy

    OpenAIRE

    Lina Wang; Di Zhang; Wolfgang Schwarz

    2014-01-01

    Low-level laser irradiation in the visible as well as infrared range is applied to skin for treatment of various diseases. Here we summarize and discuss effects of laser irradiation on mast cells that leads to degranulation of the cells. This process may contribute to initial steps in the final medical effects. We suggest that activation of TRPV channels in the mast cells forms a basis for the underlying mechanisms and that released ATP and histamine may be putative mediators for therapeutic ...

  19. Basal HIF-1a expression levels are not predictive for radiosensitivity of human cancer cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Schilling, D.; Multhoff, G. [Klinikum rechts der Isar der Technischen Univ. Muenchen (Germany). Dept. of Radiation Oncology; Helmholtz Center Munich, CCG - Innate Immunity in Tumor Biology, Munich (Germany). German Research Center for Environmental Health - Inst. of Pathology; Bayer, C.; Emmerich, K.; Molls, M.; Vaupel, P. [Klinikum rechts der Isar der Technischen Univ. Muenchen (Germany). Dept. of Radiation Oncology; Huber, R.M. [Klinikum der Univ. Muenchen (Germany). Dept. of Pneumology

    2012-04-15

    High levels of hypoxia inducible factor (HIF)-1a in tumors are reported to be associated with tumor progression and resistance to therapy. To examine the impact of HIF-1a on radioresistance under normoxia, the sensitivity towards irradiation was measured in human tumor cell lines that differ significantly in their basal HIF-1a levels. HIF-1a levels were quantified in lysates of H1339, EPLC-272H, A549, SAS, XF354, FaDu, BHY, and CX- tumor cell lines by ELISA. Protein levels of HIF-1a, HIF-2a, carbonic anhydrase IX (CA IX), and GAPDH were assessed by Western blot analysis. Knock-down experiments were performed using HIF-1a siRNA. Clonogenic survival after irradiation was determined by the colony forming assay. According to their basal HIF-1a status, the tumor cell lines were divided into low (SAS, XF354, FaDu, A549, CX-), intermediate (EPLC-272H, BHY), and high (H1339) HIF-1a expressors. The functionality of the high basal HIF-1a expression in H1339 cells was proven by reduced CA IX expression after knocking-down HIF-1a. Linear regression analysis revealed no correlation between basal HIF-1a levels and the survival fraction at either 2 or 4 Gy in all tumor cell lines investigated. Our data suggest that basal HIF-1a levels in human tumor cell lines do not predict their radiosensitivity under normoxia. (orig.)

  20. Intracellular ATP levels are a pivotal determinant of chemoresistance in colon cancer cells.

    Science.gov (United States)

    Zhou, Yunfei; Tozzi, Federico; Chen, Jinyu; Fan, Fan; Xia, Ling; Wang, Jinrong; Gao, Guang; Zhang, Aijun; Xia, Xuefeng; Brasher, Heather; Widger, William; Ellis, Lee M; Weihua, Zhang

    2012-01-01

    Altered metabolism in cancer cells is suspected to contribute to chemoresistance, but the precise mechanisms are unclear. Here, we show that intracellular ATP levels are a core determinant in the development of acquired cross-drug resistance of human colon cancer cells that harbor different genetic backgrounds. Drug-resistant cells were characterized by defective mitochondrial ATP production, elevated aerobic glycolysis, higher absolute levels of intracellular ATP, and enhanced HIF-1α-mediated signaling. Interestingly, direct delivery of ATP into cross-chemoresistant cells destabilized HIF-1α and inhibited glycolysis. Thus, drug-resistant cells exhibit a greater "ATP debt" defined as the extra amount of ATP needed to maintain homeostasis of survival pathways under genotoxic stress. Direct delivery of ATP was sufficient to render drug-sensitive cells drug resistant. Conversely, depleting ATP by cell treatment with an inhibitor of glycolysis, 3-bromopyruvate, was sufficient to sensitize cells cross-resistant to multiple chemotherapeutic drugs. In revealing that intracellular ATP levels are a core determinant of chemoresistance in colon cancer cells, our findings may offer a foundation for new improvements to colon cancer treatment. PMID:22084398

  1. Automatic Detection of Cervical Cancer Cells by a Two-Level Cascade Classification System

    Science.gov (United States)

    Su, Jie; Xu, Xuan; He, Yongjun; Song, Jinming

    2016-01-01

    We proposed a method for automatic detection of cervical cancer cells in images captured from thin liquid based cytology slides. We selected 20,000 cells in images derived from 120 different thin liquid based cytology slides, which include 5000 epithelial cells (normal 2500, abnormal 2500), lymphoid cells, neutrophils, and junk cells. We first proposed 28 features, including 20 morphologic features and 8 texture features, based on the characteristics of each cell type. We then used a two-level cascade integration system of two classifiers to classify the cervical cells into normal and abnormal epithelial cells. The results showed that the recognition rates for abnormal cervical epithelial cells were 92.7% and 93.2%, respectively, when C4.5 classifier or LR (LR: logical regression) classifier was used individually; while the recognition rate was significantly higher (95.642%) when our two-level cascade integrated classifier system was used. The false negative rate and false positive rate (both 1.44%) of the proposed automatic two-level cascade classification system are also much lower than those of traditional Pap smear review. PMID:27298758

  2. The stem cell niche: tissue physiology at a single cell level

    OpenAIRE

    Hoggatt, Jonathan; Scadden, David T.

    2012-01-01

    Stem cells are the critical unit affecting tissue maintenance, regeneration, and repair, with particular relevance to the tissues with high cell turnover. Stem cell regulation accommodates the conflicting needs of prompt responsiveness to injury and long-term preservation through quiescence. They are, in essence, the fundamental unit by which a tissue handles changing physiologic needs throughout the lifetime of the organism. As such, they are the focal point of dynamic tissue function, and t...

  3. Correlation between residual level of DNA double-strand breaks and the radiosensitivity of cancer cells

    International Nuclear Information System (INIS)

    Objective: To understand the variation of the DNA double-strand break rejoining capacity among different cultured cancer cell lines and the primary cancer cells from brain cancer patients, and to explore the predictor of radiotherapy responses of cancers. Methods: DNA double-strand breaks (DSBs) were induced by 60Co γ-irradiation. Pulsed-field gel electrophoresis was used to analyze the initial production and rejoining of DNA DSBs. Radiosensitivity was determined by in vitro assay of clonogenic-forming capacity. Results: A wide variation of radiosensitivity, e.g. the survival parameter of Do varied from 0.65 to 2.15 Gy, was displayed among the eight cell lines derived from different type of cancers. Although differential level of initial DNA DSBs induced by 20 Gy γ-rays was observed among various cell lines, it was not correlated with the radiosensitivity. The deficiency of DNA DSB rejoining in radiosensitive cell lines was shown either in the early rapid-rejoining phase (SX-10 cells) or in the late slow-rejoining phase (A2780 cells). A significant relationship was observed between the residual level of DNA DSBs measured at 2 h post-20 Gy irradiation and the cellular radiosensitivity (D0 or SF2). The kinetic curves of rejoining DNA DSBs in the primary human brain tumor cells indicated a variation on DSB rejoining capacity among different individual tumor. The residual level of DNA DSBs after 2 h of rejoining post 20 Gy irradiation in primary human brain tumor cells is compatible to the results obtained in vitro culture cancer cell lines. Conclusions: The residual level of DNA DSBs is correlated with radioresistance of cancer cells, and the residual DNA damage is a useful parameter in predicting the response of tumor tissue to radiotherapy. (authors)

  4. Analysis of gene expression levels in individual bacterial cells without image segmentation

    International Nuclear Information System (INIS)

    Highlights: ► We present a method for extracting gene expression data from images of bacterial cells. ► The method does not employ cell segmentation and does not require high magnification. ► Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. ► We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

  5. Analysis of gene expression levels in individual bacterial cells without image segmentation

    Energy Technology Data Exchange (ETDEWEB)

    Kwak, In Hae; Son, Minjun [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States); Hagen, Stephen J., E-mail: sjhagen@ufl.edu [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States)

    2012-05-11

    Highlights: Black-Right-Pointing-Pointer We present a method for extracting gene expression data from images of bacterial cells. Black-Right-Pointing-Pointer The method does not employ cell segmentation and does not require high magnification. Black-Right-Pointing-Pointer Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. Black-Right-Pointing-Pointer We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

  6. Different roles of prepubertal and postpubertal germ cells and Sertoli cells in the regulation of serum inhibin B levels

    DEFF Research Database (Denmark)

    Andersson, A M; Müller, J; Skakkebaek, N E

    1998-01-01

    To elucidate the role of germ cells in the regulation of inhibin B secretion, serum inhibin B levels in prepubertal boys and adult men whom had a concurrent testicular biopsy showing either normal or impaired testicular function were compared. In addition, by immunohistochemistry the cellular...... localization of the two subunits of inhibin B (alpha and betaB) were examined in adult testicular tissue with normal spermatogenesis, spermatogenic arrest, or Sertoli cell only tubules (SCO) as well as in normal testicular tissue from an infant and a prepubertal boy. Adult men with testicular biopsy showing...... SCO boys, inhibin B levels were undetectable as in the adult SCO men. Intense inhibin alpha-subunit immunostaining was evident in Sertoli cells in both prepubertal and adult testes. In the prepubertal testis, positive immunostaining for the betaB-subunit was observed in Sertoli cells. In the adult...

  7. Levels of Ycg1 Limit Condensin Function during the Cell Cycle

    Science.gov (United States)

    Arsenault, Heather E.; Benanti, Jennifer A.

    2016-01-01

    During mitosis chromosomes are condensed to facilitate their segregation, through a process mediated by the condensin complex. Although several factors that promote maximal condensin activity during mitosis have been identified, the mechanisms that downregulate condensin activity during interphase are largely unknown. Here, we demonstrate that Ycg1, the Cap-G subunit of budding yeast condensin, is cell cycle-regulated with levels peaking in mitosis and decreasing as cells enter G1 phase. This cyclical expression pattern is established by a combination of cell cycle-regulated transcription and constitutive degradation. Interestingly, overexpression of YCG1 and mutations that stabilize Ycg1 each result in delayed cell-cycle entry and an overall proliferation defect. Overexpression of no other condensin subunit impacts the cell cycle, suggesting that Ycg1 is limiting for condensin complex formation. Consistent with this possibility, we find that levels of intact condensin complex are reduced in G1 phase compared to mitosis, and that increased Ycg1 expression leads to increases in both levels of condensin complex and binding to chromatin in G1. Together, these results demonstrate that Ycg1 levels limit condensin function in interphase cells, and suggest that the association of condensin with chromosomes must be reduced following mitosis to enable efficient progression through the cell cycle. PMID:27463097

  8. Helicobacter pylori induces mitochondrial DNA mutation and reactive oxygen species level in AGS cells

    OpenAIRE

    Xue-Wen Huang, Rui-Hua Luo, Qi Zhao, Zhong-Ze Shen, Li-Li Huang, Xian-Yuan An, Lan-Jing Zhao, Jie Wang, Yu-Zheng Huang

    2011-01-01

    To investigate the role of ROS in the helicobacter pylori (Hp) induced mtDNA mutations, AGS cells were treated by extracts of Hp11638 or Hp11638M. The ROS levels, cytochrome C reductions, and intracellular ATP levels were measured. The coding region and the D-Loop region were amplified and sequenced. Results showed the ROS levels, cytochrome C reduction and mtDNA mutations were markedly increased and cell viability decreased after treatment with both Hp extracts, and 616 mutations were detect...

  9. Assessment of flhDC mRNA levels in Serratia liquefaciens swarm cells

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim; Christensen, Allan Beck; Holmstrøm, K.; Eberl, Leo; Rasmussen, Thomas Bovbjerg; Sternberg, Claus; Heydorn, Arne; Molin, Søren; Givskov, Michael Christian

    2000-01-01

    We reported previously that artificial overexpression of the flhDC operon in liquid-grown Serratia liquefaciens resulted in the formation of filamentous, multinucleated, and hyperflagellated cells that were indistinguishable from surface-induced swarm cells (L. Eberl, G. Christiansen, S. Molin, and...... M. Givskov, J. Bacteriol. 178:554-559, 1996). In the present report we show by means of reporter gene measurements, Northern analysis, and in situ reverse transcription-PCR that the amount of flhDC mRNA in surface-grown swarm cells does not exceed the maximum level found in nondifferentiated......, vegetative cells. This suggests that surface-induced S. liquefaciens swarm cell differentiation, although dependent on flhDC gene expression, does not occur through elevated flhDC mRNA levels....

  10. Mannosylerythritol lipid increases levels of galactoceramide in and neurite outgrowth from PC12 pheochromocytoma cells.

    Science.gov (United States)

    Shibahara, M; Zhao, X; Wakamatsu, Y; Nomura, N; Nakahara, T; Jin, C; Nagaso, H; Murata, T; Yokoyama, K K

    2000-07-01

    We report here that a microbial extracellular glycolipid,mannosylerythritol lipid (MEL), induces the outgrowth ofneurites from and enhances the activity of acetylcholinesterase(AChE) in PC12 pheochromocytoma cells. Furthermore, treatment ofPC12 cells with MEL increased levels of galactosylceramide(Galbeta1-1'Cer; GalCer). Exposure of PC12 cells to exogenous GalCer caused the dose-dependent outgrowth ofneurites. By contrast, treatment of PC12 cells with nerve growthfactor (NGF) did not increase the level of GalCer in the cells. The neurite-related morphological changes induced by GalCerdifferend from those induced by NGF, indicating differencesbetween the signal transduction pathways triggered by NGF and by GalCer. PMID:19002832

  11. Association of expression levels of pluripotency/stem cell markers with the differentiation outcome of Wharton's jelly mesenchymal stem cells into insulin producing cells.

    Science.gov (United States)

    Kassem, Dina H; Kamal, Mohamed M; El-Kholy, Abd El-Latif G; El-Mesallamy, Hala O

    2016-08-01

    Recently, there has been much attention towards generation of insulin producing cells (IPCs) from stem cells, especially from Wharton's jelly mesenchymal stem cells (WJ-MSCs). However, generation of mature IPCs remains a challenge. Assessment of generation of IPCs was usually done by examining β-cell markers, however, assessment of pluripotency/stem cell markers drew less attention. Therefore, the purpose of this study was to investigate the levels of pluripotency/stem cell markers during differentiation of WJ-MSCs into IPCs and the association of these levels with differentiation outcomes. WJ-MSCs were isolated, characterized then induced to differentiate into IPCs using three different protocols namely A, B and C. Differentiated IPCs were assessed by the expression of pluripotency/stem cell markers, together with β-cell markers using qRT-PCR, and functionally by measuring glucose stimulated insulin secretion. Differentiated cells from protocol A showed lowest expression of pluripotency/stem cell markers and relatively best GSIS. However, protocol B showed concomitant expression of pluripotency/stem cell and β-cell markers with relatively less insulin secretion as compared to protocol A. Protocol C failed to generate glucose-responsive IPCs. In conclusion, sustained expression of pluripotency/stem cell markers could be associated with the incomplete differentiation of WJ-MSCs into IPCs. A novel finding for which further investigations are warranted. PMID:27265786

  12. Laminin increases both levels and activity of tyrosine hydroxylase in calf adrenal chromaffin cells

    OpenAIRE

    1986-01-01

    We have investigated the effects of substrate-bound laminin on levels of enzymes of the catecholamine biosynthetic pathway in primary cultures of calf adrenal chromaffin cells. Laminin increases the levels of the enzymes tyrosine hydroxylase, dopamine-beta-hydroxylase, and phenylethanolamine-N-methyl-transferase. This effect is selective, in that levels of other enzymes (lactate dehydrogenase, aromatic amino acid decarboxylase, and acetylcholinesterase) are not increased. The effect of lamini...

  13. Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

    OpenAIRE

    K.S. Canuto; L.P.S. Sergio; O.R. Guimarães; Geller, M.; De Paoli, F; Fonseca, A.S.

    2015-01-01

    Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC) radiati...

  14. Hydroxyl radicals cause fluctuation in intracellular ferrous ion levels upon light exposure during photoreceptor cell death.

    Science.gov (United States)

    Imamura, Tomoyo; Hirayama, Tasuku; Tsuruma, Kazuhiro; Shimazawa, Masamitsu; Nagasawa, Hideko; Hara, Hideaki

    2014-12-01

    Iron accumulation is a potential pathogenic event often seen in age-related macular degeneration (AMD) patients. In this study, we focused on the relationship between AMD pathology and concentrations of ferrous ion, which is a highly reactive oxygen generator in biological systems. Murine cone-cells-derived 661 W cells were exposed to white fluorescence light at 2500 lx for 1, 3, 6, or 12 h. Levels of ferrous ions, reactive oxygen species (ROS), and hydroxyl radicals were detected by RhoNox-1, a novel fluorescent probe for the selective detection of ferrous ion, 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate, acetyl ester (CM-H2DCFDA), and 3'-p-(aminophenyl) fluorescein, respectively. Reduced glutathione, total iron levels and photoreceptor cell death were also measured. Two genes related to iron metabolism, transferrin receptor 1 (TfR1) and H ferritin (HFt), were quantified by RT-PCR. The effects of ferrous ion on cell death and hydroxyl radical production were determined by treatment with a ferrous ion chelating agent, 2,2'-bipyridyl. We found that the ferrous ion level decreased with light exposure in the short time frame, whereas it was upregulated during a 6-h light exposure. Total iron, ROS, cell death rate, and expression of TfR and HFt genes were significantly increased in a time-dependent manner in 661 W cells exposed to light. Chelation with 2,2'-bipyridyl reduced the level of hydroxyl radicals and protected against light-induced cell death. These results suggest that light exposure decreases ferrous ion levels and enhances iron uptake in photoreceptor cells. Ferrous ion may be involved in light-induced photoreceptor cell death through production of hydroxyl radicals. PMID:25447561

  15. Mannosylerythritol lipid increases levels of galactoceramide in and neurite outgrowth from PC12 pheochromocytoma cells

    OpenAIRE

    Shibahara, Miki; Zhao, Xiaoxian; Wakamatsu, Yoko; Nomura, Nobuhiko; Nakahara, Tadaatsu; Jin, Chunyuan; Nagaso, Hideyuki; Murata, Takehide; Yokoyama, Kazunari K.

    2000-01-01

    We report here that a microbial extracellular glycolipid,mannosylerythritol lipid (MEL), induces the outgrowth ofneurites from and enhances the activity of acetylcholinesterase(AChE) in PC12 pheochromocytoma cells. Furthermore, treatment ofPC12 cells with MEL increased levels of galactosylceramide(Galβ1-1′Cer; GalCer). Exposure of PC12 cells to exogenous GalCer caused the dose-dependent outgrowth ofneurites. By contrast, treatment of PC12 cells with nerve growthfactor (NGF) did not increase t...

  16. Bisphenol A alters transcript levels of biomarker genes for Major Depressive Disorder in vascular endothelial cells and colon cancer cells.

    Science.gov (United States)

    Ribeiro-Varandas, Edna; Pereira, H Sofia; Viegas, Wanda; Delgado, Margarida

    2016-06-01

    Bisphenol A (BPA) is capable of mimicking endogenous hormones with potential consequences for human health and BPA exposure has been associated with several human diseases including neuropsychiatric disorders. Here, quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) results show that BPA at low concentrations (10 ng/mL and 1 μg/mL) induces differential transcript levels of four biomarker genes for Major Depressive Disorder (MDD) in HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). These results substantiate increasing concerns of BPA exposure in levels currently detected in humans. PMID:27010169

  17. Effect of LLLT on the level of ATP and ROS from organ of corti cells

    Science.gov (United States)

    Rhee, ChungKu; Chang, So-Young; Ahn, Jin-Chul; Suh, Myung-Whan; Jung, Jae Yun

    2014-03-01

    It is well established that ototoxic antibiotics and acoustic trauma can damage cochlear hair cells and cause hearing loss. Previous studies using transcanal LLLT (Low level laser therapy) showed that LLLT can promote recovery of hearing thresholds and cochlear hair cells. However, its mechanism has not been studied. Aim: The aim of this study is to investigate the mechanism of hearing recovery from gentamicin induced ototoxic hearing loss by LLLT. Methods: HEI- OC1 (House ear institute organ of Corti) cells were cultured for 18 hours and ototoxicity was induced by gentamicin (GM) treatment to the cells. Cultured cells were divided into 6 groups, No treatment control, LLLT only, GM 6.6 mM and GM 13.1 mM, GM 6.6 mM+LLLT and GM 13.1 mM+LLLT cells. LD laser 808 nm, 15 mW, was irradiated to the cultured cells for 15 min, at 4 hours after GM treatment to the cells. ATP was assayed using the ATP assay Kit. ROS was measured using confocal microscope after application of H2DCFDA dye. Results: ATP was decreased in GM 13.1 mM cells and increased in LLLT only cells and GM 13.1 mM+LLLT cells compared to control and 13.1 mM cells. ROS was increased in GM 6.6 mM and GM 13.1 mM cells, and decreased in GM 6.6 mM+LLLT and GM 13.1 mM+LLLT cells compared to GM 6.6 and 13.1 mM cells immediately after laser irradiation. Conclusion: This study demonstrated that LLLT on GM treated HEI-OC1 cells increased ATP and decreased ROS that may contribute to the recovery of hearing.

  18. Emergence of cytotoxic resistance in cancer cell populations: Single-cell mechanisms and population-level consequences

    Science.gov (United States)

    Lorenzi, Tommaso; Chisholm, Rebecca H.; Lorz, Alexander; Larsen, Annette K.; de Almeida, Luís Neves; Escargueil, Alexandre; Clairambault, Jean

    2016-06-01

    We formulate an individual-based model and a population model of phenotypic evolution, under cytotoxic drugs, in a cancer cell population structured by the expression levels of survival-potential and proliferation-potential. We apply these models to a recently studied experimental system. Our results suggest that mechanisms based on fundamental laws of biology can reversibly push an actively-proliferating, and drug-sensitive, cell population to transition into a weakly-proliferative and drug-tolerant state, which will eventually facilitate the emergence of more potent, proliferating and drug-tolerant cells.

  19. How low-level laser therapy can change mechanical properties of cells

    Science.gov (United States)

    de Magalhães, Ana Carolina; Martinez, Diana; Ferreira, Marcia Z. J.; Yoshimura, Ellisabeth M.; Alencar, Adriano M.; Chavantes, Maria Cristina

    2013-03-01

    Low level laser therapy is used as a treatment of several conditions, including inflammatory processes and wound healing. Possible changes in mechanical properties of cells, caused by illumination, are investigated with optical magnetic twisting cytometry (OMTC), which is a technique used to evaluate mechanical properties in cell culture. Ferromagnetic micro beads are bound to cell cytoskeleton, the beads are magnetized vertically and a horizontal twisting magnetic field is applied causing a torque that moves the beads and deforms the cell, the beads rotate and displace. Based on the lateral displacement of the beads, elastic shear and loss moduli are obtained. Samples of human bronchial epithelial cell culture were divided in two groups: one was illuminated with a 660 nm red laser, 30 mW power, 0.75 W/cm2 irradiance, during different time intervals, and the other one, the control group, was not illuminated. The values of the mechanical constants of the cells of the control group showed a tendency of increasing with the time out of the incubator. On the other hand, the illuminated group showed constancy on the behavior of both moduli, keeping the normal conditions of the cell culture. Those results indicate that illumination can induce cells to homeostasis, and OMTC is sensitive to observe departures from the steady conditions. Hence, OMTC is an important technique which can be used to aggregate knowledge on the light effect in cell cytoskeleton and even on the low level laser therapy mechanisms in inflammatory processes and/or wound healing.

  20. The Effects of High Steady State Auxin Levels on Root Cell Elongation in Brachypodium.

    Science.gov (United States)

    Pacheco-Villalobos, David; Díaz-Moreno, Sara M; van der Schuren, Alja; Tamaki, Takayuki; Kang, Yeon Hee; Gujas, Bojan; Novak, Ondrej; Jaspert, Nina; Li, Zhenni; Wolf, Sebastian; Oecking, Claudia; Ljung, Karin; Bulone, Vincent; Hardtke, Christian S

    2016-05-01

    The long-standing Acid Growth Theory of plant cell elongation posits that auxin promotes cell elongation by stimulating cell wall acidification and thus expansin action. To date, the paucity of pertinent genetic materials has precluded thorough analysis of the importance of this concept in roots. The recent isolation of mutants of the model grass species Brachypodium distachyon with dramatically enhanced root cell elongation due to increased cellular auxin levels has allowed us to address this question. We found that the primary transcriptomic effect associated with elevated steady state auxin concentration in elongating root cells is upregulation of cell wall remodeling factors, notably expansins, while plant hormone signaling pathways maintain remarkable homeostasis. These changes are specifically accompanied by reduced cell wall arabinogalactan complexity but not by increased proton excretion. On the contrary, we observed a tendency for decreased rather than increased proton extrusion from root elongation zones with higher cellular auxin levels. Moreover, similar to Brachypodium, root cell elongation is, in general, robustly buffered against external pH fluctuation in Arabidopsis thaliana However, forced acidification through artificial proton pump activation inhibits root cell elongation. Thus, the interplay between auxin, proton pump activation, and expansin action may be more flexible in roots than in shoots. PMID:27169463

  1. The Effects of High Steady State Auxin Levels on Root Cell Elongation in Brachypodium[OPEN

    Science.gov (United States)

    Pacheco-Villalobos, David; Tamaki, Takayuki; Gujas, Bojan; Jaspert, Nina; Oecking, Claudia; Bulone, Vincent; Hardtke, Christian S.

    2016-01-01

    The long-standing Acid Growth Theory of plant cell elongation posits that auxin promotes cell elongation by stimulating cell wall acidification and thus expansin action. To date, the paucity of pertinent genetic materials has precluded thorough analysis of the importance of this concept in roots. The recent isolation of mutants of the model grass species Brachypodium distachyon with dramatically enhanced root cell elongation due to increased cellular auxin levels has allowed us to address this question. We found that the primary transcriptomic effect associated with elevated steady state auxin concentration in elongating root cells is upregulation of cell wall remodeling factors, notably expansins, while plant hormone signaling pathways maintain remarkable homeostasis. These changes are specifically accompanied by reduced cell wall arabinogalactan complexity but not by increased proton excretion. On the contrary, we observed a tendency for decreased rather than increased proton extrusion from root elongation zones with higher cellular auxin levels. Moreover, similar to Brachypodium, root cell elongation is, in general, robustly buffered against external pH fluctuation in Arabidopsis thaliana. However, forced acidification through artificial proton pump activation inhibits root cell elongation. Thus, the interplay between auxin, proton pump activation, and expansin action may be more flexible in roots than in shoots. PMID:27169463

  2. Effects of Estrogen Level on the Function of Vascular Endothelial Cells and Expression of Vascular Cell Adhesion Molecule - 1φ

    Institute of Scientific and Technical Information of China (English)

    WU Saizhu(吴赛珠); LIU Jiangguo(刘建国); TAN Jiayu(谭家余); ZHoU Kexiang(周可祥); Gorge D Webb; WEI Heming(隗和明); GUO Zhiguang(郭志刚)

    2002-01-01

    Objectives To ob- serve the effect of different estrogen levels on the se- cretory function of vascular endothelial cells of female rats, and study the effect of modulation of estrogen level on the expression of vascular cell adhesion molecule - 1 and the concentration of estrogen receptorin vascular endothelial cells. Methods Radioim-munology was used to measure the serum concentrationof endothelin and PGI2, and copper-cadmium re-duction was employed to measure the serum content ofnitrogen monoxide. Radioligand binding and flowcy-tometry were used to measure the expression of estrogenreceptor and vascular cell adhesion molecule (VCAM-1 ) of vascular endothelial cells respectively. Re-sults 1. The serum concentration of nitric oxide andPGI2 decreased when the ovaries of female rats wereremoved. In ovariectomized rats, given estrogen, theconcentration rose ( P < 0.05), but the plasma con-centration of endothelin was adverse to it. 2. Theconcentration of estrogen receptor of vascular endothe-lial cells decreased remarkably when the ovaries of fe-male rats were removed. When given estrogen, it in-creased. 3. The percent of expressed VCAM - 1 in-creased siguificantly after interleukin- lβoperated onthe cells, but 17 - βestradiol at 3 × 10-8 ~ 10-6 mol/lall decreased the percent. Conclusions Estrogenlevel can influence the secretion of nitrogen monoxide,PGI2 and endothlin of vascular endothelial cells, andalso influence the concentration of estrogen receptor ofvascular endothelial cells. 17 -β Estradiol at 3 × 10-8~ 10-6 M can decrease the elevation of VCAM - 1 ofvascular endothelial cells induced by interleukin - 1 β.

  3. Structure of the first representative of Pfam family PF04016 (DUF364) reveals enolase and Rossmann-like folds that combine to form a unique active site with a possible role in heavy-metal chelation

    International Nuclear Information System (INIS)

    The crystal structure of the first representative of DUF364 family reveals a combination of enolase N-terminal-like and C-terminal Rossmann-like folds. Analysis of the interdomain cleft combined with sequence and genome context conservation among homologs, suggests a unique catalytic site likely involved in the synthesis of a flavin or pterin derivative. The crystal structure of Dhaf4260 from Desulfitobacterium hafniense DCB-2 was determined by single-wavelength anomalous diffraction (SAD) to a resolution of 2.01 Å using the semi-automated high-throughput pipeline of the Joint Center for Structural Genomics (JCSG) as part of the NIGMS Protein Structure Initiative (PSI). This protein structure is the first representative of the PF04016 (DUF364) Pfam family and reveals a novel combination of two well known domains (an enolase N-terminal-like fold followed by a Rossmann-like domain). Structural and bioinformatic analyses reveal partial similarities to Rossmann-like methyltransferases, with residues from the enolase-like fold combining to form a unique active site that is likely to be involved in the condensation or hydrolysis of molecules implicated in the synthesis of flavins, pterins or other siderophores. The genome context of Dhaf4260 and homologs additionally supports a role in heavy-metal chelation

  4. Levels and values of circulating endothelial progenitor cells, soluble angiogenic factors, and mononuclear cell apoptosis in liver cirrhosis patients

    Directory of Open Access Journals (Sweden)

    Chen Chih-Hung

    2012-07-01

    Full Text Available Abstract Background The roles of circulating endothelial progenitor cell (EPC and mononuclear cell apoptosis (MCA in liver cirrhosis (LC patients are unknown. Moreover, vascular endothelial growth factor (VEGF and stromal cell-derived factor (SDF-1α are powerful endogenous substances enhancing EPC migration into circulation. We assessed the level and function of EPCs [CD31/CD34 (E1, KDR/CD34 (E2, CXCR4/CD34 (E3], levels of MCA, VEGF and SDF-1α in circulation of LC patients. Methods Blood sample was prospectively collected once for assessing EPC level and function, MCA, and plasma levels of VEGF and SDF-1α using flow cytometry and enzyme-linked immunosorbent assay (ELISA, respectively, in 78 LC patients and 25 age- and gender-matched healthy controls. Results Number of EPCs (E1, E2, E3 was lower (all p 2, E3 was higher but MCA was lower (all p  Conclusion The results of this study demonstrated that level, angiogenic capacity, and function of circulating EPCs were significantly reduced, whereas plasma levels of SDF-1α and circulating MCA were substantially enhanced in cirrhotic patients.

  5. Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

    Directory of Open Access Journals (Sweden)

    K.S. Canuto

    2015-01-01

    Full Text Available Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC radiation. Exponential and stationary wild-type and uvrA-deficient E. coli cells were exposed to a low-level red laser and in sequence to UVC radiation. Bacterial survival was evaluated to determine the laser protection factor (ratio between the number of viable cells after exposure to the red laser and UVC and the number of viable cells after exposure to UVC. Bacterial filaments were counted to obtain the percentage of filamentation. Area-perimeter ratios were calculated for evaluation of cellular morphology. Experiments were carried out in duplicate and the results are reported as the means of three independent assays. Pre-exposure to a red laser protected wild-type and uvrA-deficient E. coli cells against the lethal effect of UVC radiation, and increased the percentage of filamentation and the area-perimeter ratio, depending on UVC fluence and physiological conditions in the cells. Therapeutic, low-level red laser radiation can induce DNA lesions at a sub-lethal level. Consequences to cells and tissues should be considered when clinical protocols based on this laser are carried out.

  6. Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

    International Nuclear Information System (INIS)

    Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC) radiation. Exponential and stationary wild-type and uvrA-deficient E. coli cells were exposed to a low-level red laser and in sequence to UVC radiation. Bacterial survival was evaluated to determine the laser protection factor (ratio between the number of viable cells after exposure to the red laser and UVC and the number of viable cells after exposure to UVC). Bacterial filaments were counted to obtain the percentage of filamentation. Area-perimeter ratios were calculated for evaluation of cellular morphology. Experiments were carried out in duplicate and the results are reported as the means of three independent assays. Pre-exposure to a red laser protected wild-type and uvrA-deficient E. coli cells against the lethal effect of UVC radiation, and increased the percentage of filamentation and the area-perimeter ratio, depending on UVC fluence and physiological conditions in the cells. Therapeutic, low-level red laser radiation can induce DNA lesions at a sub-lethal level. Consequences to cells and tissues should be considered when clinical protocols based on this laser are carried out. (author)

  7. Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

    Energy Technology Data Exchange (ETDEWEB)

    Canuto, K.S.; Guimaraes, O.R.; Geller, M. [Centro Universitario Serra dos Orgaos, Teresopolis, RJ (Brazil). Centro de Ciencias da Saude; Sergio, L.P.S. [Instituto de Biologia Roberto Alcantara Gomes, Rio de Janeiro, RJ (Brazil). Departamento de Biofisica e Biometria; Paoli, F. [Universidade Federal de Juiz de Fora (UFJF), Juiz de Fora, MG (Brazil). Departamento de Morfologia; Fonseca, A.S., E-mail: adnfonseca@ig.com.br [Universidade Federal do Estado do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ (Brazil). Departamento de Ciencias Fisiologicas

    2015-10-15

    Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC) radiation. Exponential and stationary wild-type and uvrA-deficient E. coli cells were exposed to a low-level red laser and in sequence to UVC radiation. Bacterial survival was evaluated to determine the laser protection factor (ratio between the number of viable cells after exposure to the red laser and UVC and the number of viable cells after exposure to UVC). Bacterial filaments were counted to obtain the percentage of filamentation. Area-perimeter ratios were calculated for evaluation of cellular morphology. Experiments were carried out in duplicate and the results are reported as the means of three independent assays. Pre-exposure to a red laser protected wild-type and uvrA-deficient E. coli cells against the lethal effect of UVC radiation, and increased the percentage of filamentation and the area-perimeter ratio, depending on UVC fluence and physiological conditions in the cells. Therapeutic, low-level red laser radiation can induce DNA lesions at a sub-lethal level. Consequences to cells and tissues should be considered when clinical protocols based on this laser are carried out. (author)

  8. Helicobacter pylori induces mitochondrial DNA mutation and reactive oxygen species level in AGS cells

    Directory of Open Access Journals (Sweden)

    Xue-Wen Huang, Rui-Hua Luo, Qi Zhao, Zhong-Ze Shen, Li-Li Huang, Xian-Yuan An, Lan-Jing Zhao, Jie Wang, Yu-Zheng Huang

    2011-01-01

    Full Text Available To investigate the role of ROS in the helicobacter pylori (Hp induced mtDNA mutations, AGS cells were treated by extracts of Hp11638 or Hp11638M. The ROS levels, cytochrome C reductions, and intracellular ATP levels were measured. The coding region and the D-Loop region were amplified and sequenced. Results showed the ROS levels, cytochrome C reduction and mtDNA mutations were markedly increased and cell viability decreased after treatment with both Hp extracts, and 616 mutations were detected in D-Loop region and 3 heteroplasmic point mutations in the Cytb gene. No mutations were found in the coding region. The mutation rates of mtDNA D-Loop region were positively correlated with the ROS levels and negatively to the ATP levels.

  9. Altered intracellular pH regulation in cells with high levels of P-glycoprotein expression.

    Science.gov (United States)

    Young, Gregory; Reuss, Luis; Altenberg, Guillermo A

    2011-01-01

    P-glycoprotein is an ATP-binding-cassette transporter that pumps many structurally unrelated drugs out of cells through an ATP-dependent mechanism. As a result, multidrug-resistant cells that overexpress P-glycoprotein have reduced intracellular steady-state levels of a variety of chemotherapeutic agents. In addition, increased cytosolic pH has been a frequent finding in multidrug-resistant cells that express P-glycoprotein, and it has been proposed that this consequence of P-glycoprotein expression may contribute to the lower intracellular levels of chemotherapeutic agents. In these studies, we measured intracellular pH and the rate of acid extrusion in response to an acid load in two cells with very different levels of P-glycoprotein expression: V79 parental cells and LZ-8 multidrug resistant cells. Compared to the wild-type V79 cells, LZ-8 cells have a lower intracellular pH and a slower recovery of intracellular pH after an acid load. The data also show that LZ-8 cells have reduced ability to extrude acid, probably due to a decrease in Na(+)/H(+) exchanger activity. The alterations in intracellular pH and acid extrusion in LZ-8 cells are reversed by 24-h exposure to the multidrug-resistance modulator verapamil. The lower intracellular pH in LZ-8 indicates that intracellular alkalinization is not necessary for multidrug resistance. The reversal by verapamil of the decreased acid-extrusion suggests that P-glycoprotein can affect other membrane transport mechanism. PMID:22003434

  10. Regulation of hTERT by BCR-ABL at multiple levels in K562 cells

    Directory of Open Access Journals (Sweden)

    Chai Juin Hsien

    2011-12-01

    Full Text Available Abstract Background The cytogenetic characteristic of Chronic Myeloid Leukemia (CML is the formation of the Philadelphia chromosome gene product, BCR-ABL. Given that BCR-ABL is the specific target of Gleevec in CML treatment, we investigated the regulation of the catalytic component of telomerase, hTERT, by BCR-ABL at multiple levels in K562 cells. Methods Molecular techniques such as over expression, knockdown, real-time PCR, immunoprecipitation, western blotting, reporter assay, confocal microscopy, telomerase assays and microarray were used to suggest that hTERT expression and activity is modulated by BCR-ABL at multiple levels. Results Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at mRNA level and significantly reduced telomerase activity (TA in K562 cells, but not in HL60 or Jurkat cells (BCR-ABL negative cells. We also demonstrated that the transcription factor STAT5a plays a critical role in hTERT gene regulation in K562 cells. Knockdown of STAT5a, but not STAT5b, resulted in a marked downregulation of hTERT mRNA level, TA and hTERT protein level in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Conclusions Our data reveal that BCR-ABL can regulate TA at multiple levels, including transcription, post-translational level, and proper localization. Thus, suppression of cell growth and induction of apoptosis by Gleevec treatment may be partially due to TA inhibition. Additionally, we have identified STAT5a as critical mediator of the hTERT gene expression in BCR-ABL positive CML cells, suggesting that targeting STAT5a may be a promising therapeutic strategy for BCR-ABL positive

  11. Regulation of hTERT by BCR-ABL at multiple levels in K562 cells

    International Nuclear Information System (INIS)

    The cytogenetic characteristic of Chronic Myeloid Leukemia (CML) is the formation of the Philadelphia chromosome gene product, BCR-ABL. Given that BCR-ABL is the specific target of Gleevec in CML treatment, we investigated the regulation of the catalytic component of telomerase, hTERT, by BCR-ABL at multiple levels in K562 cells. Molecular techniques such as over expression, knockdown, real-time PCR, immunoprecipitation, western blotting, reporter assay, confocal microscopy, telomerase assays and microarray were used to suggest that hTERT expression and activity is modulated by BCR-ABL at multiple levels. Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia) cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at mRNA level and significantly reduced telomerase activity (TA) in K562 cells, but not in HL60 or Jurkat cells (BCR-ABL negative cells). We also demonstrated that the transcription factor STAT5a plays a critical role in hTERT gene regulation in K562 cells. Knockdown of STAT5a, but not STAT5b, resulted in a marked downregulation of hTERT mRNA level, TA and hTERT protein level in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Our data reveal that BCR-ABL can regulate TA at multiple levels, including transcription, post-translational level, and proper localization. Thus, suppression of cell growth and induction of apoptosis by Gleevec treatment may be partially due to TA inhibition. Additionally, we have identified STAT5a as critical mediator of the hTERT gene expression in BCR-ABL positive CML cells, suggesting that targeting STAT5a may be a promising therapeutic strategy for BCR-ABL positive CML patients

  12. Connexin50D47A Decreases Levels of Fiber Cell Connexins and Impairs Lens Fiber Cell Differentiation

    OpenAIRE

    Berthoud, Viviana M.; Minogue, Peter J.; Yu, Helena; Schroeder, Richard; Snabb, Joseph I.; Beyer, Eric C.

    2013-01-01

    Both heterozygous and homozygous Cx50D47A-expressing mice develop cataracts due to a severe decrease in the abundance of lens fiber connexins. The mutant mouse lenses also show impaired degradation of organelles and decreased levels of some crystallins implicating Cx50 in fiber cell differentiation.

  13. SINGLE-CELL LEVEL INVESTIGATION OF CYTOSKELETAL/CELLULAR RESPONSE TO EXTERNAL STIMULI

    Energy Technology Data Exchange (ETDEWEB)

    Hiddessen, A L

    2007-02-26

    A detailed understanding of the molecular mechanisms by which chemical signals control cell behavior is needed if the complex biological processes of embryogenesis, development, health and disease are to be completely understood. Yet, if we are to fully understand the molecular mechanisms controlling cell behavior, measurements at the single cell level are needed to supplement information gained from population level studies. One of the major challenges to accomplishing studies at the single cell level has been a lack of physical tools to complement the powerful molecular biological assays which have provided much of what we currently know about cell behavior. The goal of this exploratory project is the development of an experimental platform that facilitates integrated observation, tracking and analysis of the responses of many individual cells to controlled environmental factors (e.g. extracellular signals). Toward this goal, we developed chemically-patterned microarrays of both adherent and suspension mammalian cell types. A novel chemical patterning methodology, based on photocatalytic lithography, was developed to construct biomolecule and cell arrays that facilitate analysis of biological function. Our patterning techniques rely on inexpensive stamp materials and visible light, and do not necessitate mass transport or specified substrates. Patterned silicon and glass substrates are modified such that there is a non-biofouling polymer matrix surrounding the adhesive regions that target biomolecules and cells. Fluorescence and reflectance microscopy reveal successful patterning of proteins and single to small clusters of mammalian cells. In vitro assays conducted upon cells on the patterned arrays demonstrate the viability of cells interfacing with this synthetic system. Hence, we have successfully established a versatile cell measurement platform which can be used to characterize the molecular regulators of cellular behavior in a variety of important

  14. Effects of Phototherapy on Cytokines’ Levels and White Blood Cells in Term Neonate with Hyperbilirubinemia

    OpenAIRE

    Jahanshahifard, Sedigheh; Ahmadpour-Kacho, Mousa; Pasha, Yadollah Zahed

    2012-01-01

    Objective: Phototherapy is the most common treatment used for severe jaundice. There is increasing evidence that phototherapy can directly affect the expression and function of cell surface receptors including adhesion molecules, cytokines, and growth factor receptors. The aim of this study is to investigate the effect of phototherapy use on the levels of interleukin (IL)-1α, IL-6, and tumor necrosis factor (TNF)-α as cytokine expressions from keratinocytes, and also white blood cell counts i...

  15. Levels of BDNF Impact Oligodendrocyte Lineage Cells Following a Cuprizone Lesion

    OpenAIRE

    VonDran, Melissa W.; Singh, Harmandeep; Honeywell, Jean Z.; Dreyfus, Cheryl F

    2011-01-01

    Previous work in culture has shown that basal forebrain (BF) oligodendrocyte (OLG) lineage cells respond to BDNF by increasing DNA synthesis and differentiation. Further, in the BF in vivo, reduced levels of BDNF as seen in BDNF +/− mice result in reduced numbers of NG2+ cells and deficits in myelin proteins throughout development and in the adult, suggesting that BDNF impacts the proliferating population of OLGs as well as differentiation in vivo. In this study, to investigate roles BDNF may...

  16. Immune cell distribution and immunoglobulin levels change following sciatic nerve injury in a rat model

    Directory of Open Access Journals (Sweden)

    Wei Yuan

    2016-07-01

    Full Text Available Objective(s: To investigate the systemic and local immune status of two surgical rat models of sciatic nerve injury, a crushed sciatic nerve, and a sciatic nerve transection Materials and Methods:Twenty-four adult male Sprague-Dawley rats were randomly divided into three groups: sham-operation (control group, sciatic nerve crush, and sciatic nerve transaction. Sciatic nerve surgery was performed. The percentage of CD4+ cells and the CD4+/CD8+ratio were determined by flow cytometry. Serum IgM and IgG levels were analyzed by ELISA. T-cells (CD3 and macrophages (CD68 in sciatic nerve tissue sections were identified through immunohistochemistry. Results: Compared to sham-operated controls, in rats that underwent nerve injury, the percentage of CD4+ cells and the CD4+/CD8+ ratio in the peripheral blood were significantly  decreased 7 days after surgery, serum IgM levels were increased 14 days after surgery, and serum IgG levels were increased 21 days after surgery. There were a large number of CD3+ cells and a small number of CD68+ cells in sciatic nerve tissue sections 21 days after surgery, indicating T-cell and macrophage activation and infiltration. Local IgG deposition was also detected at the nerve injury site 21 days after surgery. Conclusion: Rat humoral and cellular immune status changed following sciatic nerve injury, particularly with regard to the cellular immune response at the nerve injury site.

  17. Immune cell distribution and immunoglobulin levels change following sciatic nerve injury in a rat model

    Science.gov (United States)

    Yuan, Wei; Feng, Xinhong

    2016-01-01

    Objective(s): To investigate the systemic and local immune status of two surgical rat models of sciatic nerve injury, a crushed sciatic nerve, and a sciatic nerve transection Materials and Methods: Twenty-four adult male Sprague-Dawley rats were randomly divided into three groups: sham-operation (control group), sciatic nerve crush, and sciatic nerve transaction. Sciatic nerve surgery was performed. The percentage of CD4+ cells and the CD4+/CD8+ratio were determined by flow cytometry. Serum IgM and IgG levels were analyzed by ELISA. T-cells (CD3) and macrophages (CD68) in sciatic nerve tissue sections were identified through immunohistochemistry. Results: Compared to sham-operated controls, in rats that underwent nerve injury, the percentage of CD4+ cells and the CD4+/CD8+ ratio in the peripheral blood were significantly decreased 7 days after surgery, serum IgM levels were increased 14 days after surgery, and serum IgG levels were increased 21 days after surgery. There were a large number of CD3+ cells and a small number of CD68+ cells in sciatic nerve tissue sections 21 days after surgery, indicating T-cell and macrophage activation and infiltration. Local IgG deposition was also detected at the nerve injury site 21 days after surgery. Conclusion: Rat humoral and cellular immune status changed following sciatic nerve injury, particularly with regard to the cellular immune response at the nerve injury site.

  18. Detection of the Level of Reactive Oxygen Species Induced by Ionizing Radiation in Cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jin Kyu; Chung, Dong Min; Kim, Jin-Hong [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2015-05-15

    By definition, the direct effect is referred to interaction between photon and DNA molecule, whereas the indirect effect is mediated by the reactive oxygen species (ROS) generated by radiolysis and subsequent reaction. It has been reported that ROS produced after exposure to IR can react with cellular materials such as DNA, proteins, carbohydrates and lipids. ROS is free radicals such as the superoxide anion, hydroxyl radicals and the non-radical hydrogen peroxide. Cells generate ROS during aerobic metabolism. Excessive production of ROS can lead to oxidative stress, genetic alteration and even cell death. It has been reported that ROS plays a critical role in radiation-induced cell injury. Thus, it is of great interest to determine the radiation-induced ROS level. Many kinds of methods to detect the level of ROS have been developed so far. There were random changes of fluorescence intensity in the treatment after irradiation. This result meant that this protocol was not appropriate for determination of radiation-induced ROS. On the other hand, the fluorescence intensity was increased in a dose-dependent manner when the cells were treated with the DCFH-DA solution before irradiation. Conclusions can be drawn from the experimental results of this study. In order to properly measure the ROS level in the cells exposed to ionizing radiation, the cells should be treated with the DCFH-DA solution before irradiation.

  19. Levels of expression of CD19 and CD20 in chronic B cell leukaemias.

    Science.gov (United States)

    Ginaldi, L; De Martinis, M; Matutes, E; Farahat, N; Morilla, R; Catovsky, D

    1998-01-01

    AIMS: To investigate whether the antigen levels of the B cell lineage markers CD19 and CD20 can distinguish between normal and neoplastic B cells or characterise distinct expression patterns among the chronic B cell leukaemias. METHODS: Peripheral blood cells from 70 patients with B cell disorders and 17 healthy donors were analysed by quantitative flow cytometry. Direct immunofluorescence staining was performed with phycoerythrin conjugated CD19 and CD20 monoclonal antibodies. Standard microbeads with different capacities to bind mouse immunoglobulins were used to convert the mean fluorescence intensity (MFI) values into number of antigen molecules/cell, expressed as antibody binding capacity (ABC). RESULTS: CD19 and CD20 ABC values in leukaemic B cells differed from those of normal blood B lymphocytes. The results identified distinct profiles of CD19 and CD20 expression in the various types of B cell leukaemias. In all leukaemias studied except hairy cell leukaemia (HCL), CD19 expression was significantly lower than the mean (SD) value in normal B cells (22 (7) x 10(3) molecules/cell), as follows: chronic lymphocytic leukaemia (CLL), 13 (7) x 10(3); B prolymphocytic leukaemia (B-PLL), 16 (9) x 10(3); splenic lymphoma with villous lymphocytes (SLVL), 15 (11) x 10(3); mantle cell lymphoma (MCL), 10 (7) x 10(3). In HCL there was strong CD19 expression (38 (16) x 10(3)). In contrast, the level of expression of membrane CD20 was higher than the mean (SD) value in normal B cells (94 (16) x 10(3) molecules/cell) in MCL (123 (51) x 10(3)); B-PLL (129 (47) x 10(3)); SLVL (167 (72) x 10(3)); and HCL (312 (110) x 10(3)); while it was significantly lower (65 (11) x 10(3)) in CLL compared with normal B cells and the other B cell leukaemias. CONCLUSIONS: Quantitative determination of CD19 and CD20 may provide useful diagnostic information for the study of B lymphoproliferative disorders. PMID:9708202

  20. Oral cavity squamous cell carcinoma metastatic to central compartment (level 6) lymph nodes.

    Science.gov (United States)

    Likhterov, Ilya; Rowe, Meghan E; Khorsandi, Azita S; Urken, Mark L

    2016-08-01

    Alterations to drainage pathways in the head and neck as a result of surgical manipulation are not well understood. We present two unusual cases of oral squamous cell carcinoma metastatic to the level 6 nodal compartment following extensive treatment. Both oral squamous cell carcinoma cases exhibited metastases to the central neck compartment following extensive surgery and radiation. Each patient had prior history of multifocal oral cavity disease and recurrent neck metastases requiring salvage lymphadenectomy. Surgical interventions may alter the usual lymphatic drainage patterns. In cases of extensive treatment, all levels of the neck should be monitored for lymph node recurrence. Laryngoscope, 126:1803-1805, 2016. PMID:26490846

  1. Effects of combined radiofrequency radiation exposure on levels of reactive oxygen species in neuronal cells

    International Nuclear Information System (INIS)

    The objective of this study was to investigate the effects of the combined RF radiation (837 MHz CDMA plus 1950 MHz WCDMA) signal on levels of intracellular reactive oxygen species (ROS) in neuronal cells. Exposure of the combined RF signal was conducted at specific absorption rate values of 2 W/kg of CDMA plus 2 W/kg of WCDMA for 2 h. Co-exposure to combined RF radiation with either H2O2 or menadione was also performed. The experimental exposure groups were incubator control, sham-exposed, combined RF radiation-exposed with or without either H2O2 or menadione groups. The intracellular ROS level was measured by flow cytometry using the fluorescent probe dichlorofluorescein diacetate. Intracellular ROS levels were not consistently affected by combined RF radiation exposure alone in a time-dependent manner in U87, PC12 or SH-SY5Y cells. In neuronal cells exposed to combined RF radiation with either H2O2 or menadione, intracellular ROS levels showed no statically significant alteration compared with exposure to menadione or H2O2 alone. These findings indicate that neither combined RF radiation alone nor combined RF radiation with menadione or H2O2 influences the intracellular ROS level in neuronal cells such as U87, PC12 or SH-SY5Y. (author)

  2. TRPV Channels in Mast Cells as a Target for Low-Level-Laser Therapy

    Directory of Open Access Journals (Sweden)

    Lina Wang

    2014-06-01

    Full Text Available Low-level laser irradiation in the visible as well as infrared range is applied to skin for treatment of various diseases. Here we summarize and discuss effects of laser irradiation on mast cells that leads to degranulation of the cells. This process may contribute to initial steps in the final medical effects. We suggest that activation of TRPV channels in the mast cells forms a basis for the underlying mechanisms and that released ATP and histamine may be putative mediators for therapeutic effects.

  3. Elevated levels of cell-free circulating DNA in patients with acute dengue virus infection.

    Directory of Open Access Journals (Sweden)

    Tran Thi Ngoc Ha

    Full Text Available BACKGROUND: Apoptosis is thought to play a role in the pathogenesis of severe dengue and the release of cell-free DNA into the circulatory system in several medical conditions. Therefore, we investigated circulating DNA as a potential biomarker for severe dengue. METHODS AND FINDINGS: A direct fluorometric degradation assay using PicoGreen was performed to quantify cell-free DNA from patient plasma. Circulating DNA levels were significantly higher in patients with dengue virus infection than with other febrile illnesses and healthy controls. Remarkably, the increase of DNA levels correlated with the severity of dengue. Additionally, multivariate logistic regression analysis showed that circulating DNA levels independently correlated with dengue shock syndrome. CONCLUSIONS: Circulating DNA levels were increased in dengue patients and correlated with dengue severity. Additional studies are required to show the benefits of this biomarker in early dengue diagnosis and for the prognosis of shock complication.

  4. Effect of low-level laser therapy on mesenchymal stem cell proliferation: a systematic review.

    Science.gov (United States)

    Ginani, Fernanda; Soares, Diego Moura; Barreto, Mardem Portela E Vasconcelos; Barboza, Carlos Augusto Galvão

    2015-11-01

    Low-level laser therapy (LLLT) has been used in several in vitro experiments in order to stimulate cell proliferation. Cells such as fibroblasts, keratinocytes, lymphocytes, and osteoblasts have shown increased proliferation when submitted to laser irradiation, although little is known about the effects of LLLT on stem cells. This study aims to assess, through a systematic literature review, the effects of LLLT on the in vitro proliferation of mesenchymal stem cells. Using six different terms, we conducted an electronic search in PubMed/Medline database for articles published in the last twelve years. From 463 references obtained, only 19 papers met the search criteria and were included in this review. The analysis of the papers showed a concentration of experiments using LLLT on stem cells derived from bone marrow, dental pulp, periodontal ligament, and adipose tissue. Several protocols were used to irradiate the cells, with variations on wavelength, power density, radiation time, and state of light polarization. Most studies demonstrated an increase in the proliferation rate of the irradiated cells. It can be concluded that the laser therapy positively influences the in vitro proliferation of stem cells studied, being necessary to carry out further experiments on other cell types and to uniform the methodological designs. PMID:25764448

  5. T cell Bim levels reflect responses to anti–PD-1 cancer therapy

    Science.gov (United States)

    Dronca, Roxana S.; Liu, Xin; Harrington, Susan M.; Chen, Lingling; Cao, Siyu; Kottschade, Lisa A.; McWilliams, Robert R.; Block, Matthew S.; Nevala, Wendy K.; Thompson, Michael A.; Mansfield, Aaron S.; Park, Sean S.; Markovic, Svetomir N.; Dong, Haidong

    2016-01-01

    Immune checkpoint therapy with PD-1 blockade has emerged as an effective therapy for many advanced cancers; however, only a small fraction of patients achieve durable responses. To date, there is no validated blood-based means of predicting the response to PD-1 blockade. We report that Bim is a downstream signaling molecule of the PD-1 pathway, and its detection in T cells is significantly associated with expression of PD-1 and effector T cell markers. High levels of Bim in circulating tumor-reactive (PD-1+CD11ahiCD8+) T cells were prognostic of poor survival in patients with metastatic melanoma who did not receive anti–PD-1 therapy and were also predictive of clinical benefit in patients with metastatic melanoma who were treated with anti–PD-1 therapy. Moreover, this circulating tumor-reactive T cell population significantly decreased after successful anti–PD-1 therapy. Our study supports a crucial role of Bim in both T cell activation and apoptosis as regulated by PD-1 and PD-L1 interactions in effector CD8+ T cells. Measurement of Bim levels in circulating T cells of patients with cancer may provide a less invasive strategy to predict and monitor responses to anti–PD-1 therapy, although future prospective analyses are needed to validate its utility. PMID:27182556

  6. Perovskite Solar Cells Employing Dopant-Free Organic Hole Transport Materials with Tunable Energy Levels.

    Science.gov (United States)

    Liu, Yongsheng; Hong, Ziruo; Chen, Qi; Chen, Huajun; Chang, Wei-Hsuan; Yang, Yang Michael; Song, Tze-Bin; Yang, Yang

    2016-01-20

    Conjugated small-molecule hole-transport materials (HTMs) with tunable energy levels are designed and synthesized for efficient perovskite solar cells. A champion device with efficiency of 16.2% is demonstrated using a dopant-free DERDTS-TBDT HTM, while the DORDTS-DFBT-HTM-based device shows an inferior performance of 6.2% due to its low hole mobility and unmatched HOMO level with the valence band of perovskite film. PMID:26588665

  7. 324 Building radiochemical engineering cells, high-level vault, low-level vault, and associated areas closure plan

    International Nuclear Information System (INIS)

    The Hanford Site, located adjacent to and north of Richland, Washington, is operated by the US Department of Energy, Richland Operations Office (RL). The 324 Building is located in the 300 Area of the Hanford Site. The 324 Building was constructed in the 1960s to support materials and chemical process research and development activities ranging from laboratory/bench-scale studies to full engineering-scale pilot plant demonstrations. In the mid-1990s, it was determined that dangerous waste and waste residues were being stored for greater than 90 days in the 324 Building Radiochemical Engineering Cells (REC) and in the High-Level Vault/Low-Level Vault (HLV/LLV) tanks. [These areas are not Resource Conservation and Recovery Act of 1976 (RCRA) permitted portions of the 324 Building.] Through the Hanford Federal Facility Agreement and Consent Order (Tri-Party Agreement) Milestone M-89, agreement was reached to close the nonpermitted RCRA unit in the 324 Building. This closure plan, managed under TPA Milestone M-20-55, addresses the identified building areas targeted by the Tri-Party Agreement and provides commitments to achieve the highest degree of compliance practicable, given the special technical difficulties of managing mixed waste that contains high-activity radioactive materials, and the physical limitations of working remotely in the areas within the subject closure unit. This closure plan is divided into nine chapters. Chapter 1.0 provides the introduction, historical perspective, 324 Building history and current mission, and the regulatory basis and strategy for managing the closure unit. Chapters 2.0, 3.0, 4.0, and 5.0 discuss the detailed facility description, process information, waste characteristics, and groundwater monitoring respectively. Chapter 6.0 deals with the closure strategy and performance standard, including the closure activities for the B-Cell, D-Cell, HLV, LLV; piping and miscellaneous associated building areas. Chapter 7.0 addresses the

  8. 324 Building radiochemical engineering cells, high-level vault, low-level vault, and associated areas closure plan

    Energy Technology Data Exchange (ETDEWEB)

    Barnett, J.M.

    1998-03-25

    The Hanford Site, located adjacent to and north of Richland, Washington, is operated by the US Department of Energy, Richland Operations Office (RL). The 324 Building is located in the 300 Area of the Hanford Site. The 324 Building was constructed in the 1960s to support materials and chemical process research and development activities ranging from laboratory/bench-scale studies to full engineering-scale pilot plant demonstrations. In the mid-1990s, it was determined that dangerous waste and waste residues were being stored for greater than 90 days in the 324 Building Radiochemical Engineering Cells (REC) and in the High-Level Vault/Low-Level Vault (HLV/LLV) tanks. [These areas are not Resource Conservation and Recovery Act of 1976 (RCRA) permitted portions of the 324 Building.] Through the Hanford Federal Facility Agreement and Consent Order (Tri-Party Agreement) Milestone M-89, agreement was reached to close the nonpermitted RCRA unit in the 324 Building. This closure plan, managed under TPA Milestone M-20-55, addresses the identified building areas targeted by the Tri-Party Agreement and provides commitments to achieve the highest degree of compliance practicable, given the special technical difficulties of managing mixed waste that contains high-activity radioactive materials, and the physical limitations of working remotely in the areas within the subject closure unit. This closure plan is divided into nine chapters. Chapter 1.0 provides the introduction, historical perspective, 324 Building history and current mission, and the regulatory basis and strategy for managing the closure unit. Chapters 2.0, 3.0, 4.0, and 5.0 discuss the detailed facility description, process information, waste characteristics, and groundwater monitoring respectively. Chapter 6.0 deals with the closure strategy and performance standard, including the closure activities for the B-Cell, D-Cell, HLV, LLV; piping and miscellaneous associated building areas. Chapter 7.0 addresses the

  9. Mouse T-cells restrict replication of human immunodeficiency virus at the level of integration

    Directory of Open Access Journals (Sweden)

    Goffinet Christine

    2008-07-01

    Full Text Available Abstract Background The development of an immunocompetent, genetically modified mouse model to study HIV-1 pathogenesis and to test antiviral strategies has been hampered by the fact that cells from native mice do not or only inefficiently support several steps of the HIV-1 replication cycle. Upon HIV-1 infection, mouse T-cell lines fail to express viral proteins, but the underlying replication barrier has thus far not been unambiguously identified. Here, we performed a kinetic and quantitative assessment of consecutive steps in the early phase of the HIV-1 replication cycle in T-cells from mice and humans. Results Both T-cell lines and primary T-cells from mice harbor a severe post-entry defect that is independent of potential species-specTR transactivation. Reverse transcription occurred efficiently following VSV-G-mediated entry of virions into mouse T-cells, and abundant levels of 2-LTR circles indicated successful nuclear import of the pre-integration complex. To probe the next step in the retroviral replication cycle, i.e. the integration of HIV-1 into the host cell genome, we established and validated a nested real-time PCR to specifically quantify HIV-1 integrants exploiting highly repetitive mouse B1 elements. Importantly, we demonstrate that the frequency of integrant formation is diminished 18- to > 305-fold in mouse T-cell lines compared to a human counterpart, resulting in a largely abortive infection. Moreover, differences in transgene expression from residual vector integrants, the transcription off which is cyclin T1-independent, provided evidence for an additional, peri-integrational deficit in certain mouse T-cell lines. Conclusion In contrast to earlier reports, we find that mouse T-cells efficiently support early replication steps up to and including nuclear import, but restrict HIV-1 at the level of chromosomal integration.

  10. Salt tolerance at single cell level in giant-celled Characeae

    Directory of Open Access Journals (Sweden)

    Mary Jane eBeilby

    2015-04-01

    Full Text Available Characean plants provide an excellent experimental system for electrophysiology and physiology due to: (i very large cell size, (ii position on phylogenetic tree near the origin of land plants and (iii continuous spectrum from very salt sensitive to very salt tolerant species. A range of experimental techniques is described, some unique to characean plants. Application of these methods provided electrical characteristics of membrane transporters, which dominate the membrane conductance under different outside conditions. With this considerable background knowledge the electrophysiology of salt sensitive and salt tolerant genera can be compared under salt and/or osmotic stress. Both salt tolerant and salt sensitive Characeae show a rise in membrane conductance and simultaneous increase in Na+ influx upon exposure to saline medium. Salt tolerant Chara longifolia and Lamprothamnium sp. exhibit proton pump stimulation upon both turgor decrease and salinity increase, allowing the membrane PD to remain negative. The turgor is regulated through the inward K+ rectifier and 2H+/Cl- symporter. Lamprothamnium plants can survive in hypersaline media up to twice seawater strength and withstand large sudden changes in salinity. Salt-sensitive Chara australis succumbs to 50 - 100 mM NaCl in few days. Cells exhibit no pump stimulation upon turgor decrease and at best transient pump stimulation upon salinity increase. Turgor is not regulated. The membrane PD exhibits characteristic noise upon exposure to salinity. Depolarization of membrane PD to excitation threshold sets off trains of action potentials, leading to further loses of K+ and Cl-. In final stages of salt damage the H+/OH- channels are thought to become the dominant transporter, dissipating the proton gradient and bringing the cell PD close to 0. The differences in transporter electrophysiology and their synergy under osmotic and/or saline stress in salt sensitive and salt tolerant characean cells

  11. Elevated levels of 14-3-3 proteins, serotonin, gamma enolase and pyruvate kinase identified in clinical samples from patients diagnosed with colorectal cancer

    Czech Academy of Sciences Publication Activity Database

    Dowling, P.; Hughes, D. J.; Larkin, A.M.; Meiller, J.; Henry, M.; Meleady, P.; Lynch, V.; Pardini, B.; Naccarati, A.; Levý, M.; Vodička, Pavel; Neary, P.; Clynes, M.

    2015-01-01

    Roč. 441, feb. (2015), s. 133-141. ISSN 0009-8981 Institutional support: RVO:68378041 Keywords : biomarkers * colorectal cancer * proteomics * mass spectrometry * 14-3-3 proteins * pyruvate kinase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.824, year: 2014

  12. Hematopoietic cell transplantation in murine globoid cell leukodystrophy (the twitcher mouse): effects on levels of galactosylceramidase, psychosine, and galactocerebrosides

    International Nuclear Information System (INIS)

    Hematopoietic cell transplantation (HCT) prolongs survival in the twitcher mouse, an authentic animal model of human globoid cell leukodystrophy (Krabbe disease: galactosylceramidase deficiency), but the effects of HCT on levels of galactosylceramidase, psychosine, and cerebrosides in the tissues of twitcher mice have not been previously studied. Galactosylceramidase was less than 8% of control activity in tissues of untreated twitcher mice but reached normal values in brain and spleen and 20-30% of control kidney of 100-day-old twitchers that received HCT at age 10 days. Using a recently developed method for the simultaneous determination of psychosine and cerebrosides, the authors measured the tissue levels of these lipids in the above animals. The levels of psychosine in brain, sciatic nerve, and kidney of untreated twitcher mice were 44, 200, and 12 times control values, respectively, in 30-day-old animals and 69,500, and 14 times control levels in 40-day-old mice. On the other hand, levels of cerebroside were approximately 35% of control values in sciatic nerve, remained about the same in the brain, and were elevated 10-fold in the kidney of twitcher mice. After HCT, psychosine levels in the brains of 30-day-old twitchers were lowered to 30-35% of values in untreated twitchers, and the levels remained in that range during the post-HCT period. Similarly, brain cerebroside levels remained low in HCT-treated twitcher mice. It is not known whether the extremely high levels of psychosine in sciatic nerves ultimately contribute to the death of twitcher mice after HCT

  13. Quantitative investigation of bacterial chemotaxis at the single-cell level

    Science.gov (United States)

    Min, Taejin

    Living cells sense and respond to constantly changing environmental conditions. Depending on the type of stimuli, the cell may response by altering gene expression pattern, secreting molecules, or migrating to a different environment. Directed movement of cells in response to chemical stimuli is called chemotaxis. In bacterial chemotaxis, small extracellular molecules bind receptor proteins embedded in the cell membrane, which then transmit the signal inside the cell through a cascade of protein-protein interactions. This chain of events influences the behavior of motor proteins that drive the rotation of helical filaments called flagella. Individual cells of the gut-dwelling bacteria Escherichia coli (E. coli) have many such flagella, whose collective action results in the swimming behavior of the cell. A recent study found that in absence of chemical stimuli, fluctuations in the protein cascade can cause non-Poissonian switching behavior in the flagellar motor (2). A corollary was that extension of such behavior to the whole-cell swimming level would have implications for E. coli's foraging strategy. However, existence of such behavior at the swimming cell level could not be predicted a priori, since the mapping from single flagellum behavior to the swimming behavior of a multi-flagellated cell is complex and poorly understood (3, 4). Here we characterize the chemotactic behavior of swimming E. coli cells using a novel optical trap-based measurement technique. This technique allows us to trap individual cells and monitor their swimming behavior over long time periods with high temporal resolution. We find that swimming cells exhibit non-Poissonian switching statistics between different swimming states, in a manner similar to the rotational direction-switching behavior seen in individual flagella. Furthermore, we develop a data analysis routine that allows us to characterize higher order swimming features such as reversal of swimming direction and existence of

  14. Hazard Identification on a Single Cell Level Using a Laser Beam

    Directory of Open Access Journals (Sweden)

    Xing-Zheng Wu

    2007-01-01

    Full Text Available This research shows a novel method for hazard identification of a chemical and UV light on a single cell level with a laser probe beam. The laser probe beam was passed through interface of cell membrane/culture medium of a cultured human hepatoblastoma cell line HepG2. Deflection of the laser probe beam, which was induced by changes in concentration gradients due to the active materials movement across the cell membrane, was monitored. When a toxic hazard existed, a living cell was expected to be killed or injured, or cellular behaviors to be changed greatly. Then, the changing deflection signal from the living cell would become unchanged or altered in a different way. This was successfully demonstrated with cytotoxity of UV light and H2O2. Most of the cultured HepG2 cells showed changing defl ection signals after 10 min illumination of UV-visible light longer than 370 nm, while almost all HepG2 cells showed unchanged deflection signal after 10 min illumination of UV-visible light with wavelength longer than 330 nm. The results suggested that UV light between 330–370 nm could kill the cells. Additions of H2O2 solution with different concentrations to the cell cultures caused the changing deflection signal from a living cell either unchanged or changed in different trend, suggesting toxicity of H2O2 to the cells. The results from the beam deflection detection agreed well with those obtained by the conventional trypane blue method.

  15. Multi-level communication of human retinal pigment epithelial cells via tunneling nanotubes.

    Directory of Open Access Journals (Sweden)

    Dierk Wittig

    Full Text Available BACKGROUND: Tunneling nanotubes (TNTs may offer a very specific and effective way of intercellular communication. Here we investigated TNTs in the human retinal pigment epithelial (RPE cell line ARPE-19. Morphology of TNTs was examined by immunostaining and scanning electron microscopy. To determine the function of TNTs between cells, we studied the TNT-dependent intercellular communication at different levels including electrical and calcium signalling, small molecular diffusion as well as mitochondrial re-localization. Further, intercellular organelles transfer was assayed by FACS analysis. METHODOLOGY AND PRINCIPAL FINDINGS: Microscopy showed that cultured ARPE-19 cells are frequently connected by TNTs, which are not attached to the substratum. The TNTs were straight connections between cells, had a typical diameter of 50 to 300 nm and a length of up to 120 µm. We observed de novo formation of TNTs by diverging from migrating cells after a short time of interaction. Scanning electron microscopy confirmed characteristic features of TNTs. Fluorescence microscopy revealed that TNTs between ARPE-19 cells contain F-actin but no microtubules. Depolymerisation of F-actin, induced by addition of latrunculin-B, led to disappearance of TNTs. Importantly, these TNTs could function as channels for the diffusion of small molecules such as Lucifer Yellow, but not for large molecules like Dextran Red. Further, organelle exchange between cells via TNTs was observed by microscopy. Using Ca²⁺ imaging we show the intercellular transmission of calcium signals through TNTs. Mechanical stimulation led to membrane depolarisation, which expand through TNT connections between ARPE-19 cells. We further demonstrate that TNTs can mediate electrical coupling between distant cells. Immunolabelling for Cx43 showed that this gap junction protein is interposed at one end of 44% of TNTs between ARPE-19 cells. CONCLUSIONS AND SIGNIFICANCE: Our observations indicate that

  16. Analysis of 4070A envelope levels in retroviral preparations and effect on target cell transduction efficiency.

    Science.gov (United States)

    Slingsby, J H; Baban, D; Sutton, J; Esapa, M; Price, T; Kingsman, S M; Kingsman, A J; Slade, A

    2000-07-01

    A number of stable producer cell lines for high-titer Mo-MuLV vectors have been constructed. Development has previously centered on increasing end-point titers by producing maximal levels of Mo-MuLV Gag/Pol, envelope glycoproteins, and retroviral RNA genomes. We describe the production yields and transduction efficiency characteristics of two Mo-MuLV packaging cell lines, FLYA13 and TEFLYA. Although they both produce 4070A-pseudotyped retroviral vectors reproducibly at >1 x 10(6) LFU ml(-1), the transduction efficiency of unconcentrated and concentrated virus from FLYA13 lines is poor compared with vector preparations from TEFLYA lines. A powerful inhibitor of retroviral transduction is secreted by FLYA13 packaging cells. We show that the inhibitory factor does not affect transduction of target cells by RD114-pseudotyped vectors. This suggests that the inhibitory factor functions at the level of envelope-receptor interactions. Phosphate starvation of target cells shows a two-fold increase in Pit2 receptor mRNA and causes some improvement in FLYA13 virus transduction efficiency. Western blots show that FLYA13 viral samples contain an eight-fold higher ratio of 4070A envelope to p30gag than that of virus produced by TEFLYA producer cell lines. This study correlates overexpression of 4070A envelope glycoprotein in retroviral preparations with a reduction of transduction efficiency at high multiplicities of infection. We suggest that TEFLYA packaging cells express preferable levels of 4070A compared with FLYA13, which not only enables high-titer stocks to be generated, but also facilitates a high efficiency of transduction of target cells. PMID:10910141

  17. Human and great ape red blood cells differ in plasmalogen levels and composition

    Directory of Open Access Journals (Sweden)

    Ely John J

    2011-06-01

    Full Text Available Abstract Background Plasmalogens are ether phospholipids required for normal mammalian developmental, physiological, and cognitive functions. They have been proposed to act as membrane antioxidants and reservoirs of polyunsaturated fatty acids as well as influence intracellular signaling and membrane dynamics. Plasmalogens are particularly enriched in cells and tissues of the human nervous, immune, and cardiovascular systems. Humans with severely reduced plasmalogen levels have reduced life spans, abnormal neurological development, skeletal dysplasia, impaired respiration, and cataracts. Plasmalogen deficiency is also found in the brain tissue of individuals with Alzheimer disease. Results In a human and great ape cohort, we measured the red blood cell (RBC levels of the most abundant types of plasmalogens. Total RBC plasmalogen levels were lower in humans than bonobos, chimpanzees, and gorillas, but higher than orangutans. There were especially pronounced cross-species differences in the levels of plasmalogens with a C16:0 moiety at the sn-1 position. Humans on Western or vegan diets had comparable total RBC plasmalogen levels, but the latter group showed moderately higher levels of plasmalogens with a C18:1 moiety at the sn-1 position. We did not find robust sex-specific differences in human or chimpanzee RBC plasmalogen levels or composition. Furthermore, human and great ape skin fibroblasts showed only modest differences in peroxisomal plasmalogen biosynthetic activity. Human and chimpanzee microarray data indicated that genes involved in plasmalogen biosynthesis show cross-species differential expression in multiple tissues. Conclusion We propose that the observed differences in human and great ape RBC plasmalogens are primarily caused by their rates of biosynthesis and/or turnover. Gene expression data raise the possibility that other human and great ape cells and tissues differ in plasmalogen levels. Based on the phenotypes of humans and

  18. The TGFβ pathway stimulates ovarian cancer cell proliferation by increasing IGF1R levels.

    Science.gov (United States)

    Alsina-Sanchis, Elisenda; Figueras, Agnès; Lahiguera, Álvaro; Vidal, August; Casanovas, Oriol; Graupera, Mariona; Villanueva, Alberto; Viñals, Francesc

    2016-10-15

    In a search for new therapeutic targets for treating epithelial ovarian cancer, we analyzed the Transforming Growth Factor Beta (TGFβ) signaling pathway in these tumors. Using a TMA with patient samples we found high Smad2 phosphorylation in ovarian cancer tumoral cells, independently of tumor subtype (high-grade serous or endometrioid). To evaluate the impact of TGFβ receptor inhibition on tumoral growth, we used different models of human ovarian cancer orthotopically grown in nude mice (OVAs). Treatment with a TGFβRI&II dual inhibitor, LY2109761, caused a significant reduction in tumor size in all these models, affecting cell proliferation rate. We identified Insulin Growth Factor (IGF)1 receptor as the signal positively regulated by TGFβ implicated in ovarian tumor cell proliferation. Inhibition of IGF1R activity by treatment with a blocker antibody (IMC-A12) or with a tyrosine kinase inhibitor (linsitinib) inhibited ovarian tumoral growth in vivo. When IGF1R levels were decreased by shRNA treatment, LY2109761 lost its capacity to block tumoral ovarian cell proliferation. At the molecular level TGFβ induced mRNA IGF1R levels. Overall, our results suggest an important role for the TGFβ signaling pathway in ovarian tumor cell growth through the control of IGF1R signaling pathway. Moreover, it identifies anti-TGFβ inhibitors as being of potential use in new therapies for ovarian cancer patients as an alternative to IGF1R inhibition. PMID:27299695

  19. Manganese superoxide dismutase level in blood cells of patients with breast cancer

    International Nuclear Information System (INIS)

    The intracellular MnSOD levels were determined in peripheral blood cells obtained from two age groups of patients with breast cancer (BC): 30-45 year old patients (premenopausal, n=7, clinical stage 1 to 3) and 46-60 year old patients (peri- and postmenopausal women, n=12, clinical stage 3 or 4), at diagnosis, prior to any clinical treatment. The respective healthy women groups were used as controls. Blood cells were also irradiated in vitro with 2- and 9- Gy of gamma-ray radiation from 60Co source. The MnSOD levels were determined by the specific immunostaining and quantified by the laser densitometry. The MANOVA analysis and Tukey post-hoc test indicated significantly higher MnSOD levels in both groups of BC patients in relation to the respective controls ( F=25.166, p<0.001). The data indicated that the increased initial MnSOD levels in peripheral blood cells may be related to the presence of BC i.e. they may reflect the system response to the presence of malignant tumour. In addition to that, in vitro radiation challenge of blood cells indicate that MnSOD overexpression may be a good indicator for selection of BC patients that would express increased resistance to oxidative stress imposed by the clinical treatment. (author)

  20. The Effect of Low-Level Laser Therapy on Human Leukemic Cells

    OpenAIRE

    Dastanpour, Somayeh; Momen Beitollahi, Jalil; Saber, Kazem

    2015-01-01

    Introduction: Laser phototherapy is used for the treatment of chemotherapy-induced oral mucositis in patients with leukemia, although there are limited data supporting the safety of this method. This study aimed to evaluate the effect of different doses of low-level laser on proliferation of acute myeloid leukemia (AML) cell line (KG-1a) in vitro.

  1. Cytochrome P450 levels are altered in patients with esophageal squamous-cell carcinoma

    DEFF Research Database (Denmark)

    Bergheim, I.; Wolfgarten, E.; Bollschweiler, E.;

    2007-01-01

    AIM: To investigate the role of cytochrome P450 (CYP) in the carcinogenesis of squamous-cell carcinoma (SCC) in human esophagus by determining expression patterns and protein levels of representative CYPs in esophageal tissue of patients with SCC and controls. METHODS: mRNA expression of CYP2E1, ...

  2. Elevated circulating stromal-derived factor-1 levels in sickle cell disease

    NARCIS (Netherlands)

    Landburg, P P; Nur, E; Maria, N; Brandjes, D P M; Biemond, B J; Schnog, J B; Duits, A J

    2009-01-01

    Inflammation and angiogenesis are of importance in the pathophysiology of sickle cell disease (SCD). Recently, the chemokine stromal-derived factor-1 (SDF-1) has been shown to be a key mediator of angiogenesis and inflammation. In this study we determined serum SDF-1 levels in consecutive adult sick

  3. Two outward potassium current types are expressed during the neural differentiation of neural stem cells**

    Institute of Scientific and Technical Information of China (English)

    Ruiying Bai; Guowei Gao; Ying Xing; Hong Xue

    2013-01-01

    The electrophysiological properties of potassium ion channels are regarded as a basic index for determining the functional differentiation of neural stem cells. In this study, neural stem cells from the hippocampus of newborn rats were induced to differentiate with neurotrophic growth factor, and the electrophysiological properties of the voltage-gated potassium ion channels were observed. Immunofluorescence staining showed that the rapidly proliferating neural stem cells formed spheres in vitro that expressed high levels of nestin. The differentiated neurons were shown to express neuron-specific enolase. Flow cytometric analysis revealed that the neural stem cells were actively dividing and the percentage of cells in the S + G2/M phase was high. However, the ratio of cells in the S + G2/M phase decreased obviously as differentiation proceeded. Whole-cellpatch-clamp re-cordings revealed apparent changes in potassium ion currents as the neurons differentiated. The potassium ion currents consisted of one transient outward potassium ion current and one delayed rectifier potassium ion current, which were blocked by 4-aminopyridine and tetraethylammonium, respectively. The experimental findings indicate that neural stem cells from newborn rat hippo-campus could be cultured and induced to differentiate into functional neurons under defined condi-tions in vitro. The differentiated neurons expressed two types of outward potassium ion currents similar to those of mature neurons in vivo.

  4. Therapeutic and diagnostic set for irradiation the cell lines in low level laser therapy

    Science.gov (United States)

    Gryko, Lukasz; Zajac, Andrzej; Gilewski, Marian; Szymanska, Justyna; Goralczyk, Krzysztof

    2014-05-01

    In the paper is presented optoelectronic diagnostic set for standardization the biostimulation procedures performed on cell lines. The basic functional components of the therapeutic set are two digitally controlled illuminators. They are composed of the sets of semiconductor emitters - medium power laser diodes and high power LEDs emitting radiation in wide spectral range from 600 nm to 1000 nm. Emitters are coupled with applicator by fibre optic and optical systems that provides uniform irradiation of vessel with cell culture samples. Integrated spectrometer and optical power meter allow to control the energy and spectral parameters of electromagnetic radiation during the Low Level Light Therapy procedure. Dedicated power supplies and digital controlling system allow independent power of each emitter . It was developed active temperature stabilization system to thermal adjust spectral line of emitted radiation to more efficient association with absorption spectra of biological acceptors. Using the set to controlled irradiation and allowing to measure absorption spectrum of biological medium it is possible to carry out objective assessment the impact of the exposure parameters on the state cells subjected to Low Level Light Therapy. That procedure allows comparing the biological response of cell lines after irradiation with radiation of variable spectral and energetic parameters. Researches were carried out on vascular endothelial cell lines. Cells proliferations after irradiation of LEDs: 645 nm, 680 nm, 740 nm, 780 nm, 830 nm, 870 nm, 890 nm, 970 nm and lasers 650 nm and 830 nm were examined.

  5. Effect of low-level laser irradiation on odontoblast-like cells

    International Nuclear Information System (INIS)

    Low-level laser therapy (LLLT), also referred to as therapeutic laser, has been recommended for a wide array of clinical procedures, among which the treatment of dentinal hypersensitivity. However, the mechanism that guides this process remains unknown. Therefore, the objective of this study was to evaluate in vitro the effects of LLL irradiation on cell metabolism (MTT assay), alkaline phosphatase (ALP) expression and total protein synthesis. The expression of genes that encode for collagen type-1 (Col-1) and fibronectin (FN) was analyzed by RT-PCR. For such purposes, odontoblast-like cell line (MDPC-23) was previously cultured in Petri dishes (15000 cells/cm2) and submitted to stress conditions during 12 h. Thereafter, 6 applications with a monochromatic near infrared radiation (GaAlAs) set at predetermined parameters were performed at 12-h intervals. Non-irradiated cells served as a control group. Neither the MTT values nor the total protein levels of the irradiated group differed significantly from those of the control group (Mann-Whitney test; p > 0.05). On the other hand, the irradiated cells showed a decrease in ALP activity (Mann-Whitney test; p 0.05). It may be concluded that, under the tested conditions, the LLLT parameters used in the present study did not influence cell metabolism, but reduced slightly the expression of some specific proteins

  6. Visualization and measurement of ATP levels in living cells replicating hepatitis C virus genome RNA.

    Directory of Open Access Journals (Sweden)

    Tomomi Ando

    Full Text Available Adenosine 5'-triphosphate (ATP is the primary energy currency of all living organisms and participates in a variety of cellular processes. Although ATP requirements during viral lifecycles have been examined in a number of studies, a method by which ATP production can be monitored in real-time, and by which ATP can be quantified in individual cells and subcellular compartments, is lacking, thereby hindering studies aimed at elucidating the precise mechanisms by which viral replication energized by ATP is controlled. In this study, we investigated the fluctuation and distribution of ATP in cells during RNA replication of the hepatitis C virus (HCV, a member of the Flaviviridae family. We demonstrated that cells involved in viral RNA replication actively consumed ATP, thereby reducing cytoplasmic ATP levels. Subsequently, a method to measure ATP levels at putative subcellular sites of HCV RNA replication in living cells was developed by introducing a recently-established Förster resonance energy transfer (FRET-based ATP indicator, called ATeam, into the NS5A coding region of the HCV replicon. Using this method, we were able to observe the formation of ATP-enriched dot-like structures, which co-localize with non-structural viral proteins, within the cytoplasm of HCV-replicating cells but not in non-replicating cells. The obtained FRET signals allowed us to estimate ATP concentrations within HCV replicating cells as ∼5 mM at possible replicating sites and ∼1 mM at peripheral sites that did not appear to be involved in HCV replication. In contrast, cytoplasmic ATP levels in non-replicating Huh-7 cells were estimated as ∼2 mM. To our knowledge, this is the first study to demonstrate changes in ATP concentration within cells during replication of the HCV genome and increased ATP levels at distinct sites within replicating cells. ATeam may be a powerful tool for the study of energy metabolism during replication of the viral genome.

  7. Platelet-rich fibrin-induced bone marrow mesenchymal stem cell differentiation into osteoblast-like cells and neural cells

    Institute of Scientific and Technical Information of China (English)

    Qi Li; Yajun Geng; Lei Lu; Tingting Yang; Mingrui Zhang; Yanmin Zhou

    2011-01-01

    Bone marrow mesenchymal stem cells were allowed to develop for 14 days in a platelet-rich fibrin environment. Results demonstrated that platelet-rich fibrin significantly promoted bone marrow mesenchymal stem cell proliferation. In addition, there was a dose-dependent increase in Runt-related transcription factor-2 and bone morphogenetic protein-2 mRNA expression, as well as neuron-specific enolase and glial acidic protein. Results showed that platelet-rich fibrin promoted bone marrow mesenchymal stem cell proliferation and differentiation of osteoblastlike cells and neural cells in a dose-dependent manner.

  8. Relationship of chromosomal damage induced by caffeine to growth temperature and ATP level in proliferating cells.

    Science.gov (United States)

    Hernández, P; Mingo, R; González-Fernández, A; López-Sáez, J F

    1986-10-01

    Caffeine is known to induce chromosomal aberrations in proliferating cells when they are incubated during G2 and mitotic prophase. In the present paper, this caffeine effect has been analyzed in Allium cepa root meristems growing at different culture temperatures under steady-state kinetics. Caffeine (1-10 mM) induces chromosomal aberrations in a dose-dependent manner, and the treatment efficiency correlates linearly with the square of caffeine concentration. The efficiency of caffeine incubations, within the range 5-25 degrees C during equivalent cycle time periods has also been studied. It has been found that the lower the culture temperature, the higher the level of chromosomal aberrations. Moreover, at different temperatures, the level of chromosomal aberrations is a simple function of caffeine concentration and the ATP level. Therefore, the efficiency of caffeine treatment appears to be determined by some interaction between caffeine concentration and cellular ATP level. Our present results demonstrate that the influence of growth temperature on the chromosome-breaking effect of caffeine can be, at least partially, explained by the ATP levels during the incubation periods. In short, under different kinetics of plant cell proliferation, the ATP level, and/or something correlating with it, could explain the efficiency of caffeine in inducing chromosomal aberrations: the lower the ATP level, the higher the caffeine efficiency. PMID:3773927

  9. Measuring the dynamics of cyclic adenosine monophosphate level in living cells induced by low-level laser irradiation using bioluminescence resonance energy transfer

    Science.gov (United States)

    Huang, Yimei; Zheng, Liqin; Yang, Hongqin; Chen, Jiangxu; Wang, Yuhua; Li, Hui; Xie, Shusen; Zeng, Haishan

    2015-05-01

    Several studies demonstrated that the cyclic adenosine monophosphate (cAMP), an important second messenger, is involved in the mechanism of low-level laser irradiation (LLLI) treatment. However, most of these studies obtained the cAMP level in cell culture extracts or supernatant. In this study, the cAMP level in living cells was measured with bioluminescence resonance energy transfer (BRET). The effect of LLLI on cAMP level in living cells with adenosine receptors blocked was explored to identify the role of adenosine receptors in LLLI. The results showed that LLLI increased the cAMP level. Moreover, the rise of cAMP level was light dose dependent but wavelength independent for 658-, 785-, and 830-nm laser light. The results also exhibited that the adenosine receptors, a class of G protein-coupled receptor (GPCR), modulated the increase of cAMP level induced by LLLI. The cAMP level increased more significantly when the A3 adenosine receptors (A3R) were blocked by A3R antagonist compared with A1 adenosine receptor or A2a adenosine receptor blocked in HEK293T cells after LLLI, which was in good agreement with the adenosine receptors' expressions. All these results suggested that measuring the cAMP level with BRET could be a useful technique to study the role of GPCRs in living cells under LLLI.

  10. Oxidized low density lipoprotein increases RANKL level in human vascular cells. Involvement of oxidative stress

    International Nuclear Information System (INIS)

    Highlights: •Oxidized LDL enhances RANKL level in human smooth muscle cells. •The effect of OxLDL is mediated by the transcription factor NFAT. •UVA, H2O2 and buthionine sulfoximine also increase RANKL level. •All these effects are observed in human fibroblasts and endothelial cells. -- Abstract: Receptor Activator of NFκB Ligand (RANKL) and its decoy receptor osteoprotegerin (OPG) have been shown to play a role not only in bone remodeling but also in inflammation, arterial calcification and atherosclerotic plaque rupture. In human smooth muscle cells, Cu2+-oxidized LDL (CuLDL) 10–50 μg/ml increased reactive oxygen species (ROS) and RANKL level in a dose-dependent manner, whereas OPG level was not affected. The lipid extract of CuLDL reproduced the effects of the whole particle. Vivit, an inhibitor of the transcription factor NFAT, reduced the CuLDL-induced increase in RANKL, whereas PKA and NFκB inhibitors were ineffective. LDL oxidized by myeloperoxidase (MPO-LDL), or other pro-oxidant conditions such as ultraviolet A (UVA) irradiation, incubation with H2O2 or with buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, also induced an oxidative stress and enhanced RANKL level. The increase in RANKL in pro-oxidant conditions was also observed in fibroblasts and endothelial cells. Since RANKL is involved in myocardial inflammation, vascular calcification and plaque rupture, this study highlights a new mechanism whereby OxLDL might, by generation of an oxidative stress, exert a deleterious effect on different cell types of the arterial wall

  11. Low level light promotes the proliferation and differentiation of bone marrow derived mesenchymal stem cells

    Science.gov (United States)

    Ahn, Jin-Chul; Rhee, Yun-Hee; Choi, Sun-Hyang; Kim, Dae Yu; Chung, Phil-Sang

    2015-03-01

    Low-level light irradiation (LLLI) reported to stimulate the proliferation or differentiation of a variety of cell types. However, very little is known about the effect of light therapy on stem cells. The aim of the present study was to evaluate the effect of LLLI on the molecular physiological change of human bone marrow derived stem cells (hBMSC) by wavelength (470, 630, 660, 740 and 850, 50mW). The laser diode was performed with different time interval (0, 7.5, 15, 30J/cm2, 50mW) on hBMSC. To determine the molecular physiological changes of cellular level of hBMSC, the clonogenic assay, ATP assay, reactive oxygen species (ROS) detection, mitochondria membrane potential (MMPΦ) staining and calcium efflux assay were assessed after irradiation. There was a difference between with and without irradiation on hBMSCs. An energy density up to 30 J/cm² improved the cell proliferation in comparison to the control group. Among these irradiated group, 630 and 660nm were significantly increased the cell proliferation. The cellular level of ATP and calcium influx was increased with energy dose-dependent in all LLLI groups. Meanwhile, ROS and MMPΦ were also increased after irradiation except 470nm. It can be concluded that LLLI using infrared light and an energy density up to 30 J/cm² has a positive stimulatory effect on the proliferation or differentiation of hBMSCs. Our results suggest that LLLI may influence to the mitochondrial membrane potential activity through ATP synthesis and increased cell metabolism which leads to cell proliferation and differentiation.

  12. Serum ferritin levels in adults with sickle cell disease in Lagos, Nigeria

    Directory of Open Access Journals (Sweden)

    Akinbami AA

    2013-05-01

    Full Text Available Akinsegun A Akinbami,1 Adedoyin O Dosunmu,1 Adewumi A Adediran,3 Olajumoke O Oshinaike,2 Vincent O Osunkalu,1 Sarah O Ajibola,3 Olanrewaju M Arogundade11Department of Haematology and Blood Transfusion, 2Department of Medicine, Lagos State University, College of Medicine, Ikeja, Nigeria; 3Department of Haematology and Blood Transfusion, Faculty of Clinical Sciences, College of Medicine, Idiaraba, NigeriaBackground: Serum ferritin is considered to be one of the most important tools in the measurement of iron balance in steady-state sickle cell disease. Increased gastrointestinal absorption of iron has been reported in sickle cell disease because of the associated chronic hemolysis, and it is also thought that repeated red cell transfusion consequent to chronic hemolysis and anemia causes excessive iron levels. The aim of this study was to determine overall and gender-specific mean ferritin levels in patients with steady-state sickle cell disease in order to establish the prevalence of iron deficiency and overload.Methods: This was a cross-sectional study in homozygous patients with sickle cell disease attending the sickle cell clinic at Lagos State University Teaching Hospital, Ikeja. A 5 mL blood sample was collected in plain bottles from consenting participants during steady-state periods. The serum was separated and analyzed for ferritin by enzyme-linked immunosorbent assay. Another 5 mL sample was collected for a full blood count, done on the same day of collection, to determine red blood cell indices, ie, mean cell volume, mean cell hemoglobin concentration, and mean corpuscular hemoglobin concentration. The Pearson Chi-square test was used for statistical analysis. The differences were considered to be statistically significant when P was 300 ng/mL. Ninety-three subjects (90.29% had serum ferritin within the normal reference range of 15–300 ng/mL.Conclusion: In this study, 90% of subjects with sickle cell disease had normal iron stores

  13. Small Cell Carcinoma of the Urinary Bladder: KIT and PDGFRA Gene Mutations

    OpenAIRE

    Nuket Eliyakin; Hakan Postaci; Yasemin Baskin; Zafer Kozacioğlu

    2015-01-01

    Primary small cell carcinoma of the urinary bladder is very rare. A 72-year-old was admitted to our hospital because of hematuria and dysuria. Cystoscopy revealed a bladder full of multiple, solid and papillary tumors. Biopsies from the deep and papillary tumors were taken. Histologically, tumor was pure small cell carcinoma. Immunohistochemically, the tumor cells were positive for cytokeratin, chromogranin, synaptophysin, neuron-specific enolase, CD56, CD117 and Ki67 (labeling 70%). The tumo...

  14. Unraveling the genetic driving forces enabling antibiotic resistance at the single cell level

    Science.gov (United States)

    Bos, Julia

    Bacteria are champions at finding ways to quickly respond and adapt to environments like the human gut, known as the epicentre of antibiotic resistance. How do they do it? Combining molecular biology tools to microfluidic and fluorescence microscopy technologies, we monitor the behavior of bacteria at the single cell level in the presence of non-toxic doses of antibiotics. By tracking the chromosome dynamics of Escherichia coli cells upon antibiotic treatment, we examine the changes in the number, localization and content of the chromosome copies within one cell compartment or between adjacent cells. I will discuss how our work pictures the bacterial genomic plasticity as a driving force in evolution and how it provides access to the mechanisms controlling the subtle balance between genetic diversity and stability in the development of antibiotic resistance.

  15. Childrens' learning and behaviour and the association with cheek cell polyunsaturated fatty acid levels.

    Science.gov (United States)

    Kirby, A; Woodward, A; Jackson, S; Wang, Y; Crawford, M A

    2010-01-01

    Increasing interest in the role of omega-3 fatty acids in relation to neurodevelopmental disorders (e.g. ADHD, dyslexia, autism) has occurred as a consequence of some international studies highlighting this link. In particular, some studies have shown that children with ADHD may have lower concentrations of polyunsaturated fatty acids (PUFAs), particularly omega-3, in their red blood cells and plasma, and that supplementation with omega-3 fatty acids may alleviate behavioural symptoms in this population. However, in order to compare levels it seems appropriate to establish fatty acid levels in a mainstream school aged population and if levels relate to learning and behaviour. To date no study has established this. For this study, cheek cell samples from 411 typically developing school children were collected and analysed for PUFA content, in order to establish the range in this population. In addition, measures of general classroom attention and behaviour were assessed in these children by teachers and parents. Cognitive performance tests were also administered in order to explore whether an association between behaviour and/or cognitive performance and PUFA levels exists. Relationships between PUFA levels and socio-economic status were also explored. Measures of reading, spelling and intelligence did not show any association with PUFA levels, but some associations were noted with the level of omega-3 fatty acids and teacher and parental reports of behaviour, with some evidence that higher omega-3 levels were associated with decreased levels of inattention, hyperactivity, emotional and conduct difficulties and increased levels of prosocial behaviour. These findings are discussed in relation to previous findings from omega-3 supplementation studies with children. PMID:20172688

  16. The Radiotherapeutic Significance of Serum NSE Level in Non-Small Cell Lung Cancers (NSCLC)

    International Nuclear Information System (INIS)

    From December 1989 to February 1993, 108 patients with Non-Small Cell Lung Cancers(NSCLC) were studied retrospectively to evaluate radiotherapeutic significance of serum levels of NSE. We considered elevated serum neuron specific pathologic evaluation revealed 86 squamous cell carcinomas, 11 adenocarcinomas, 3 large cell carcinomas, 3 mucoepidermoid carcinomas, and 5 unknown pathology. Eight patients had stage I, 40 stage IIIA, and 60 stage IIIB. S-NSE level greater than 15 ng/ml was considered as elevated, and below this considered as normal. All patients received radiotherapy as primary treatment modality. The responders to radiotherapy had significantly higher mean S-NSE level than on-responders (28.5 ng/ml vs 20 ng/ml, p=0.01). Overall 2-year survival rate (YSR) was 23.6%. According to radiotherapy response, 2 YSR for patients with CR, PR, and NR were 39.2%, 28.6%, and 6.2% respectively (p=0.001). 2 YSR for patients with elevated and normal S-NSE were 14.6% and 31.7%(p=0.02). The patients with NR showed no difference in survival according to S-NSE level. When we considered all patients, S-NSE level showed no significant impact on response. But for squamous cell cardinomas alone, patients with elevated S-NSE had more patients with higher nodal stage. Based on our and other data, NSCLSC with neuroendocrine features have different response to treatment and clinical behavior compared to other NSCLSC. Thus, this subgroup may need different treatment modality, and S-NSE level may have prognostic significance

  17. Low-level laser irradiation effect on endothelial cells under conditions of hyperglycemia.

    Science.gov (United States)

    Góralczyk, Krzysztof; Szymańska, Justyna; Szot, Katarzyna; Fisz, Jacek; Rość, Danuta

    2016-07-01

    Diabetes mellitus is considered to be a very serious lifestyle disease leading to cardiovascular complications and impaired wound healing observed in the diabetic foot syndrome. Chronic hyperglycemia is the source of the endothelial activation. The inflammatory process in diabetes is associated with the secretion of inflammatory cytokines by endothelial cells, e.g., tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6). The method of phototherapy using laser beam of low power (LLLT-low-level laser therapy) effectively supports the conventional treatment of diabetic vascular complications such as diabetic foot syndrome. The aim of our study was to evaluate the effect of low-power laser irradiation at two wavelengths (635 and 830 nm) on the secretion of inflammatory factors (TNF-α and IL-6) by the endothelial cell culture-HUVEC line (human umbilical vein endothelial cell)-under conditions of hyperglycemia. It is considered that adverse effects of hyperglycemia on vascular endothelial cells may be corrected by the action of LLLT, especially with the wavelength of 830 nm. It leads to the reduction of TNF-α concentration in the supernatant and enhancement of cell proliferation. Endothelial cells play an important role in the pathogenesis of diabetes; however, a small number of studies evaluate an impact of LLLT on these cells under conditions of hyperglycemia. Further work on this subject is warranted. PMID:26861982

  18. Biological effects of low-level laser irradiation on umbilical cord mesenchymal stem cells

    Science.gov (United States)

    Chen, Hongli; Wang, Hong; Li, Yingxin; Liu, Weichao; Wang, Chao; Chen, Zhuying

    2016-04-01

    Low-level laser irradiation (LLLI) can enhance stem cell (SC) activity by increasing migration and proliferation. This study investigated the effects of LLLI on proliferation, enzymatic activity, and growth factor production in human umbilical cord mesenchymal SCs (hUC-MSCs) as well as the underlying mechanisms. hUC-MSCs were assigned to a control group (non-irradiation group) and three LLLI treatment groups (635 nm group, 808 nm group, and 635/808 nm group). Laser power density and energy density of 20 mW/cm2 and 12 J/cm2, respectively, were used for each experiment. The proliferation rate was higher in the 635 nm as compared to the other groups. LLLI at 808 nm did not induce cell proliferation. ROS levels in cells exposed to 635, 808, and 635/808 nm radiation were increased by 52.81%, 26.89%, and 21.15%, respectively, relative to the control group. CAT, tGPx, and SOD activity was increased. LLLI at 808 nm increased the levels of IL-1, IL-6, and NFκB but not VEGF. LLLI improved hUC-MSCs function and increased antioxidant activity. Dual-wavelength LLLI had more potent effects on hUC-MSCs than single-wavelength treatment. LLLI has potential applications in the preconditioning of hUC-MSCs in vitro prior to transplantation, which could improve the regenerative capacity of cells.

  19. Biological effects of low-level laser irradiation on umbilical cord mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Hongli Chen

    2016-04-01

    Full Text Available Low-level laser irradiation (LLLI can enhance stem cell (SC activity by increasing migration and proliferation. This study investigated the effects of LLLI on proliferation, enzymatic activity, and growth factor production in human umbilical cord mesenchymal SCs (hUC-MSCs as well as the underlying mechanisms. hUC-MSCs were assigned to a control group (non-irradiation group and three LLLI treatment groups (635 nm group, 808 nm group, and 635/808 nm group. Laser power density and energy density of 20 mW/cm2 and 12 J/cm2, respectively, were used for each experiment. The proliferation rate was higher in the 635 nm as compared to the other groups. LLLI at 808 nm did not induce cell proliferation. ROS levels in cells exposed to 635, 808, and 635/808 nm radiation were increased by 52.81%, 26.89%, and 21.15%, respectively, relative to the control group. CAT, tGPx, and SOD activity was increased. LLLI at 808 nm increased the levels of IL-1, IL-6, and NFκB but not VEGF. LLLI improved hUC-MSCs function and increased antioxidant activity. Dual-wavelength LLLI had more potent effects on hUC-MSCs than single-wavelength treatment. LLLI has potential applications in the preconditioning of hUC-MSCs in vitro prior to transplantation, which could improve the regenerative capacity of cells.

  20. Zinc finger protein 521 overexpression increased transcript levels of Fndc5 in mouse embryonic stem cells

    Indian Academy of Sciences (India)

    Motahere-Sadat Hashemi; Abbas Kiani Esfahani; Maryam Peymani; Alireza Shoaraye Nejati; Kamran Ghaedi; Mohammad Hossein Nasr-Esfahani; Hossein Baharvand

    2016-03-01

    Zinc finger protein 521 is highly expressed in brain, neural stem cells and early progenitors of the human hematopoietic cells. Zfp521 triggers the cascade of neurogenesis inmouse embryonic stemcells through inducing expression of the early neuroectodermal genes Sox1, Sox3 and Pax6. Fndc5, a precursor of Irisin has inducing effects on the expression level of brain derived neurotrophic factor in hippocampus. Therefore, it is most likely that Fndc5 may play an important role in neural differentiation. To exhibit whether the expression of this protein is under regulation with Zfp521, we overexpressed Zfp521 in a stable transformants of mESCs expressing EGFP under control of Fndc5 promoter. Increased expression of Zfp521 enhanced transcription levels of both EGFP and endogenous Fndc5. This result was confirmed by overexpression the aforementioned vectors in HEK cells and indicated that Zfp521 functions upstream of Fndc5 expression. It is most likely that Zfp521 may act through the binding to its response element on Fndc5 core promoter. Therefore it is concluding that an enhanced expression of Fndc5 in neural progenitor cells is stimulated by Zfp521 overexpression in these cells.

  1. Effect of vitamin E levels on the cell-mediated immunity of broilers vaccinated against coccidiosis

    Directory of Open Access Journals (Sweden)

    ICM da Silva

    2011-03-01

    Full Text Available Studies on the relationships between animal nutrition and immunity have sought reliable methodologies to measure responses. Cell-mediated immune response is similarly studied in humans. The cutaneous basophil hypersensitivity test (CBH is one of the methods to measure that response and consists in the infiltration of inflammatory cells, particularly of lymphocytes and basophils, as result of the application of substances capable of inducing cell proliferation in determined sites, such as wings, wattle, and interdigital space in birds. CBH is considered a simple and fast method and can be applied in birds of different ages. In immunocompetence studies with poultry, phytohemagglutinin-P (PHA-P is a commonly used substance, despite the variability of the response related to the method of application (intradermal injection and the antigens used. In the present experiment, PHA-P was used to observe the cell-mediated immune response of 216 chicks fed three dietary levels of vitamin E from 1 to 36 days of age. All birds were immunologically challenged by vaccination against coccidiosis at three days of age and against Newcastle Disease (NCD at 14 and 30 days of age. At 36 days of age, birds were submitted to the CBH test according to the methodology of Corrier & DeLoach (1990. Birds fed 65mg/kg of vitamin E presented lasting cell reaction (p<0.08, which indicates that this vitamin E level improved cell immune response of birds due to its antioxidant and immunomodulating properties. The use of this vitamin E level can be considered by nutritionists under practical conditions, aiming to improve broiler immunity.

  2. Reduced Levels of Proteasome Products in a Mouse Striatal Cell Model of Huntington's Disease.

    Directory of Open Access Journals (Sweden)

    Sayani Dasgupta

    Full Text Available Huntington's disease is the result of a long polyglutamine tract in the gene encoding huntingtin protein, which in turn causes a large number of cellular changes and ultimately results in neurodegeneration of striatal neurons. Although many theories have been proposed, the precise mechanism by which the polyglutamine expansion causes cellular changes is not certain. Some evidence supports the hypothesis that the long polyglutamine tract inhibits the proteasome, a multiprotein complex involved in protein degradation. However, other studies report normal proteasome function in cells expressing long polyglutamine tracts. The controversy may be due to the methods used to examine proteasome activity in each of the previous studies. In the present study, we measured proteasome function by examining levels of endogenous peptides that are products of proteasome cleavage. Peptide levels were compared among mouse striatal cell lines expressing either 7 glutamines (STHdhQ7/Q7 or 111 glutamines in the huntingtin protein, either heterozygous (STHdhQ7/Q111 or homozygous (STHdhQ111/Q111. Both of the cell lines expressing huntingtin with 111 glutamines showed a large reduction in nearly all of the peptides detected in the cells, relative to levels of these peptides in cells homozygous for 7 glutamines. Treatment of STHdhQ7/Q7 cells with proteasome inhibitors epoxomicin or bortezomib also caused a large reduction in most of these peptides, suggesting that they are products of proteasome-mediated cleavage of cellular proteins. Taken together, these results support the hypothesis that proteasome function is impaired by the expression of huntingtin protein containing long polyglutamine tracts.

  3. Defect levels in Cu2ZnSn(SxSe1-x)4 solar cells probed by current-mode deep level transient spectroscopy

    Science.gov (United States)

    Das, Sandip; Chaudhuri, Sandeep K.; Bhattacharya, Raghu N.; Mandal, Krishna C.

    2014-05-01

    Defect levels in kesterite Cu2ZnSn(S,Se)4 (CZTSSe) solar cells have been investigated by current-mode deep level transient spectroscopy. Experiments were carried out on two CZTSSe cells with photoconversion efficiencies of 4.1% and 7.1% measured under AM 1.5 illumination. The absorber layer of the 4.1% efficiency cell was prepared by annealing evaporated ZnS/Cu/Sn stacked precursor under S/Se vapor, while the absorber of the 7.1% efficiency cell was prepared by co-evaporation of the constituent elements. The 4.1% efficiency CZTSSe cell with a S/(S + Se) ratio of 0.58 exhibited two dominant deep acceptor levels at Ev + 0.12 eV, and Ev + 0.32 eV identified as CuZn(-/0) and CuSn(2-/-) antisite defects, respectively. The 7.1% efficiency cell with purely Se composition S/(S + Se) = 0 showed only one shallow level at Ev + 0.03 eV corresponding to Cu-vacancy (VCu). Our results revealed that VCu is the primary defect center in the high-efficiency kesterite solar cell in contrast to the detrimental CuZn and CuSn antisites found in the low efficiency CZTSSe cells limiting the device performance.

  4. Effects of arsenite on UROtsa cells: low-level arsenite causes accumulation of ubiquitinated proteins that is enhanced by reduction in cellular glutathione levels

    International Nuclear Information System (INIS)

    Chronic arsenic exposure increases risk for the development of diabetes, vascular disease, and cancers of the skin, lung, kidney, and bladder. This study investigates the effects of arsenite [As(III)] on human urothelial cells (UROtsa). As(III) toxicity was determined by exposing confluent UROtsa cells to As(III) (0.5-200 μM). Depleting cellular glutathione levels with buthionine sulfoximine (BSO) potentiated the toxicity of As(III). Cell viability was assessed with the (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. UROtsa cell ability to biotransform As(III) was determined by dosing cells with environmentally relevant concentrations of As(III) followed by HPLC/ICP-MS analysis of cell media and lysate. Both pentavalent and trivalent monomethylated products were detected. Although cytotoxicity was observed at high doses of As(III) (approximately 100 μM) in UROtsa cells, perturbations of a variety of molecular processes occurred at much lower doses. Exposure to low-level As(III) (0.5-25 μM) causes an accumulation of ubiquitin (Ub)-conjugated proteins. This effect is enhanced when cellular glutathione levels have been reduced with BSO treatment. Because As(III) has many effects on UROtsa cells, a greater understanding of how As(III) is affecting cellular proteins in a target tissue will lead to a better understanding of the mechanism of toxicity and pathogenesis for low-level As(III)

  5. Low-level laser therapy in 3D cell culture model using gingival fibroblasts.

    Science.gov (United States)

    Basso, Fernanda G; Soares, Diana G; de Souza Costa, Carlos Alberto; Hebling, Josimeri

    2016-07-01

    Besides extensive data about the effects of low-level laser therapy (LLLT) on different cell types, so far, these results were obtained from monolayer cell culture models, which have limitations in terms of cell morphology and phenotype expression. Therefore, for better in vitro evaluation of the effects of LLLT, this study was performed with a 3D cell culture model, where gingival fibroblasts were seeded in collagen matrix. Cells isolated from a healthy patient were seeded in wells of 24-well plates with culture medium (DMEM) supplemented with 10 % fetal bovine serum and collagen type I solution. After 5 days, a serum-free DMEM was added to the matrices with cells that were subjected or not to three consecutive irradiations of LLLT by means of the LaserTABLE diode device (780 nm, 25 mW) at 0.5, 1.5, and 3 J/cm(2). Twenty-four hours after the last irradiation, cell viability and morphology as well as gene expression of growth factors were assessed. Histological evaluation of matrices demonstrated uniform distribution and morphology of gingival fibroblasts within the collagen matrix. LLLT at 3 J/cm(2) increased gingival fibroblast viability. Enhanced gene expression of hCOL-I and hEGF was observed for 0.5 J/cm(2), while no significant changes were detected for the other irradiation densities tested. In conclusion, LLLT promoted biostimulation of gingival fibroblasts seeded in a 3D cell culture model, demonstrating that this model can be applied for phototherapy studies and that LLLT could penetrate the collagen matrix to increase cell functions related to tissue repair. PMID:27126408

  6. Increased viability of odontoblast-like cells subjected to low-level laser irradiation

    Science.gov (United States)

    Oliveira, C. F.; Basso, F. G.; Lins, E. C.; Kurachi, C.; Hebling, J.; Bagnato, V. S.; de Souza Costa, C. A.

    2010-07-01

    Studies have shown that the increase of cell metabolism depends on the low level laser therapy (LLLT) parameters used to irradiate the cells. However, the optimal laser dose to up-regulate pulp cell activity remains unknown. Consequently, the aim of this study was to evaluate the metabolic response of odontoblast-like cells (MDPC-23) exposed to different LLLT doses. Cells at 20000 cells/cm2 were seeded in 24-well plates using plain culture medium (DMEM) and were incubated in a humidified incubator with 5% CO2 at 37°C. After 24 h, the culture medium was replaced by fresh DMEM supplemented with 5% (stress by nutritional deficit) or 10% fetal bovine serum (FBS). The cells were exposed to different laser doses from a near infrared diode laser prototype designed to provide a uniform irradiation of the wells. The experimental groups were: G1: 1.5 J/cm2 + 5% FBS; G2: 1.5 J/cm2 + 10% FBS; G3: 5 J/cm2 + 5% FBS; G4: 5 J/cm2 + 10% FBS; G5: 19 J/cm2 + 5% FBS; G6: 19 J/cm2 + 10% FBS. LLLT was performed in 3 consecutive irradiation cycles with a 24-hour interval. Non-irradiated cells cultured in DMEM supplemented with either 5 or 10% FBS served as control groups. The analysis of the metabolic response was performed by the MTT assay 3 h after the last irradiation. G1 presented an increase in SDH enzyme activity and differed significantly (Mann-Whitney test, p LLLT stimulated the metabolic activity of MDPC-23 cultured in DMEM supplemented with 5% FBS and exposed to a laser dose of 1.5 J/cm2. These findings are relevant for further studies on the action of near infrared lasers on cells with odontoblast phenotype.

  7. Relationships between cell cycle regulator gene copy numbers and protein expression levels in Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Ayako Chino

    Full Text Available We previously determined the copy number limits of overexpression for cell division cycle (cdc regulatory genes in the fission yeast Schizosaccharomyces pombe using the "genetic tug-of-war" (gTOW method. In this study, we measured the levels of tandem affinity purification (TAP-tagged target proteins when their copy numbers are increased in gTOW. Twenty analyzed genes showed roughly linear correlations between increased protein levels and gene copy numbers, which suggested a general lack of compensation for gene dosage in S. pombe. Cdc16 and Sid2 protein levels but not their mRNA levels were much lower than that expected by their copy numbers, which suggested the existence of a post-transcriptional down regulation of these genes. The cyclin Cig1 protein level and its mRNA level were much higher than that expected by its copy numbers, which suggested a positive feedback mechanism for its expression. A higher Cdc10 protein level and its mRNA level, probably due to cloning its gene into a plasmid, indicated that Cdc10 regulation was more robust than that previously predicted.

  8. Relationships between cell cycle regulator gene copy numbers and protein expression levels in Schizosaccharomyces pombe.

    Science.gov (United States)

    Chino, Ayako; Makanae, Koji; Moriya, Hisao

    2013-01-01

    We previously determined the copy number limits of overexpression for cell division cycle (cdc) regulatory genes in the fission yeast Schizosaccharomyces pombe using the "genetic tug-of-war" (gTOW) method. In this study, we measured the levels of tandem affinity purification (TAP)-tagged target proteins when their copy numbers are increased in gTOW. Twenty analyzed genes showed roughly linear correlations between increased protein levels and gene copy numbers, which suggested a general lack of compensation for gene dosage in S. pombe. Cdc16 and Sid2 protein levels but not their mRNA levels were much lower than that expected by their copy numbers, which suggested the existence of a post-transcriptional down regulation of these genes. The cyclin Cig1 protein level and its mRNA level were much higher than that expected by its copy numbers, which suggested a positive feedback mechanism for its expression. A higher Cdc10 protein level and its mRNA level, probably due to cloning its gene into a plasmid, indicated that Cdc10 regulation was more robust than that previously predicted. PMID:24019917

  9. Efficient Quantitative Analysis of Carboxyalkylpyrrole Ethanolamine Phospholipids: Elevated Levels in Sickle Cell Disease Blood.

    Science.gov (United States)

    Guo, Junhong; Wang, Hua; Hrinczenko, Borys; Salomon, Robert G

    2016-07-18

    γ-Hydroxy-α,β-unsaturated aldehydes, generated by oxidative damage of polyunsaturated phospholipids, form pyrrole derivatives that incorporate the ethanolamine phospholipid (EP) amino group such as 2-pentylpyrrole (PP)-EP and 2-(ω-carboxyalkyl)pyrrole (CAP)-EP derivatives: 2-(ω-carboxyethyl)pyrrole (CEP)-EP, 2-(ω-carboxypropyl)pyrrole (CPP)-EP, and 2-(ω-carboxyheptyl)pyrrole (CHP)-EP. Because EPs occur in vivo in various forms, a complex mixture of pyrrole-modified EPs with different molecular weights is expected to be generated. To provide a sensitive index of oxidative stress, all of the differences in mass related to the glycerophospholipid moieties were removed by releasing a single CAP-ethanolamine (ETN) or PP-ETN from each mixture by treatment with phospholipase D. Accurate quantization was achieved using the corresponding ethanolamine-d4 pyrroles as internal standards. The product mixture obtained by phospholipolysis of total blood phospholipids from sickle cell disease (SCD) patients was analyzed by LC-MS/MS. The method was applied to measure CAP-EP and PP-EP levels in blood plasma from clinical monitoring of SCD patients. We found uniformly elevated blood levels of CEP-EP (63.9 ± 9.7 nM) similar to mean levels in blood from age-related macular degeneration (AMD) patients (56.3 ± 37.1 nM), and 2-fold lower levels (27.6 ± 3.6 nM, n = 5) were detected in plasma from SCD patients hospitalized to treat a sickle cell crisis, although mean levels remain higher than those (12.1 ± 10.5 nM) detected in blood from healthy controls. Plasma levels of CPP-EPs from SCD clinic patients were 4-fold higher than those of SCD patients hospitalized to treat a sickle cell crisis (45.1 ± 10.9 nM, n = 5 versus 10.9 ± 3.4 nM, n = 6; p < 0.002). PP-EP concentration in plasma from SCD clinic patients is nearly 4.8-fold higher than its level in plasma samples from SCD patients hospitalized to treat a sickle cell crisis (7.06 ± 4.05 vs 1.48 ± 0.92 nM; p < 0.05). Because

  10. Changes in peripheral blood level of regulatory T cells in patients with malignant melanoma during treatment with dendritic cell vaccination and low-dose IL-2

    DEFF Research Database (Denmark)

    Bjoern, J; Brimnes, M K; Andersen, M H;

    2011-01-01

    In this study, changes in peripheral blood regulatory T cell (Treg) levels were evaluated in 46 progressive patients with melanoma treated with a dendritic cell-based vaccine and concomitant low-dose IFN-a and IL-2. The regulatory subset of CD4 T cells, characterized by CD25(high) , was prospecti......In this study, changes in peripheral blood regulatory T cell (Treg) levels were evaluated in 46 progressive patients with melanoma treated with a dendritic cell-based vaccine and concomitant low-dose IFN-a and IL-2. The regulatory subset of CD4 T cells, characterized by CD25(high) , was...

  11. Changes in peripheral blood level of regulatory T cells in patients with malignant melanoma during treatment with dendritic cell vaccination and low-dose IL-2

    DEFF Research Database (Denmark)

    Bjoern, J; Brimnes, M K; Andersen, M H;

    2011-01-01

    In this study, changes in peripheral blood regulatory T cell (Treg) levels were evaluated in 46 progressive patients with melanoma treated with a dendritic cell-based vaccine and concomitant low-dose IFN-α and IL-2. The regulatory subset of CD4 T cells, characterized by CD25(high) , was prospecti......In this study, changes in peripheral blood regulatory T cell (Treg) levels were evaluated in 46 progressive patients with melanoma treated with a dendritic cell-based vaccine and concomitant low-dose IFN-α and IL-2. The regulatory subset of CD4 T cells, characterized by CD25(high) , was...

  12. High-throughput imaging method for direct assessment of GM1 ganglioside levels in mammalian cells

    Directory of Open Access Journals (Sweden)

    Walter Acosta

    2016-03-01

    Full Text Available GM1-gangliosidosis is an inherited autosomal recessive disorder caused by mutations in the gene GLB1, which encodes acid β-galactosidase (β-gal. The lack of activity in this lysosomal enzyme leads to accumulation of GM1 gangliosides (GM1 in cells. We have developed a high-content-imaging method to assess GM1 levels in fibroblasts that can be used to evaluate substrate reduction in treated GLB1−/− cells [1]. This assay allows fluorescent quantification in a multi-well system which generates unbiased and statistically significant data. Fluorescently labeled Cholera Toxin B subunit (CTXB, which specifically binds to GM1 gangliosides, was used to detect in situ GM1 levels in a fixed monolayer of fibroblasts. This sensitive, rapid, and inexpensive method facilitates in vitro drug screening in a format that allows a high number of replicates using low working volumes.

  13. High-throughput imaging method for direct assessment of GM1 ganglioside levels in mammalian cells.

    Science.gov (United States)

    Acosta, Walter; Martin, Reid; Radin, David N; Cramer, Carole L

    2016-03-01

    GM1-gangliosidosis is an inherited autosomal recessive disorder caused by mutations in the gene GLB1, which encodes acid β-galactosidase (β-gal). The lack of activity in this lysosomal enzyme leads to accumulation of GM1 gangliosides (GM1) in cells. We have developed a high-content-imaging method to assess GM1 levels in fibroblasts that can be used to evaluate substrate reduction in treated GLB1(-/-) cells [1]. This assay allows fluorescent quantification in a multi-well system which generates unbiased and statistically significant data. Fluorescently labeled Cholera Toxin B subunit (CTXB), which specifically binds to GM1 gangliosides, was used to detect in situ GM1 levels in a fixed monolayer of fibroblasts. This sensitive, rapid, and inexpensive method facilitates in vitro drug screening in a format that allows a high number of replicates using low working volumes. PMID:26958633

  14. The levels of HDAC1 and thioredoxin1 are related to the death of mesothelioma cells by suberoylanilide hydroxamic acid.

    Science.gov (United States)

    You, Bo Ra; Park, Woo Hyun

    2016-05-01

    Mesothelioma is an aggressive tumor which is mainly derived from the pleura of lung. In the present study, we evaluated the anticancer effect of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor on human mesothelioma cells in relation to the levels of HDAC1, reactive oxygen species (ROS) and thioredoxin (Trx). While 1 µM SAHA inhibited cell growth in Phi and ROB cells at 24 h, it did not affect the growth in ADA and Mill cells. Notably, the level of HDAC1 was relatively overexpressed among Phi, REN and ROB cells. SAHA induced necrosis and apoptosis, which was accompanied by the cleavages of PARP and caspase-3 in Phi cells. This agent also increased the loss of mitochondrial membrane potential (MMP, ΔΨm) in Phi cells. All the tested caspase inhibitors attenuated apoptosis in SAHA-treated Phi cells whereas HDAC1 siRNA enhanced the apoptotic cell death. SAHA increased intracellular ROS levels including O2•- in Phi cells. N-acetyl cysteine (NAC) and vitamin C (Vit.C) significantly reduced the growth inhibition and death of Phi cells caused by SAHA. This drug decreased the mRNA and protein levels of Trx1 in Phi and ROB cells. Furthermore, Trx1 siRNA increased cell death and O2•- level in SAHA-treated Phi cells. In conclusion, SAHA selectively inhibited the growth of Phi and ROB mesothelioma cells, which showed the higher basal level of HDAC1. SAHA-induced Phi cell death was related to oxidative stress and Trx1 levels. PMID:26936390

  15. The effects of insulin on the expression levels of ADAMTS6 & 19 in OUMS-27 cell

    Directory of Open Access Journals (Sweden)

    Veli Uğurcu

    2014-09-01

    Full Text Available Objective: A Disintegrin-like Metalloproteinase with Thrombospondin Motifs (ADAMTS proteins are kind of matrix metalloproteinase enzymes that primarily founds in the extracellular matrix (ECM. Insulin is an important anabolic hormone, which acts on many tissues. The aim of this study is to evaluate the time-dependent effects of insulin on the two functionally unknown enzyme expressions (ADAMTS6 & 19 in OUMS-27 human chondrosarcoma cell line. Methods:OUMS-27 cells were cultured in Dulbecco’s modified Eagle’ medium (DMEM alone and DMEM containing 10 μg/mL insulin. The medium was changed every other day up to 11th day. Cells were harvested at 1, 3, 7,and 11th days and RNA isolation was performed at appropriate times according to study setup. The levels of RNA expression of ADAMTS6 and 19 were estimated by qRT-PCR using appropriate primers. Results: According to qRT-PCR analysis, ADAMTS6 mRNA expression was found to be decreased as early as one day after insulin application and continued up to day 11, the last day of insulin induction (p=0.008. The ratio of ADAMTS6 in insulin-applied groups was changed between 1/2 and 1/4 of control values. The changes in ADAMTS19 mRNA levels in insulin-applied cells were not statistically significant compared to control cell group. Conclusion:Our results demonstrated that insulin has a potential effect on alleviation of loss of extracellular matrix compounds by diminishing ADAMTS6 levels. To test this hypothesis and finding, more investigations are needed to recognize the real functions of orphan ADAMTS proteins.

  16. Protection against UVA-induced photooxidative damage in mammalian cell lines expressing increased levels of metallothionein

    International Nuclear Information System (INIS)

    Metallothionein (MT) is an endogenous low molecular weight protein that is inducible in a variety of eukaryotic cells and has the ability to selectivity bind heavy metal ions such as zinc and the cadmium. Although the exact physiological role of MT is still not understood, there is strong evidence that MT is involved in providing cellular resistance against the damaging effects of heavy metals and in the regulation of intracellular zinc and copper. Recently, it has been demonstrated that MT can scavenge radiation-induced reactive oxygen intermediates in vitro, specifically hydroxyl and superoxide radicals, and because of these observations it has been suggested that MT may provide protection against radiation-induced oxidative stress in vivo. Cell lines expressing increased levels of MT have demonstrated resistance to ionizing radiation, to ultraviolet radiation, and also to various DNA damaging agents including melphalan and cis-diaminedichloroplatinum. It is therefore important to gain some insight into the relationship between cellular MT content and cellular resistance to radiation and other DNA damaging agents. In this study we investigated the role of MT in providing protection against monochromatic 365-nm UVA radiation, which is known to generate intracellular reactive oxygen species that are involved in both DNA damage and cell killing. For this purpose, we used zinc acetate, a potent inducer of MT, to elevate MT levels in V79 Chinese hamster fibroblasts prior to UVA exposure and determined cell survival for uninduced and induced cultures. In order to eliminate any zinc effects other than MT induction, we also isolated and characterized cadmium chloride-resistant clones of V79 cells that have increased steady-state levels of both MT mRNA and protein, and we examined their survival characteristics against 365-nm radiation in the absence of zinc acetate. 14 refs., 3 figs

  17. Deep level transient spectroscopy measurements on CuInS2-thin film solar cells

    International Nuclear Information System (INIS)

    During the last decade CuInS2 was investigated for its use as absorber in thin film solar cells. Now these cells are ready for volume production. The advantages against already used materials are e. g. high absorbing capacity and cost-efficient and sustainable production. Because of the great discrepancy between predicted degree of efficiency and the already reached degree more investigations are necessary. To get a better understanding of the electron transport and recombination in order to arise efficiency we characterize the solar cells by deep level transient spectroscopy (DLTS). This method gives information about crystal defects depending on their electric position. Transient capacity measurements in the range of 25 K and 350 K allow us to determine activation energy and concentration of electron traps

  18. Expression Level of IL-6 Secreted by Bone Marrow Stromal Cells in Mice with Aplastic Anemia

    OpenAIRE

    Yong Feng Chen; Zhong Min Wu; Cong Xie; Shi Bai; Li Dong Zhao

    2013-01-01

    Parasecretion of the hematopoietic cytokines is considered as one of the mechanisms account for bone marrow hematopoiesis disorder. In this study, the level of IL-6 secreted by bone marrow stromal cells from a mouse model of aplastic anemia was analyzed. The aplastic anemia mouse model was established with cyclophosphamide in combination with chloramphenicol and 60Co γ radiation. The impairment of bone marrow hematopoiesis induced by irradiation and chemotherapeutic drugs was subsequently cha...

  19. Effect of T cell subset and inflammatory cytokine levels on prognosis in patients with pulmonary tuberculosis

    Institute of Scientific and Technical Information of China (English)

    Cheng-Zhou Wu; Yan-Qiao Wu

    2016-01-01

    Objective:To explore the effect of T cell subset and inflammatory cytokine levels on the prognosis in patients with pulmonary tuberculosis.Methods:A total of 72 patients confirmed with pulmonary tuberculosis who were admitted in our hospital from February, 2013 to February, 2015 were included in the study and served as the experiment group, among which 58 cases had active tuberculosis, and 14 cases had static tuberculosis; while 50 healthy individuals who came for physical examinations were served as the control group. The sputum bacteria before treatment and 6 months after treatment in the two groups were detected. The sputum negative conversion rate was recorded. The absorption of pulmonary lesions and the closure of tuberculosis cavity were examined. The immune cell function of T cell subset was detected again.Results: The peripheral blood CD3, CD4, CD8, and CD4/CD8 levels in the experiment group were significantly lower than those in the control group, especially for the active tuberculosis patients (P<0.01). The peripheral blood CD4/CD8 levels in the static tuberculosis patients was lower than that in the control group, but was significantly higher than that in active tuberculosis patients (P<0.01). The serum IL-1, IL-6, and TNF-α levels in the experiment group were significantly higher than those in the control group, especially for the active tuberculosis patients (P<0.01).Conclusions:The cell subsets and inflammatory cytokines play an important role in patients with pulmonary tuberculosis, whose dynamic change can effectively display the immune function and severity degree, which is of great value in estimating the condition and assessing the prognosis; therefore, it deserves to be further explored in the clinic.

  20. Ambient Levels of Air Pollution Induce Goblet-Cell Hyperplasia in Human Conjunctival Epithelium

    Science.gov (United States)

    Novaes, Priscila; do Nascimento Saldiva, Paulo Hilário; Kara-José, Newton; Macchione, Mariângela; Matsuda, Monique; Racca, Lourdes; Berra, Alejandro

    2007-01-01

    Background Ocular mucosa is exposed constantly to the external environment, and chronic exposure to air pollution may affect the ocular surface. Objective We assessed the effect of air pollution on the ocular surface by combining determinations of individual exposure and conjunctival impression cytology. Methods A panel study was conducted with 29 volunteers recruited in two locations with different pollution levels: São Paulo (n = 13) and Divinolândia (n = 16). We assessed mean individual levels of nitrogen dioxide (NO2) exposure for 7 days, using a passive sampler. Impression cytology samples were obtained from inferior tarsal conjunctiva. Comparisons between the two groups in terms of NO2 exposure and goblet-cell counts were performed using the Student t-test. Correlations between goblet-cells counts and corresponding individual NO2 exposure levels were determined using Spearman’s correlation. Results Individuals living in São Paulo received a significantly (p = 0.005) higher dose of NO2 (mean 32.47; SD 9.83) than those living in Divinolândia (mean 19.33; SD 5.24). There was a steady increase in goblet-cell counts, proportional to NO2 exposure (Spearman’s correlation = 0.566, p = 0.001), with a dose–response pattern. Conclusions A positive and significant association between exposure to air pollution and goblet-cell hyperplasia in human conjunctiva was detected. The combination of simple measurements of exposure and impression cytology was an effective and noninvasive approach for characterizing human response to ambient levels of air pollution. PMID:18087595

  1. ATP consumption of eukaryotic flagella measured at a single-cell level

    OpenAIRE

    Chen, Daniel T. N.; Heymann, Michael; Fraden, Seth; Nicastro, Daniela; Dogic, Zvonimir

    2015-01-01

    The motility of cilia and flagella is driven by thousands of dynein motors that hydrolyze adenosine triphosphate (ATP). Despite decades of genetic, biochemical, structural and biophysical studies, some aspects of ciliary motility remain elusive, such as the regulation of beating patterns and the energetic efficiency of these nanomachines. Here, we introduce an experimental method to measure ATP consumption of actively beating axonemes on a single-cell level. We encapsulated individual sea urc...

  2. Nutrient Starvation Decreases Cx43 Levels and Limits Intercellular Communication in Primary Bovine Corneal Endothelial Cells.

    Science.gov (United States)

    D'hondt, Catheleyne; Iyyathurai, Jegan; Welkenhuyzen, Kirsten; Himpens, Bernard; Leybaert, Luc; Bultynck, Geert

    2016-06-01

    Connexin (Cx) proteins form large conductance channels which function as regulators of communication between neighboring cells via gap junctions and/or hemichannels. Intercellular communication is essential to coordinate cellular responses in tissues and organs, thereby fulfilling an essential role in the spreading of signaling, survival and death processes. Connexin 43 (Cx43), a major connexin isoform in brain and heart, is rapidly turned over. Recent studies implicated that autophagy, a lysosomal degradation pathway induced upon nutrient starvation, mediates connexins, including Cx43, degradation. Here, we examined the impact of nutrient starvation on endogenous Cx43-protein levels and endogenous Cx43-driven intercellular communication in primary bovine corneal endothelial cells (BCECs). Hank's Balanced Salt Solution (HBSS) was used as a starvation condition that induces autophagic flux without impacting the survival of the BCECs. Nutrient starvation of BCECs caused a rapid decline in Cx43-protein levels, both as gap junctions and as hemichannels. The time course of the decline in Cx43-protein levels coincided with the time course of the decline in intercellular communication, assessed as intercellular Ca(2+)-wave propagation in BCECs exposed to a single-cell mechanical stimulus. The decline in Cx43-protein levels, both as gap junctions and as hemichannels, could be prevented by the addition of bafilomycin A1, a lysosomal inhibitor, during the complete nutrient starvation period. Consistent with this, bafilomycin A1 significantly alleviated the decrease in intercellular Ca(2+)-wave propagation. This study further underpins the importance of autophagy as an important degradation pathway for Cx43 proteins during periods of nutrient deprivation, thereby impacting the ability of cells to perform intercellular communication. PMID:26873723

  3. Effects of ionizing radiation. The measurable consequences at the level of the cell

    International Nuclear Information System (INIS)

    The exposition of an organism to ionizing radiations has measurable consequences at the DNA level, but also other cell structures. After the bringing to the fore of received doses indicators based on the observation of damaged chromosomes, the evolution of molecular biology techniques and their sensitivity have allowed to develop biological indicators of cellular effects. Without having the specificity of previous ones, these ones lead to better understand the radioinduced effects to reach diagnosis and prognosis factors of irradiation. (N.C.)

  4. Change of nucleolus characteristic of fish embryo cells under the influence of low-level radiation

    International Nuclear Information System (INIS)

    The nucleolus activity of fish embryo cells was stimulated by low-level radiation at a dose rate of 2-13 mGy/h. The size of nucleoli generally increased in embryos of Cyprinus carpio, whereas the number of nucleoli was greater in embryos of Carassius auratus gibelio. The higher the functional activity of nucleolus is, the more pronounced are changes in the characteristics. The size of single nucleolus at gastrulation is the most sensitive characteristic. 16 refs.; 1 tab

  5. Protective Pleiotropic Effect of Flavonoids on NAD + Levels in Endothelial Cells Exposed to High Glucose

    OpenAIRE

    Boesten, Daniëlle M. P. H. J.; von Ungern-Sternberg, Saskia N. I.; den Hartog, Gertjan J. M.; Aalt Bast

    2015-01-01

    NAD+ is important for oxidative metabolism by serving as an electron transporter. Hyperglycemia decreases NAD+ levels by activation of the polyol pathway and by overactivation of poly(ADP-ribose)-polymerase (PARP). We examined the protective role of three structurally related flavonoids (rutin, quercetin, and flavone) during high glucose conditions in an in vitro model using human umbilical vein endothelial cells (HUVECs). Additionally we assessed the ability of these flavonoids to inhibit al...

  6. Clinical significance of determination of serum leptin level, peripheral B cell number, T cell subset distribution type in patients with pregnancy induced hypertension

    International Nuclear Information System (INIS)

    Objective: To investigate the clinical significance of changes of serum leptin level, B cell number, T cell subsets in patients with pregnancy induced hypertension (PIH). Methods: Serum leptin levels (with RIA), peripheral B cell number, T cell subsets distribution type (with monoclonal anti-body technic) were determined in 32 patients with pregnaney induced hypertension (PIH) and 35 controls. Results: The serum leptin levels and B cell percentage were significantly higher in patients with PIH than those in controls (P<0.01), while the CD3, CD4 percentage and CD4/CD8 ratio were significantly lower (P<0.01). Conclusion: Determination of serum leptin levels and peripheral B cell number, T cell subsets distribution type might demonstrate immuno-disturbances in patients with pregnaney induced hypertension. (authors)

  7. High miR-196a levels promote the oncogenic phenotype of colorectal cancer cells

    Institute of Scientific and Technical Information of China (English)

    Carl Christoph Schimanski; Kirsten Frerichs; Fareed Rahman; Martin Berger; Hauke Lang; Peter R Galle; Markus Moehler; Ines Gockel

    2009-01-01

    AIM: To analyze the relevance of the microRNA miR- 196a for colorectal oncogenesis. METHODS: The impact of miR-196a on the restriction targets HoxA7, HoxB8, HoxC8 and HoxD8 was analyzed by reverse transcription polymerase chain reaction (RT-PCR) after transient transfection of SW480 cancer cells. The miR-196a transcription profile in colorectal cancer samples, mucosa samples and diverse cancer cell lines was quantified by RT-PCR. Transiently miR- 196a-transfected colorectal cancer cells were used for diverse functional assays in vitro and for a xenograft lung metastasis model in vivo. RESULTS: HoxA7, HoxB8, HoxC8 and HoxD8 were restricted by miR-196a in a dose-dependent and gene-specific manner. High levels of miR-196a activated the AKT signaling pathway as indicated by increased phosphorylation of AKT. In addition, high levels of miR-196a promoted cancer cell detachment, migration, invasion and chemosensitivity towards platin derivatives but did not impact on proliferation or apoptosis. Furthermore, miR-196a increased the development of lung metastases in mice after tail vein injection. CONCLUSION: miR-196a exerts a pro-oncogenic influence in colorectal cancer.

  8. Understanding epigenetic regulation: Tracking protein levels across multiple generations of cells

    Science.gov (United States)

    Rowat, A. C.; Weitz, D. A.

    2009-11-01

    Cells and organisms are remarkably robust: they alter the variety and levels of expressed genes and proteins in response to environmental stimuli, including temperature, chemicals, and the stiffness of their surroundings. Ultimately changes in gene and protein expression can result in a distinct phenotypic state, which in some cases is maintained over multiple generations; the ability to pass on a particular phenotypic state to progeny cells is critical for differentiation. Moreover, epigenetic regulation of phenotype is also thought to provide an evolutionary advantage for a population of cells adapting to a fluctuating environment on faster timescales than the occurrence of genetic mutations. However, simple methods to study patterns of gene and protein expression on multi-generational timescales are sparse. Here we describe a technique to study lineages of single cells over multiple generations using a microfluidic device; this reveals patterns of expression where protein levels are correlated across multiple generations. Such quantitative information of protein expression in the context of pedigree remains hidden when studying the population as an ensemble.

  9. 324 Radiochemical engineering cells and high level vault tanks mixed waste compliance status

    International Nuclear Information System (INIS)

    The 324 Building in the Hanford 300 Area contains Radiochemical Engineering Cells and High Level Vault tanks (the open-quotes REC/HLVclose quotes) for research and development activities involving radioactive materials. Radioactive mixed waste within this research installation, found primarily in B-Cell and three of the high level vault tanks, is subject to RCRA/DWR (open-quotes RCRAclose quotes) regulations for storage. This white paper provides a baseline RCRA compliance summary of MW management in the REC/HLV, based on best available knowledge. The REC/HLV compliance project, of which this paper is a part, is intended to achieve the highest degree of compliance practicable given the special technical difficulties of managing high activity radioactive materials, and to assure protection of human health and safety and the environment. The REC/HLV was constructed in 1965 to strict standards for the safe management of highly radioactive materials. Mixed waste in the REC/HLV consists of discarded tools and equipment, dried feed stock from nuclear waste melting experiments, contaminated particulate matter, and liquid feed stock from various experimental programs in the vault tanks. B-Cell contains most of these materials. Total radiological inventory in B-Cell is estimated at 3 MCi, about half of which is potentially open-quotes dispersibleclose quotes, that is, it is in small pieces or mobile particles. Most of the mixed waste currently in the REC/HLV was generated or introduced before mixed wastes were subjected to RCRA in 1987

  10. Correlation between the Sensitivity to TRAIL and the Expression Level of DR5 on the Surface of Tumor Cells

    Institute of Scientific and Technical Information of China (English)

    Yuanfang Ma; Jun Zhang; Yueping Zhao

    2005-01-01

    OBJECTIVE To investigate the correlation between the sensitivity to the tumor necrosis factor- related apoptosis inducing ligand (TRAIL) and the level of expression of the death receptor 5 (DR5) on the surface of tumor cells.METHODS Anti-DR5 mAbs were used to directly detect the level of expression of DR5 on the surface of tumor cells. Using a TRAIL apoptosis kit and flow cytometry, the sensitivity of the tumor cells to TRAIL-induced apoptosis was determined and the correlation between DR5 expression and sensitivity to TRAIL analyzed.RESULTS The expression level of DR5 on the surface of different tumor cells was as follows: 97.9% in U937 cells, 95.1% in Jurkat cells, 93.8% in SW480 cells, 86.2% in HCT116 cells, 64.2% in HL-60 cells, 46.6% in Hela cells and 13.1% in K562 cells. The TRAIL-induced apoptotic rate was 72.6% in U937 cells, 85.2% in Jurkat cells, 78.6% in SW480 cells, 70.2% in HCT116 cells,60.1% in HL-60 cells, 45.4% in Hela cells and 12.3% in K562 cells. Statistical analysis showed there was a significant positive correlation (r=0.997, P<0.001) between DR5 expression and sensitivity to TRAIL.CONCLUSION The sensitivity of tumor cells to TRAIL is related to the level of expression of DR5 on the surface of tumor cells. These results confirm the importance of DR5 expression for induction of apoptosis by TRAIL.

  11. Helicobacter pylori Infection in Association with Cell Proliferation,Apoptosis and Prostaglandin E2 Levels

    Institute of Scientific and Technical Information of China (English)

    PAN Kai-feng; ZHANG Yang; ZHANG Lian; MA Jun-ling; FENG Guo-shuang; ZHOU Tong; YOU Wei-cheng

    2007-01-01

    Objective: To evaluate the relationship between H. pylori infection with cell proliferation, apoptosis and PGE2 levels. Methods: A population-based study was conducted in Linqu, a high-risk area of gastric cancer in China. A total of 1523 subjects, aged 35-64, participating in a gastric cancer screening survey were investigated. H. pylori status were determined by 13C-urea breath test, expressions of Ki-67 were assessed by immunohistochemistry, apoptotic cells were detected by terminal deoxynucleotide transferase mediated dUTP nick end-labeling (TUNEL) method, and PGE2 levels were measured by enzyme immunoassay. Results: H. pylori infection was positively associated with cell proliferation activity. The mean and median percentage of Ki-67 labeling index (LI) in subjects with H. pylori positive were 14.1±10.3 and 12.0, significantly higher than those with H. pylori negative (-x±s: 8.4±7.0;median: 5.8;P<0.0001). Moreover, the prevalence rates of H. pylori infection showed a tendency to increase according to severity score of cell apoptosis (Ptrend <0.0001), from score 0 to 3, the percentage of H. pylori positivity increased from 67.5% to 96.7%. Furthermore, The mean and median of PGE2 concentration were 628.84±726.40 pg/mL and 411.33 pg/mL among subjects with H. pylori positive compared with 658.19±575.91pg/mL and 455.97 pg/mL among those with H. pylori negative (P=0.209). Conclusion: H. pylori infection was positively associated with increased cell proliferation and apoptosis activity, suggesting that H. pylori infection plays an important role in the gastric epithelial cell malignant transformation.

  12. Determination of plasma heparin level improves identification of systemic mast cell activation disease.

    Directory of Open Access Journals (Sweden)

    Milda Vysniauskaite

    Full Text Available Diagnosis of mast cell activation disease (MCAD, i.e. systemic mastocytosis (SM and idiopathic systemic mast cell activation syndrome (MCAS, usually requires demonstration of increased mast cell (MC mediator release. Since only a few MC mediators are currently established as biomarkers of MCAD, the sensitivity of plasma heparin level (pHL as an indicator of increased MC activation was compared with that of serum tryptase, chromogranin A and urinary N-methylhistamine levels in 257 MCAD patients. Basal pHL had a sensitivity of 41% in MCAS patients and 27% in SM patients. Non-pharmacologic stimulation of MC degranulation by obstruction of venous flow for 10 minutes increased the sensitivity of pHL in MCAS patients to 59% and in SM patients to 47%. In MCAS patients tryptase, chromogranin A, and N-methylhistamine levels exhibited low sensitivities (10%, 12%, and 22%, respectively, whereas sensitivities for SM were higher (73%, 63%, and 43%, respectively. Taken together, these data suggest pHL appears more sensitive than the other mediators for detecting systemic MC activity in patients with MCAS. The simple, brief venous occlusion test appears to be a useful indicator of the presence of pathologically irritable MCs, at least in the obstructed compartment of the body.

  13. Efficient handling of high-level radioactive cell waste in a vitrification facility analytical laboratory

    International Nuclear Information System (INIS)

    The Savannah River Site''s (SRS) Defense Waste Processing Facility (DWPF) near Aiken, South Carolina, is the world''s largest and the United State''s first high level waste vitrification facility. For the past 1.5 years, DWPF has been vitrifying high level radioactive liquid waste left over from the Cold War. The vitrification process involves the stabilization of high level radioactive liquid waste into borosilicate glass. The glass is contained in stainless steel canisters. DWPF has filled more than 200 canisters 3.05 meters (10 feet) long and 0.61 meters (2 foot) diameter. Since operations began at DWPF in March of 1996, high level radioactive solid waste continues to be generated due to operating the facility''s analytical laboratory. The waste is referred to as cell waste and is routinely removed from the analytical laboratories. Through facility design, engineering controls, and administrative controls, DWPF has established efficient methods of handling the high level waste generated in its laboratory facility. These methods have resulted in the prevention of undue radiation exposure, wasted man-hours, expenses due to waste disposal, and the spread of contamination. This level of efficiency was not reached overnight, but it involved the collaboration of Radiological Control Operations and Laboratory personnel working together to devise methods that best benefited the facility. This paper discusses the methods that have been incorporated at DWPF for the handling of cell waste. The objective of this paper is to provide insight to good radiological and safety practices that were incorporated to handle high level radioactive waste in a laboratory setting

  14. Alterations of T cell activation signalling and cytokine production by postmenopausal estrogen levels

    Directory of Open Access Journals (Sweden)

    Taylor Douglas D

    2009-03-01

    Full Text Available Abstract Background Immunosenescence is an age-associated disorder occurring primarily in T cell compartments, including altered subset composition, functions, and activation. In women, evidence implicates diminished estrogen in the postmenopausal period as a contributing factor to diminished T cell responsiveness. Since hypoestrogenism is present in postmenopausal women, our objective focused on whether T cell activation, defined as signalling molecule expressions and activation, and function, identified as IL-2 production, were affected by low estrogen. Methods Using Jurkat 6.1 T cells, consequences of 4 pg/ml (corresponding to postmenopausal levels or 40 pg/ml (premenopausal levels of estradiol (E2 were analyzed on signalling proteins, CD3-zeta, JAK2, and JAK3, determined by Western immunoblotting. These consequences were correlated with corresponding gene expressions, quantified by real time-polymerase chain reaction. Tyrosine phosphorylation of CD3-zeta was defined by immunoprecipitation and western immunoblotting following activation by T cell receptor (TcR cross-linking. CD3-zeta expression and modulation was also confirmed in T cells from pre- and postmenopausal women. To assess functional consequences, IL-2 production, induced by PMA and ionomycin, was determined using enzyme-linked immunosorbent spot assay (ELISpot. Results At 40 pg/ml E2, the level of signalling protein CD3-zeta was elevated 1.57-fold, compared with cells exposed to 4 pg/ml E2. The CD3-zeta proteins also exhibited altered levels of activation-induced phosphorylation in the presence of 40 pg/ml E2 versus 4 pg/ml: 23 kD phosphorylated form increased 2.64-fold and the 21 kD form was elevated 2.95-fold. Examination of kinases associated with activation signalling also demonstrated that, in the presence of 40 pg/ml E2, JAK2 protein expression was increased 1.64-fold (p 2 (2.39, 2.01, and 2.21 fold, respectively versus 4 pg/ml. These findings were confirmed in vivo, since T

  15. Decreased circulating endothelial progenitor cell levels and function in patients with nonalcoholic fatty liver disease.

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    Chia-Hung Chiang

    Full Text Available OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD is associated with advanced atherosclerosis and a higher risk of cardiovascular disease. Increasing evidence suggests that injured endothelial monolayer is regenerated by circulating bone marrow derived-endothelial progenitor cells (EPCs, and levels of circulating EPCs reflect vascular repair capacity. However, the relation between NAFLD and EPC remains unclear. Here, we tested the hypothesis that patients with nonalcoholic fatty liver disease (NAFLD might have decreased endothelial progenitor cell (EPC levels and attenuated EPC function. METHODS AND RESULTS: A total of 312 consecutive patients undergoing elective coronary angiography because of suspected coronary artery disease were screened and received examinations of abdominal ultrasonography between July 2009 and November 2010. Finally, 34 patients with an ultrasonographic diagnosis of NAFLD, and 68 age- and sex-matched controls without NAFLD were enrolled. Flow cytometry with quantification of EPC markers (defined as CD34(+, CD34(+KDR(+, and CD34(+KDR(+CD133(+ in peripheral blood samples was used to assess circulating EPC numbers. The adhesive function, and migration, and tube formation capacities of EPCs were also determined in NAFLD patients and controls. Patients with NAFLD had a significantly higher incidence of metabolic syndrome, previous myocardial infarction, hyperuricemia, and higher waist circumference, body mass index, fasting glucose and triglyceride levels. In addition, patients with NAFLD had significantly decreased circulating EPC levels (all P<0.05, attenuated EPC functions, and enhanced systemic inflammation compared to controls. Multivariate logistic regression analysis showed that circulating EPC level (CD34(+KDR(+ [cells/10(5 events] was an independent reverse predictor of NAFLD (Odds ratio: 0.78; 95% confidence interval: 0.69-0.89, P<0.001. CONCLUSIONS: NAFLD patients have decreased circulating EPC numbers and

  16. Fatigue, serum cytokine levels, and blood cell counts during radiotherapy of patients with breast cancer

    International Nuclear Information System (INIS)

    Purpose: To assess the level of fatigue during the course of adjuvant radiotherapy (RT) of breast cancer patients and its relation to anxiety, depression, serum cytokines, and blood count levels. Methods and Materials: Forty-one patients who received adjuvant RT after breast-conserving surgery were prospectively studied. All patients underwent RT without concomitant chemotherapy. Patients rated their fatigue with two standardized self-assessment instruments, the Fatigue Assessment Questionnaire and a visual analog scale on fatigue intensity, before RT, during weeks 1-5 of RT, and 2 months after RT completion. In addition, the anxiety and depression levels were assessed with the Hospital Anxiety and Depression Scale. A differential blood cell count and the serum levels of the cytokines interleukin (IL)-1β, IL-6, and tumor necrosis factor-α were determined in parallel to the fatigue assessments. Results: Fatigue intensity as assessed with the visual analog scale increased (p<0.001) until treatment week 4 and remained elevated until week 5. Two months after RT, the values had fallen to the pretreatment levels. Fatigue measured with the Fatigue Assessment Questionnaire did not increase significantly during treatment, but the subscores on physical (p=0.035) and cognitive (p=0.015) fatigue were elevated during treatment weeks 4 and 5. Affective fatigue did not change significantly. Anxiety, as rated with the Hospital Anxiety and Depression Scale, declined during RT (p=0.002), but the Hospital Anxiety and Depression Scale depression score did not change significantly. IL-1β, IL-6, and tumor necrosis factor-α levels did not change during therapy and did not correlate with fatigue. Peripheral blood cell levels declined significantly during therapy and were still low 2 months after treatment. Until treatment week 5, lymphocytes were reduced to almost 50% of their initial values. Hemoglobin levels did not correlate with fatigue. Conclusions: We observed an increase in

  17. Clonal-level responses of functionally distinct hematopoietic stem cells to trophic factors.

    Science.gov (United States)

    Mallaney, Cates; Kothari, Alok; Martens, Andrew; Challen, Grant A

    2014-04-01

    Recent findings from several groups have identified distinct classes of hematopoietic stem cells (HSCs) in the bone marrow, each with inherent functional biases in terms of their differentiation, self-renewal, proliferation, and lifespan. It has previously been demonstrated that myeloid- and lymphoid-biased HSCs can be prospectively enriched based on their degree of Hoechst dye efflux. In the present study, we used differential Hoechst efflux to enrich lineage-biased HSC subtypes and analyzed their functional potentials. Despite similar outputs in vitro, bone marrow transplantation assays revealed contrasting lineage differentiation in vivo. To stratify the molecular differences underlying these contrasting functional potentials at the clonal level, single-cell gene expression analysis was performed using the Fluidigm BioMark system and revealed dynamic expression of genes including Meis1, CEBP/α, Sfpi1, and Dnmt3a. Finally, single-cell gene expression analysis was used to unravel the opposing proliferative responses of lineage-biased HSCs to the growth factor TGF-β1, revealing a potential role for the cell cycle inhibitor Cdkn1c as molecular mediator. This work lends further credence to the concept of HSC heterogeneity, and it presents unprecedented molecular resolution of the HSC response to trophic factors using single-cell gene expression analysis. PMID:24373928

  18. Interleukin-18 alters protein expressions of neurodegenerative diseases-linked proteins in human SH-SY5Y neuron-like cells

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    Elina M Sutinen

    2014-08-01

    Full Text Available Chronic inflammation and oxidative stress (OS are present in Alzheimer´s disease (AD brains in addition to neuronal loss, Amyloid-β (Aβ plaques and hyperphosphorylated tau-protein neurofibrillary tangles. Previously we showed that levels of the pro-inflammatory cytokine, interleukin-18 (IL-18, are elevated in post-mortem AD brains. IL-18 can modulate the tau kinases, Cdk5 and GSK3β, as well as Aβ-production. IL-18 levels are also increased in AD risk diseases, including type-2 diabetes and obesity. Here, we explored other IL-18 regulated proteins in neuron-like SH-SY5Y cells. Differentiated SH-SY5Y cells, incubated with IL-18 for 24, 48 or 72h, were analyzed by two-dimensional gel electrophoresis (2D-DIGE. Specific altered protein spots were chosen and identified with mass spectrometry and verified by western immunoblotting. IL-18 had time-dependent effects on the SH-SY5Y proteome, modulating numerous protein levels/modifications. We concentrated on those related to OS (DDAH2, peroxiredoxins 2, 3 and 6, DJ-1, BLVRA, Aβ-degradation (MMP14, TIMP2, Aβ-aggregation (Septin-2 and modifications of axon growth and guidance associated, collapsing response mediator protein 2 (CRMP2. IL-18 significantly increased antioxidative enzymes, indicative of OS, and altered levels of glycolytic α- and γ-enolase and multifunctional 14-3-3γ and -ε, commonly affected in neurodegenerative diseases. MMP14, TIMP2, α-enolase and 14-3-3ε, indirectly involved in Aβ metabolism, as well as Septin-2 showed changes that increase Aβ levels. Increased 14-3-3γ may contribute to GSK3β driven tau hyperphosphorylation and CRMP2 Thr514 and Ser522 phosphorylation with the Thr555-site, a target for Rho kinase, showing time-dependent changes. IL-18 also increased caspase-1 levels and vacuolization of the cells. Although our SH-SY5Y cells were not aged, as neurons in AD, our work suggests that heightened or prolonged IL-18 levels can drive protein changes of known

  19. Radiation-induced alterations of histone post-translational modification levels in lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Radiation-induced alterations in posttranslational histone modifications (PTMs) may affect the cellular response to radiation damage in the DNA. If not reverted appropriately, altered PTM patterns may cause long-term alterations in gene expression regulation and thus lead to cancer. It is therefore important to characterize radiation-induced alterations in PTM patterns and the factors affecting them. A lymphoblastoid cell line established from a normal donor was used to screen for alterations in methylation levels at H3K4, H3K9, H3K27, and H4K20, as well as acetylation at H3K9, H3K56, H4K5, and H4K16, by quantitative Western Blot analysis at 15 min, 1 h and 24 h after irradiation with 2 Gy and 10 Gy. The variability of alterations in acetylation marks was in addition investigated in a panel of lymphoblastoid cell lines with differing radiosensitivity established from lung cancer patients. The screening procedure demonstrated consistent hypomethylation at H3K4me3 and hypoacetylation at all acetylation marks tested. In the panel of lymphoblastoid cell lines, however, a high degree of inter-individual variability became apparent. Radiosensitive cell lines showed more pronounced and longer lasting H4K16 hypoacetylation than radioresistant lines, which correlates with higher levels of residual γ-H2AX foci after 24 h. So far, the factors affecting extent and duration of radiation-induced histone alterations are poorly defined. The present work hints at a high degree of inter-individual variability and a potential correlation of DNA damage repair capacity and alterations in PTM levels

  20. Motor axon synapses on renshaw cells contain higher levels of aspartate than glutamate.

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    Dannette S Richards

    Full Text Available Motoneuron synapses on spinal cord interneurons known as Renshaw cells activate nicotinic, AMPA and NMDA receptors consistent with co-release of acetylcholine and excitatory amino acids (EAA. However, whether these synapses express vesicular glutamate transporters (VGLUTs capable of accumulating glutamate into synaptic vesicles is controversial. An alternative possibility is that these synapses release other EAAs, like aspartate, not dependent on VGLUTs. To clarify the exact EAA concentrated at motor axon synapses we performed a quantitative postembedding colloidal gold immunoelectron analysis for aspartate and glutamate on motor axon synapses (identified by immunoreactivity to the vesicular acetylcholine transporter; VAChT contacting calbindin-immunoreactive (-IR Renshaw cell dendrites. The results show that 71% to 80% of motor axon synaptic boutons on Renshaw cells contained aspartate immunolabeling two standard deviations above average neuropil labeling. Moreover, VAChT-IR synapses on Renshaw cells contained, on average, aspartate immunolabeling at 2.5 to 2.8 times above the average neuropil level. In contrast, glutamate enrichment was lower; 21% to 44% of VAChT-IR synapses showed glutamate-IR two standard deviations above average neuropil labeling and average glutamate immunogold density was 1.7 to 2.0 times the neuropil level. The results were not influenced by antibody affinities because glutamate antibodies detected glutamate-enriched brain homogenates more efficiently than aspartate antibodies detecting aspartate-enriched brain homogenates. Furthermore, synaptic boutons with ultrastructural features of Type I excitatory synapses were always labeled by glutamate antibodies at higher density than motor axon synapses. We conclude that motor axon synapses co-express aspartate and glutamate, but aspartate is concentrated at higher levels than glutamate.

  1. Conceptual designs for utility load-leveling battery with Li/FeS cells

    Energy Technology Data Exchange (ETDEWEB)

    Zivi, S. M.; Kacinskas, H.; Pollack, I.; Chilenskas, A. A.; Grieve, W.; McFarland, B. L.; Sudar, S.

    1980-07-01

    In 1978, a conceptual design of a 100 MW-h load-leveling battery system having Li alloy/FeS cells was developed as a result of a joint effort between ANL and Rockwell International. In this conceptual design, the submodule, which was the basic replaceable unit for the system, had a capacity of 240 kW-h and consisted of ninety-six 2.5 kW-h cells. However, a study by Rockwell indicated that the cost for battery hardware, $60 to 80/kW-h (cells and converters not included), was too high. Most of this cost was contributed by the submodule structure and the charge equalization scheme, which was the same as that developed for electric-vehicle batteries. In 1979, subsequent efforts were concentrated on lowering these hardware costs and resulted in the development of three modified designs, which are presented in this report. The first, developed at ANL, consisted of a 30 kW-h cell/submodule and the electric-vehicle equalization scheme. The hardware cost for this modified design was quite low, about $25/kW-h; however, this design was eventually rejected owing to the apparent impracticality of such a large cell. The two other modified designs had more conservative cell designs. One of them, developed at ANL, consisted of a 120 kW-h submodule consisting of one hundred 1.2 kW-h cells; the other, developed at Rockwell, consisted of a 1020 kW-h submodule consisting of four hundred and eight 2.5 kW-h cells. For both of these designs, an alternative method of equalization, in which fixed resistance shunts are used on each cell, was proposed; this equalization method adds little equipment cost to the system and only sacrifices about 4% of the coulombic and energy efficiencies. The cost of battery hardware for these two designs was estimated to be acceptable, about $22 to 60/kW-h. Some questions remain on the assumed capabilities of the cells and the feasibility of the battery hardware.

  2. Diethylstilbestrol alters the morphology and calcium levels of growth cones of PC12 cells in vitro.

    Science.gov (United States)

    Janevski, J; Choh, V; Stopper, H; Schiffmann, D; De Boni, U

    1993-01-01

    Diethylstilbestrol (DES) is a synthetic estrogen with carcinogenic properties. DES is known to alter cytoskeletal components, including the organization of actin stress fibres in C6 rat glioma cells. In a test of the hypothesis that DES disrupts actin filaments of growth cones in neuron-like cells, DES-induced changes in filopodial lengths were quantified in rat pheochromocytoma (PC12) cells in vitro. DES significantly altered growth cone morphology, with collapse of growth cone filopodia and neurite retraction invariably occurring at a concentration of 10 microM. At 5 microM DES, transient reductions in total filopodial lengths occurred. At DES concentrations of 0.1 nM and 1 nM, reductions in total filopodial lengths occurred in a fraction of growth cones. Evidence exists which shows that growth cone activity and morphology are intimately linked to levels of intracellular, free calcium and that DES increases such levels. Measurements of free intracellular calcium levels by fluorescence microscopy, at times concurrent with the DES-induced reduction in total filopodial lengths, showed that calcium levels were indeed significantly increased by 10 microM DES. Labelling of filamentous actin (f-actin) with FITC-phalloidin showed that the f-actin distribution in growth cones exposed to DES could not be differentiated from the distribution found in spontaneously retracting growth cones. Together with evidence which showed that growth cone motility was not affected, the results are taken to indicate that DES, rather than acting directly on the cytoskeleton, exerts its effects indirectly, by a calcium-induced destabilization of actin filaments in the growth cone. PMID:8164893

  3. Constructing disease-specific gene networks using pair-wise relevance metric: Application to colon cancer identifies interleukin 8, desmin and enolase 1 as the central elements

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    Jiang Wei

    2008-08-01

    Full Text Available Abstract Background With the advance of large-scale omics technologies, it is now feasible to reversely engineer the underlying genetic networks that describe the complex interplays of molecular elements that lead to complex diseases. Current networking approaches are mainly focusing on building genetic networks at large without probing the interaction mechanisms specific to a physiological or disease condition. The aim of this study was thus to develop such a novel networking approach based on the relevance concept, which is ideal to reveal integrative effects of multiple genes in the underlying genetic circuit for complex diseases. Results The approach started with identification of multiple disease pathways, called a gene forest, in which the genes extracted from the decision forest constructed by supervised learning of the genome-wide transcriptional profiles for patients and normal samples. Based on the newly identified disease mechanisms, a novel pair-wise relevance metric, adjusted frequency value, was used to define the degree of genetic relationship between two molecular determinants. We applied the proposed method to analyze a publicly available microarray dataset for colon cancer. The results demonstrated that the colon cancer-specific gene network captured the most important genetic interactions in several cellular processes, such as proliferation, apoptosis, differentiation, mitogenesis and immunity, which are known to be pivotal for tumourigenesis. Further analysis of the topological architecture of the network identified three known hub cancer genes [interleukin 8 (IL8 (p ≈ 0, desmin (DES (p = 2.71 × 10-6 and enolase 1 (ENO1 (p = 4.19 × 10-5], while two novel hub genes [RNA binding motif protein 9 (RBM9 (p = 1.50 × 10-4 and ribosomal protein L30 (RPL30 (p = 1.50 × 10-4] may define new central elements in the gene network specific to colon cancer. Gene Ontology (GO based analysis of the colon cancer-specific gene network and

  4. Study on serum TNF-α level, B-cell count and T-cell subsets distribution in peripheral blood in patients with rheumatoid arthritis

    International Nuclear Information System (INIS)

    Objective: To study the changes of serum TNF-α levels, B-cell count and T-cell subsets distribution in peripheral blood in patients with rheumatoid arthritis. Methods: Serum TNF-α levels (with RIA), B cell as well as T cell subsets distribution type (with monoclonal antibody technique) were examined in 37 patients with rheumatoid arthritis and 30 controls. Results Serum TNF-α levels and B lymphocytes count were significantly higher in the patients than those in controls (P3, CD4 and CD4/CD8 were obviously lower (P<0.01). Conclusion: Rheumatoid arthritis is an autoimmune disease with abnormal immunoregulation. (authors)

  5. Radiosensitizing Effects of Ectopic miR-101 on Non–Small-Cell Lung Cancer Cells Depend on the Endogenous miR-101 Level

    International Nuclear Information System (INIS)

    Purpose: Previously, we showed that ectopic miR-101 could sensitize human tumor cells to radiation by targeting ATM and DNA-PK catalytic subunit (DNA-PKcs) to inhibit DNA repair, as the endogenous miR-101 levels are low in tumors in general. However, the heterogeneity of human cancers may result in an exception. The purpose of this study was to test the hypothesis that a few tumor cell lines with a high level of endogenous miR-101 would prove less response to ectopic miR-101. Methods and Materials: Fourteeen non–small-cell lung cancer (NSCLC) cell lines and one immortalized non-malignant lung epithelial cell line (NL20) were used for comparing endogenous miR-101 levels by real-time reverse transcription–polymerase chain reaction. Based on the different miR-101 levels, four cell lines with different miR-101 levels were chosen for transfection with a green fluorescent protein–lentiviral plasmid encoding miR-101. The target protein levels were measured by using Western blotting. The radiosensitizing effects of ectopic miR-101 on these NSCLC cell lines were determined by a clonogenic assay and xenograft mouse model. Results: The endogenous miR-101 level was similar or lower in 13 NSCLC cell lines but was 11-fold higher in one cell line (H157) than in NL20 cells. Although ectopic miR-101 efficiently decreased the ATM and DNA-PKcs levels and increased the radiosensitization level in H1299, H1975, and A549 cells, it did not change the levels of the miR-101 targets or radiosensitivity in H157 cells. Similar results were observed in xenograft mice. Conclusions: A small number of NSCLC cell lines could have a high level of endogenous miR-101. The ectopic miR-101 was able to radiosensitize most NSCLC cells, except for the NSCLC cell lines that had a much higher endogenous miR-101 level. These results suggest that when we choose one miRNA as a therapeutic tool, the endogenous level of the miRNA in each tumor should be considered.

  6. Radiosensitizing Effects of Ectopic miR-101 on Non-Small-Cell Lung Cancer Cells Depend on the Endogenous miR-101 Level

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Susie; Wang Hongyan; Ng, Wooi Loon; Curran, Walter J. [Department of Radiation Oncology, School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, GA (United States); Wang Ya, E-mail: ywang94@emory.edu [Department of Radiation Oncology, School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, GA (United States)

    2011-12-01

    Purpose: Previously, we showed that ectopic miR-101 could sensitize human tumor cells to radiation by targeting ATM and DNA-PK catalytic subunit (DNA-PKcs) to inhibit DNA repair, as the endogenous miR-101 levels are low in tumors in general. However, the heterogeneity of human cancers may result in an exception. The purpose of this study was to test the hypothesis that a few tumor cell lines with a high level of endogenous miR-101 would prove less response to ectopic miR-101. Methods and Materials: Fourteeen non-small-cell lung cancer (NSCLC) cell lines and one immortalized non-malignant lung epithelial cell line (NL20) were used for comparing endogenous miR-101 levels by real-time reverse transcription-polymerase chain reaction. Based on the different miR-101 levels, four cell lines with different miR-101 levels were chosen for transfection with a green fluorescent protein-lentiviral plasmid encoding miR-101. The target protein levels were measured by using Western blotting. The radiosensitizing effects of ectopic miR-101 on these NSCLC cell lines were determined by a clonogenic assay and xenograft mouse model. Results: The endogenous miR-101 level was similar or lower in 13 NSCLC cell lines but was 11-fold higher in one cell line (H157) than in NL20 cells. Although ectopic miR-101 efficiently decreased the ATM and DNA-PKcs levels and increased the radiosensitization level in H1299, H1975, and A549 cells, it did not change the levels of the miR-101 targets or radiosensitivity in H157 cells. Similar results were observed in xenograft mice. Conclusions: A small number of NSCLC cell lines could have a high level of endogenous miR-101. The ectopic miR-101 was able to radiosensitize most NSCLC cells, except for the NSCLC cell lines that had a much higher endogenous miR-101 level. These results suggest that when we choose one miRNA as a therapeutic tool, the endogenous level of the miRNA in each tumor should be considered.

  7. Influence of free radicals generated by gamma irradiation on the trehalose levels in Saccharomyces boulardii leaven cells

    International Nuclear Information System (INIS)

    Lyophilized cells of yeast Saccharomyces boulardii were submit to the gamma irradiation (60 Cobalt). It was noted that the yeast cells surviving to gamma irradiation had more trehalose than control cells. The incubation of cells with H202 induces the synthesis of trehalose but the exposition to CdCl2 induces the accumulation of trehalose only in low concentrations. Apparently there is a positive correlation between of trehalose level and gamma irradiation resistance of S. boulardii cells. It was not observed any correlation with the level of trehalose determined and the oxidative stress tested. (author)

  8. Non increased neuron-specific enolase concentration in cerebrospinal fluid during first febrile seizures and a year follow-up in pediatric patients No incrementos en la concentración de enolasa específica de neurona en el líquido cefalorraquídeo durante el primer ataque febril y al año en pacientes pediátricos

    OpenAIRE

    Dorta-Contreras, Alberto J; EDITH TABÍO-VALDÉS; ALINA TABÍO-VALDÉS; CARIDAD DELGADO-FERNÁNDEZ; HANSOTTO REIBER

    1998-01-01

    Febrile seizures are the commonest acute neurological disorder of early childhood. Studies suggested that febrile seizures are previous acute events from a more serious neurological problem. Due to neuron-specific enolase is generally accepted as a marker for neuropathological processes in the brain, 16 pediatric patients were studied during their first seizures and a year after it. Neuron-specific enolase in cerebrospinal fluid and blood were analysed by an immune enzyme assay. Non pathologi...

  9. Human peripheral blood mononuclear cells exhibit heterogeneous CD52 expression levels and show differential sensitivity to alemtuzumab mediated cytolysis.

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    Sambasiva P Rao

    Full Text Available Alemtuzumab is a monoclonal antibody that targets cell surface CD52 and is effective in depleting lymphocytes by cytolytic effects in vivo. Although the cytolytic effects of alemtuzumab are dependent on the density of CD52 antigen on cells, there is scant information regarding the expression levels of CD52 on different cell types. In this study, CD52 expression was assessed on phenotypically distinct subsets of lymphoid and myeloid cells in peripheral blood mononuclear cells (PBMCs from normal donors. Results demonstrate that subsets of PBMCs express differing levels of CD52. Quantitative analysis showed that memory B cells and myeloid dendritic cells (mDCs display the highest number while natural killer (NK cells, plasmacytoid dendritic cells (pDCs and basophils have the lowest number of CD52 molecules per cell amongst lymphoid and myeloid cell populations respectively. Results of complement dependent cytolysis (CDC studies indicated that alemtuzumab mediated profound cytolytic effects on B and T cells with minimal effect on NK cells, basophils and pDCs, correlating with the density of CD52 on these cells. Interestingly, despite high CD52 levels, mDCs and monocytes were less susceptible to alemtuzumab-mediated CDC indicating that antigen density alone does not define susceptibility. Additional studies indicated that higher expression levels of complement inhibitory proteins (CIPs on these cells partially contributes to their resistance to alemtuzumab mediated CDC. These results indicate that alemtuzumab is most effective in depleting cells of the adaptive immune system while leaving innate immune cells relatively intact.

  10. Effects of low-level laser therapy on stem cells from human exfoliated deciduous teeth

    Science.gov (United States)

    FERNANDES, Ana Paula; JUNQUEIRA, Marina de Azevedo; MARQUES, Nádia Carolina Teixeira; MACHADO, Maria Aparecida Andrade Moreira; SANTOS, Carlos Ferreira; OLIVEIRA, Thais Marchini; SAKAI, Vivien Thiemy

    2016-01-01

    ABSTRACT Low-Level Laser Therapy stimulates the proliferation of a variety of types of cells. However, very little is known about its effect on stem cells from human exfoliated deciduous teeth (SHED). Objective This study aimed to evaluate the influence of different laser therapy energy densities on SHED viability and proliferation. Material and Methods SHED were irradiated according to the groups: I (1.2 J/cm2 - 0.5 mW – 10 s), II (2.5 J/cm2 – 10 mW – 10 s), III (3.7 J/cm2 – 15 mW – 10 s), IV (5.0 J/cm2 – 20 mW – 10 s), V (6.2 J/cm2 – 25 mW – 10 s), and VI (not irradiated – control group). Cell viability was assessed 6 and 24 h after irradiation measuring the mitochondrial activity and using the Crystal Violet assay. Cell proliferation was assessed after 24, 48, and 72 h of irradiation by SRB assay. Results MTT assay demonstrated differences from 6 to 24 hours after irradiation. After 24 h, groups I and IV showed higher absorbance values than those of control group. Crystal Violet assay showed statistically differences in the absorbance rate from 6 to 24 h after irradiation for groups III and VI. At 24 h after irradiation, Group III absorbance rate was greater than that of groups I, II, and IV. Group VI absorbance rate was greater than that of groups I and IV. SRB assay showed that the group I had higher rates than those of groups II, III, V, and VI, at 24 h after irradiation. After 48 h, group I exhibited the greatest cell proliferation rate followed by groups III, V, and VI. After 72 h, group III exhibited the lowest cell proliferation rate than those of groups II, IV, and V. Conclusions The Low-Level Laser Therapy energy densities used in this study did not cause loss of cell viability and stimulated SHED proliferation within the parameters described in this study. PMID:27556203

  11. Dengue viral RNA levels in peripheral blood mononuclear cells are associated with disease severity and preexisting dengue immune status.

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    Anon Srikiatkhachorn

    Full Text Available BACKGROUND: Infection with dengue viruses (DENV causes a wide range of manifestations from asymptomatic infection to a febrile illness called dengue fever (DF, to dengue hemorrhagic fever (DHF. The in vivo targets of DENV and the relation between the viral burden in these cells and disease severity are not known. METHOD: The levels of positive and negative strand viral RNA in peripheral blood monocytes, T/NK cells, and B cells and in plasma of DF and DHF cases were measured by quantitative RT-PCR. RESULTS: Positive strand viral RNA was detected in monocytes, T/NK cells and B cells with the highest amounts found in B cells. Viral RNA levels in CD14+ cells and plasma were significantly higher in DHF compared to DF, and in cases with a secondary infection compared to those undergoing a primary infection. The distribution of viral RNA among cell subpopulations was similar in DF and DHF cases. Small amounts of negative strand RNA were found in a few cases only. The severity of plasma leakage correlated with viral RNA levels in plasma and in CD14+ cells. CONCLUSIONS: B cells were the principal cells containing DENV RNA in peripheral blood, but overall there was little active DENV RNA replication detectable in peripheral blood mononuclear cells (PBMC. Secondary infection and DHF were associated with higher viral burden in PBMC populations, especially CD14+ monocytes, suggesting that viral infection of these cells may be involved in disease pathogenesis.

  12. Sputum and BAL Clara cell secretory protein and surfactant protein D levels in asthma.

    Science.gov (United States)

    Emmanouil, P; Loukides, S; Kostikas, K; Papatheodorou, G; Papaporfyriou, A; Hillas, G; Vamvakaris, I; Triggidou, R; Katafigiotis, P; Kokkini, A; Papiris, S; Koulouris, N; Bakakos, P

    2015-06-01

    Clara cell secretory protein (CC16) is associated with Th2 modulation. Surfactant protein D (SPD) plays an important role in surfactant homeostasis and eosinophil chemotaxis. We measured CC16 and SPD in sputum supernatants of 84 asthmatic patients and 12 healthy controls. In 22 asthmatics, we additionally measured CC16 and SPD levels in BAL and assessed smooth muscle area (SMA), reticular basement membrane (RBM) thickness, and epithelial detachment (ED) in bronchial biopsies. Induced sputum CC16 and SPD were significantly higher in patients with severe asthma (SRA) compared to mild-moderate and healthy controls. BAL CC16 and SPD levels were also higher in SRA compared to mild-moderate asthma. CC16 BAL levels correlated with ED, while SPD BAL levels correlated with SMA and RBM. Severity represented a significant covariate for these associations. CC16 and SPD levels are upregulated in SRA and correlate with remodeling indices, suggesting a possible role of these biomarkers in the remodeling process. PMID:25728058

  13. Caffeic Acid Phenethyl Ester Regulates PPAR's Levels in Stem Cells-Derived Adipocytes.

    Science.gov (United States)

    Vanella, Luca; Tibullo, Daniele; Godos, Justyna; Pluchinotta, Francesca Romana; Di Giacomo, Claudia; Sorrenti, Valeria; Acquaviva, Rosaria; Russo, Alessandra; Li Volti, Giovanni; Barbagallo, Ignazio

    2016-01-01

    Hypertrophic obesity inhibits activation of peroxisome proliferators-activated receptor gamma (PPARγ), considered the key mediator of the fully differentiated and insulin sensitive adipocyte phenotype. We examined the effects of Caffeic Acid Phenethyl Ester (Cape), isolated from propolis, a honeybee hive product, on Adipose Stem Cells (ASCs) differentiation to the adipocyte lineage. Finally we tested the effects of Cape on insulin-resistant adipocytes. Quantification of Oil Red O-stained cells showed that lipid droplets decreased following Cape treatment as well as radical oxygen species formation. Additionally, exposure of ASC to high glucose levels decreased adiponectin and increased proinflammatory cytokines mRNA levels, which were reversed by Cape-mediated increase of insulin sensitivity. Cape treatment resulted in decreased triglycerides synthesis and increased beta-oxidation. Exposure of ASCs to Lipopolysaccharide (LPS) induced a reduction of PPARγ, an increase of IL-6 levels associated with a well-known stimulation of lipolysis; Cape partially attenuated the LPS-mediated effects. These observations reveal the main role of PPARγ in the adipocyte function and during ASC differentiation. As there is now substantial interest in functional food and nutraceutical products, the observed therapeutic value of Cape in insulin-resistance related diseases should be taken into consideration. PMID:26904104

  14. The Effect of Low Level Laser Irradiation on Human Embryonic Stem Cells

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    Hossein Baharvand

    2005-01-01

    Full Text Available Introduction: Different effects of low level laser irradiation (LLLI on various cell types have already been demonstrated. However, its effects on embryonic stem cells have not yet been shown. The present study evaluates the morphological and immunocytochemical effects of LLLI on human embryonic stem cell (hESC colonies. Material and Methods: Equal-sized pieces of hESC line (Royan H1 were irradiated with a single dose of 830-nm Ga-Al-As diode laser (3, 5, and 8 jcm-2, 30mW and cultured on mouse embryonic fibroblasts. The morphology of the colonies was evaluated qualitatively by observation under an inverted microscope (grades A, B, C, and D exhibited 0-30%, 30-50%, 50-80%, and 80-100% differentiation, respectively. The stemness area was assessed by expression of surface antigens using anti-Tra-1-60 and anti-Tra-1-81. Results: Our data demonstrated a dose-dependent stimulatory effect of LLLI on hESC differentiation. Two doses of 5 and 8jcm-2 induced statistically significant differentiation (grades C and D. Conclusions: These data showed that LLLI influenced hESC differentiation, which might be used for cell therapy after transplantation

  15. Development of a unit cell model for interim performance assessment of vitrified low level waste disposal

    International Nuclear Information System (INIS)

    The unit cell modeling approach has been developed and used in analysis of some design options for a vitrified low level waste disposal facility. The unit cell modeling approach is likely to be useful in interim performance assessment for the facility. The present unit cell model will probably need to be refitted in terms of some model parameters for the latter purpose. Two present disposal facility concepts differ in the length of a capillary barrier proposed to limit effective recharge through the top of the facility. Results of the study summarized herein suggest design of a capillary barrier which can reduce a recharge rate of 0.1 cm/yr by one or two orders of magnitude seems feasible for both concepts. A benchmark comparison of the unit cell model against a full facility model shows comparable predictive accuracy in less than one percent of the computer time. Results suggest that model parameters include capillary barrier performance, inter-canister spacing, rate of moisture withdrawal due to glass corrosion, contaminant inventory, and the well interceptor factor. It is also important that variations of waste form hydraulic parameters suggest that transport through the waste form is dominated by diffusion

  16. Dynamic expression of the translational machinery during Bacillus subtilis life cycle at a single cell level.

    Directory of Open Access Journals (Sweden)

    Alex Rosenberg

    Full Text Available The ability of bacteria to responsively regulate the expression of translation components is crucial for rapid adaptation to fluctuating environments. Utilizing Bacillus subtilis (B. subtilis as a model organism, we followed the dynamics of the translational machinery at a single cell resolution during growth and differentiation. By comprehensive monitoring the activity of the major rrn promoters and ribosomal protein production, we revealed diverse dynamics between cells grown in rich and poor medium, with the most prominent dissimilarities exhibited during deep stationary phase. Further, the variability pattern of translational activity varied among the cells, being affected by nutrient availability. We have monitored for the first time translational dynamics during the developmental process of sporulation within the two distinct cellular compartments of forespore and mother-cell. Our study uncovers a transient forespore specific increase in expression of translational components. Finally, the contribution of each rrn promoter throughout the bacterium life cycle was found to be relatively constant, implying that differential expression is not the main purpose for the existence of multiple rrn genes. Instead, we propose that coordination of the rrn operons serves as a strategy to rapidly fine tune translational activities in a synchronized fashion to achieve an optimal translation level for a given condition.

  17. A low-level diode laser therapy reduces the lipopolysaccharide (LPS)-induced periodontal ligament cell inflammation

    International Nuclear Information System (INIS)

    The purpose of this study was to investigate the cytologic effects of inflammatory periodontal ligament cells in vitro after low-level laser therapy. Human periodontal ligament cells were cultured, exposed to lipopolysaccharide and subjected to low-level laser treatment of 5 J cm−2 or 10 J cm−2 using a 920 nm diode laser. A periodontal ligament cell attachment was observed under a microscope, and the cell viability was quantified by a mitochondrial colorimetric assay. Lipopolysaccharide-treated periodontal ligament cells were irradiated with the low-level laser, and the expression levels of several inflammatory markers, iNOS, TNF-α and IL-1, and pErk kinase, were analyzed by reverse transcription polymerase chain reaction and western blot. The data were collected and analyzed by one-way analysis of variance; p < 0.05 indicated a statistically significant difference. The low-level laser treatment of periodontal ligament cells increased their ability to attach and survive. After irradiation, the expression levels of iNOS, TNF-α and IL-1 in lipopolysaccharide-exposed periodontal ligament cells decreased over time (p < 0.05). In periodontal ligament cells, low-level diode laser treatment increased the cells’ proliferative ability and decreased the expression of the examined inflammatory mediators. (letters)

  18. Variation of DNA damage levels in peripheral blood mononuclear cells isolated in different laboratories

    DEFF Research Database (Denmark)

    Godschalk, Roger W L; Ersson, Clara; Stępnik, Maciej;

    2014-01-01

    This study investigated the levels of DNA strand breaks and formamidopyrimidine DNA glycosylase (FPG) sensitive sites, as assessed by the comet assay, in peripheral blood mononuclear cells (PBMC) from healthy women from five different countries in Europe. The laboratory in each country (referred to...... as 'centre') collected and cryopreserved PBMC samples from three donors, using a standardised cell isolation protocol. The samples were analysed in 13 different laboratories for DNA damage, which is measured by the comet assay. The study aim was to assess variation in DNA damage in PBMC samples that...... were collected in the same way and processed using the same blood isolation procedure. The inter-laboratory variation was the prominent contributor to the overall variation. The inter-laboratory coefficient of variation decreased for both DNA strand breaks (from 68 to 26%) and FPG sensitive sites (from...

  19. Supraphysiological Levels of Quercetin Glycosides are Required to Alter Mineralization in Saos2 Cells.

    Science.gov (United States)

    Nash, Leslie A; Peters, Sandra J; Sullivan, Philip J; Ward, Wendy E

    2016-01-01

    Flavonoid intake is positively correlated to bone mineral density (BMD) in women. Flavonoids such as quercetin exhibit strong anti-oxidant and anti-inflammatory activity that may be beneficial for bone health. Quercetin, previously shown to positively influence osteoblasts, is metabolized into glycosides including rutin and hyperoside. We compared the effects of these glycosides on mineralization in human osteoblast (Saos2) cells. Administration of rutin (≥25 µM) and hyperoside (≥5 µM) resulted in higher mineral content, determined using the alizarin red assay. This was accompanied by higher alkaline phosphatase activity with no cell toxicity. The expression of osteopontin, sclerostin, TNFα and IL6, known stimuli for decreasing osteoblast activity, were reduced with the addition of rutin or hyperoside. In summary, rutin and hyperoside require supraphysiological levels, when administered individually, to positively influence osteoblast activity. This information may be useful in developing nutraceuticals to support bone health. PMID:27136576

  20. Supraphysiological Levels of Quercetin Glycosides are Required to Alter Mineralization in Saos2 Cells

    Science.gov (United States)

    Nash, Leslie A.; Peters, Sandra J.; Sullivan, Philip J.; Ward, Wendy E.

    2016-01-01

    Flavonoid intake is positively correlated to bone mineral density (BMD) in women. Flavonoids such as quercetin exhibit strong anti-oxidant and anti-inflammatory activity that may be beneficial for bone health. Quercetin, previously shown to positively influence osteoblasts, is metabolized into glycosides including rutin and hyperoside. We compared the effects of these glycosides on mineralization in human osteoblast (Saos2) cells. Administration of rutin (≥25 µM) and hyperoside (≥5 µM) resulted in higher mineral content, determined using the alizarin red assay. This was accompanied by higher alkaline phosphatase activity with no cell toxicity. The expression of osteopontin, sclerostin, TNFα and IL6, known stimuli for decreasing osteoblast activity, were reduced with the addition of rutin or hyperoside. In summary, rutin and hyperoside require supraphysiological levels, when administered individually, to positively influence osteoblast activity. This information may be useful in developing nutraceuticals to support bone health. PMID:27136576

  1. Migration and differentiation of bone marrow-derived multipotent adult progenitor cells through tail vein injection in a rat model of cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Lei Lei; Ruixiang Zhou

    2009-01-01

    BACKGROUND: Multipotent adult progenitor cells (MAPCs) from the bone marrow have been shown to differentiate into neurons.OBJECTIVE: To observe migration, survival, and neuronal-like differentiation of MAPCs by tail vein injection.DESIGN, TIME AND SETTING: Randomized, controlled experiment of neural tissue engineering was performed at the Laboratory for Cardio-Cerebrovascular Disease, Hospital of Integrated Traditional and Western Medicine, Tongji Medical College of Huazhong University of Science and Technology between September 2006 and August 2007.MATERIALS: Eighty Sprague Dawley rats, 3-6 months old, underwent cerebral ischemia/reperfusion by thread technique, and were randomly divided into model and MAPCs groups (n = 40).METHODS: Mononuclear cells were harvested from bone marrow using the Ficoll-Paque density gradient centrifugation method. After removing CD45 and glycophorin A-positive cells (GLYA+) with immunomagnetic beads, CD45 GLYA adult progenitor cells were labeled with bromodeoxyuridine (5-bromo-2-deoxyuridine, BrdU). A total of 1 mL cell suspension, containing 5 ×106 MAPCs, was injected into the MAPCs group through the tail vein. A total of 1 mL normal saline was injected into the model rats.MAIN OUTCOME MEASURES: After 60 days, BrdU and neuron-specific enolase double-positive cells were observed using immunofluorescence. Cell morphology was observed under electron microscopy, and nerve growth factor mRNA was measured through RT-PCR. In addition, rat neurological functions were measured with behavioral tests.RESULTS: Immunofluorescence revealed that MAPCs positive for BrdU and neuron specific enolase were found surrounding the ischemic focus in the MAPCs group. Microscopic observation suggested that MAPCs-derived neuronal-like cells connected with other nerve cells to form synapses, Compared with the model animals, the level of nerve growth factor mRNA was significantly upregulated in rats injected with MAPCs (P < 0.05). In addition, rats in the MAPCs

  2. Nuclear Factor κB1/RelA Mediates Inflammation in Human Lung Epithelial Cells at Atmospheric Oxygen Levels.

    Science.gov (United States)

    Jagannathan, Lakshmanan; Jose, Cynthia C; Arita, Adriana; Kluz, Thomas; Sun, Hong; Zhang, Xiaoru; Yao, Yixin; Kartashov, Andrey V; Barski, Artem; Costa, Max; Cuddapah, Suresh

    2016-07-01

    Oxygen levels range from 2% to 9% in vivo. Atmospheric O2 levels (21%) are known to induce cell proliferation defects and cellular senescence in primary cell cultures. However, the mechanistic basis of the deleterious effects of higher O2 levels is not fully understood. On the other hand, immortalized cells including cancer cell lines, which evade cellular senescence are normally cultured at 21% O2 and the effects of higher O2 on these cells are understudied. Here, we addressed this problem by culturing immortalized human bronchial epithelial (BEAS-2B) cells at ambient atmospheric, 21% O2 and lower, 10% O2. Our results show increased inflammatory response at 21% O2 but not at 10% O2. We found higher RelA binding at the NF-κB1/RelA target gene promoters as well as upregulation of several pro-inflammatory cytokines in cells cultured at 21% O2. RelA knockdown prevented the upregulation of the pro-inflammatory cytokines at 21% O2, suggesting NF-κB1/RelA as a major mediator of inflammatory response in cells cultured at 21% O2. Interestingly, unlike the 21% O2 cultured cells, exposure of 10% O2 cultured cells to H2O2 did not elicit inflammatory response, suggesting increased ability to tolerate oxidative stress in cells cultured at lower O2 levels. PMID:26588041

  3. Fluorescent reporter signals, EGFP and DsRed, encoded in HIV-1 facilitate the detection of productively infected cells and cell-associated viral replication levels

    Directory of Open Access Journals (Sweden)

    Kazutaka eTerahara

    2012-01-01

    Full Text Available Flow cytometric analysis is a reliable and convenient method for investigating molecules at the single cell level. Previously, recombinant human immunodeficiency virus type 1 (HIV-1 strains were constructed that express a fluorescent reporter, either enhanced green fluorescent protein or DsRed, which allow the monitoring of HIV-1-infected cells by flow cytometry. The present study further investigated the potential of these recombinant viruses in terms of whether the HIV-1 fluorescent reporters would be helpful in evaluating viral replication based on fluorescence intensity. When primary CD4+ T cells were infected with recombinant viruses, the fluorescent reporter intensity measured by flow cytometry was associated with the level of CD4 downmodulation and Gag p24 expression in infected cells. Interestingly, some HIV-1-infected cells, in which CD4 was only moderately downmodulated, were reporter-positive but Gag p24-negative. Furthermore, when the activation status of primary CD4+ T cells was modulated by T cell receptor-mediated stimulation, we confirmed the preferential viral production upon strong stimulation and showed that the intensity of the fluorescent reporter within a proportion of HIV-1-infected cells was correlated with the viral replication level. These findings indicate that a fluorescent reporter encoded within HIV-1 is useful for the sensitive detection of productively-infected cells at different stages of infection and for evaluating cell-associated viral replication at the single cell level.

  4. Naringin promotes differentiation of bone marrow stem cells into osteoblasts by upregulating the expression levels of microRNA-20a and downregulating the expression levels of PPARγ.

    Science.gov (United States)

    Fan, Jifeng; Li, Jie; Fan, Qinbo

    2015-09-01

    Naringin is a dihydrotestosterone flavonoid compound that significantly inhibits bone loss, improves bone density, and enhances biomechanical anti‑compression performance. Previous studies have demonstrated that naringin improves the activity levels of osteocalcin (OC) and alkaline phosphatase (ALP) in MC3T3‑E1 osteoblast precursor cells. The present study investigated the effects of naringin on osteoblastic differentiation and inhibition of adipocyte formation in bone marrow stem cells (BMSCs). The levels of osteogenesis were modulated via upregulation of the expression levels of microRNA (miR)‑20a, and downregulation of the expression levels of peroxisome proliferator‑activated receptor γ (PPARγ). The results indicated that naringin significantly enhanced BMSC proliferation in a dose‑dependent manner. In addition, naringin significantly increased the mRNA expression levels of OC, ALP, and collagen type I. Furthermore, naringin decreased the protein expression levels of PPARγ, and increased the expression levels of miR‑20a in the BMSCs. These results suggested that miR‑20a may regulate the expression of PPARγ in BMSCs. To our knowledge, this is the first study to report naringin‑induced osteogenesis via upregulation of the expression levels of miR‑20a, and downregulation of the expression levels of PPARγ. These results indicated the important role of naringin in BMSC differentiation. PMID:26126997

  5. Cleavage efficient 2A peptides for high level monoclonal antibody expression in CHO cells.

    Science.gov (United States)

    Chng, Jake; Wang, Tianhua; Nian, Rui; Lau, Ally; Hoi, Kong Meng; Ho, Steven C L; Gagnon, Peter; Bi, Xuezhi; Yang, Yuansheng

    2015-01-01

    Linking the heavy chain (HC) and light chain (LC) genes required for monoclonal antibodies (mAb) production on a single cassette using 2A peptides allows control of LC and HC ratio and reduces non-expressing cells. Four 2A peptides derived from the foot-and-mouth disease virus (F2A), equine rhinitis A virus (E2A), porcine teschovirus-1 (P2A) and Thosea asigna virus (T2A), respectively, were compared for expression of 3 biosimilar IgG1 mAbs in Chinese hamster ovary (CHO) cell lines. HC and LC were linked by different 2A peptides both in the absence and presence of GSG linkers. Insertion of a furin recognition site upstream of 2A allowed removal of 2A residues that would otherwise be attached to the HC. Different 2A peptides exhibited different cleavage efficiencies that correlated to the mAb expression level. The relative cleavage efficiency of each 2A peptide remains similar for expression of different IgG1 mAbs in different CHO cells. While complete cleavage was not observed for any of the 2A peptides, GSG linkers did enhance the cleavage efficiency and thus the mAb expression level. T2A with the GSG linker (GT2A) exhibited the highest cleavage efficiency and mAb expression level. Stably amplified CHO DG44 pools generated using GT2A had titers 357, 416 and 600 mg/L for the 3 mAbs in shake flask batch cultures. Incomplete cleavage likely resulted in incorrectly processed mAb species and aggregates, which were removed with a chromatin-directed clarification method and protein A purification. The vector and methods presented provide an easy process beneficial for both mAb development and manufacturing. PMID:25621616

  6. Cancer Stem Cells in Small Cell Lung Cancer Cell Line H446: Higher Dependency on Oxidative Phosphorylation and Mitochondrial Substrate-Level Phosphorylation than Non-Stem Cancer Cells

    OpenAIRE

    Gao, Cuicui; Shen, Yao; Jin, Fang; Miao, Yajing; Qiu, Xiaofei

    2016-01-01

    Recently, targeting cancer stem cells (CSCs) metabolism is becoming a promising therapeutic approach to improve cancer treatment outcomes. However, knowledge of the metabolic state of CSCs in small cell lung cancer is still lacking. In this study, we found that CSCs had significantly lower oxygen consumption rate and extracellular acidification rate than non-stem cancer cells. Meanwhile, this subpopulation of cells consumed less glucose, produced less lactate and maintained lower ATP levels. ...

  7. The active principle region of Buyang Huanwu decoction induced differentiation of bone marrow-derived mesenchymal stem cells into neural-like cells Superior effects over original formula of Buyang Huanwu decoction

    Institute of Scientific and Technical Information of China (English)

    Jinghui Zheng; Yi Wan; Jianhuai Chi; Dekai Shen; Tingting Wu; Weimin Li; Pengcheng Du

    2012-01-01

    The present study induced in vitro-cultured passage 4 bone marrow-derived mesenchymal stem cells to differentiate into neural-like cells with a mixture of alkaloid, polysaccharide, aglycone, glycoside, essential oils, and effective components of Buyang Huanwu decoction (active principle region of decoction for invigorating yang for recuperation). After 28 days, nestin and neuron-specific enolase were expressed in the cytoplasm. Reverse transcription-PCR and western blot analyses showed that nestin and neuron-specific enolase mRNA and protein expression was greater in the active principle region group compared with the original formula group. Results demonstrated that the active principle region of Buyang Huanwu decoction induced greater differentiation of rat bone marrow-derived mesenchymal stem cells into neural-like cells in vitro than the original Buyang Huanwu decoction formula.

  8. Selenium supplementation in patients undergoing hematopoietic stem cell transplantation: effects on pro-inflammatory cytokines levels

    OpenAIRE

    Daeian, Nesa; Radfar, Mania; Jahangard-Rafsanjani, Zahra; Hadjibabaie, Molouk; Ghavamzadeh, Ardeshir

    2014-01-01

    Background Pro-inflammatory cytokines including tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) play an important role in the development of hematopoietic stem cell transplantation (HSCT) complications. We explored the effect of Selenium as an antioxidant and anti-inflammatory agent on pro-inflammatory cytokines levels in HSCT candidates. Findings Plasma concentrations of TNF-α, IL-1β and IL-6 were measured in 74 patients from a double-blind, randomized, p...

  9. Metabolism of red blood cells in chronic renal failure. I. Glycolytic enzyme levels.

    Science.gov (United States)

    Rodríguez-Commes, J L; Tabernero, J M; Martin-Vasallo, P; De Castro, S; Battaner, E

    1979-01-01

    This paper starts a series on red blood cell (RBC) metabolism in patients with chronic renal failure (CRF). The glycolytic enzyme levels and in vitro half-lives of these patients' RBCs were determined. A number of enzymes (hexokinase, glucose-6-phosphate isomerase, fructose-6-phosphate kinase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase) showed higher activities than in normal control RBCs. Other enzyme activities were normal. These results were discussed and several possible mechanisms considered. We favour the point of view of a shortened life span of the RBCs in CRF, making the most unstable enzymes of the glycolytic sequence appear increase as compared with normal controls. PMID:226898

  10. The effect of ipt transgene expression on a single-cell level

    Czech Academy of Sciences Publication Activity Database

    Křížková, Lucie; Petrášek, Jan; Březinová, Alena; Zažímalová, Eva

    Habana: Centro de Ingeniería Genética y Biotecnología, 2002. s. -. [La Agrobiotecnología en el Nuevo Milenio. 24.11.2002-29.11.2002, Habana] R&D Projects: GA ČR GV206/02/P106; GA AV ČR IAB6038203; GA MŠk LN00A081 Grant ostatní: INCO Copernicus(BE) IC15-CT98-0118 Keywords : transgene expression * single cell level * ipt Subject RIV: EB - Genetics ; Molecular Biology

  11. Comparison of the nonradiative deep levels in silicon solar cells made of monocrystalline, polycrystalline and amorphous silicon using deep level transient spectroscopy (DLTS)

    International Nuclear Information System (INIS)

    The aim of this work is to study the defects in solar cells fabricated from crystalline, polycrystalline and amorphous silicon. Using Deep Level Transient Spectroscopy technique, (DLTS), we have determined their activation energies, concentrations and their effect on the solar cell efficiency. Our results show a DLTS peak in crystalline silicon which we could attribute to tow peaks originating from iron contamination. In the polycrystalline based solar cells we observed a series of non conventional DLTS peaks while in amorphous silicon we observed a peak using low measurement frequencies (between 8 kHz and 20 kHz). We studied these defects and determined their activation energies as well as the capture cross section for one of them. We suggest a possible configuration of these defects. We cannot able to study the effect of these defects on the solar cell efficiency because we have not the experimental set-up which measure the solar cell efficiency. (Authors)

  12. Insulin stimulates endothelin-1 secretion from human endothelial cells and modulates its circulating levels in vivo.

    Science.gov (United States)

    Ferri, C; Pittoni, V; Piccoli, A; Laurenti, O; Cassone, M R; Bellini, C; Properzi, G; Valesini, G; De Mattia, G; Santucci, A

    1995-03-01

    Endothelin-1 (ET-1) is a potent vasoactive and mitogenic peptide produced by the vascular endothelium. In this study, we evaluated whether insulin stimulates ET-1 secretion by human endothelial cells derived from umbilical cord veins and by human permanent endothelial hybrid cells Ea.hy 926. Moreover, to provide evidence that insulin may stimulate ET-1 secretion in vivo, plasma ET-1 levels were evaluated in 7 type II diabetic normotensive males (mean age, 54.3 +/- 4.0 yr) during 2-h hyperinsulinemic euglycemic clamps (287 pmol insulin/m2.min-1) as well as in 12 obese hypertensive males (mean age, 44.2 +/- 4.6 yr) before and after a 12-week period of caloric restriction. Our results showed that insulin stimulated ET-1 release from cultured endothelial cells in a dose-dependent fashion. ET-1 release persisted for 24 h and was also observed at physiological insulin concentrations (10(-9) mol/L). The insulin-induced ET-1 secretion was inhibited by genistein, a tyrosine kinase inhibitor, and by cycloheximide, a protein synthesis inhibitor, suggesting that it requires de novo protein synthesis rather than ET-1 release from intracellular stores. In the in vivo experiments, plasma ET-1 levels rapidly increased during euglycemic hyperinsulinemic clamps (from 0.76 +/- 0.18 pg/mL at time zero to 1.65 +/- 0.21 pg/mL at 60 min; P < 0.05) and persisted elevated until the end of insulin infusion (1.37 +/- 0.37 pg/mL at 120 min; P < 0.05 vs. time zero). In obese hypertensives, plasma ET-1 levels significantly decreased after 12 weeks of caloric restriction (from 0.85 +/- 0.51 to 0.48 +/- 0.28 pg/mL; P < 0.04). The decrease in body weight induced by caloric restriction was accompanied by a significant reduction in fasting insulin levels (from 167.2 +/- 94.0 to 98.9 +/- 44.9 pmol/L; P < 0.05) which correlated with the reduction in plasma ET-1 levels (r = 0.78; P < 0.003). In conclusion, our data show that insulin stimulates both in vitro and in vivo ET-1 secretion. Such interaction

  13. Effects of low level radiation upon the hematopoietic stem cell: Implications for leukemogenesis

    International Nuclear Information System (INIS)

    These studies have addressed firstly the effect of single small doses of X-rays upon murine hematopoietic stem cells to obtain a better estimate of the Dq. It is small, of the order of 20 rad. Secondly, a dose fractionation schedule that does not kill or perturb the kinetcs of hemopoietic cell proliferation was sought in order to investigate the leukemogenic potential of low level radiation upon an unperturbed hemopoietic system. Doses used by others in past radiation leukemogenesis studies clearly perturb hemopoiesis and kill a detectable fraction of stem cells. The studies reported herein show that 1.25 rad every day decrease the CFU-S content of bone marrow by the time 80 rads are accumulated. Higher daily doses as used in published studies on radiation leukemogenesis produce greater effects. Studies on the effect of 0.5, 1.0, 2.0, and 3.0 rad 3 times per week are under way. Two rad 3 times per week produced a modest decrease in CFU-S content of bone marrow after an accumulation of 68 rad. With 3.0 rad 3 times per week an accumulation of 102 rad produced a significant decrease in CFU-S content of bone marrow. Dose fractionation at 0.5 and 1.0 rad 3 times per week has not produced a CFU-S depression after accumulation of 17 and 34 rad. Radiation leukemogenesis studies published to date have utilized single doses and chronic exposure schedules that probably have significantly perturbed the kinetcs of hematopoietic stem cells. Whether radiation will produce leukemia in animal models with dose schedules that do not perturb kinetics of hematopoietic stem cells remains to be seen. (orig.)

  14. Effects of low level radiation upon the hematopoietic stem cell: Implications for leukemogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Cronkite, E.P.; Bond, V.P.; Carsten, A.L.; Miller, M.E.; Bullis, J.E.; Inoue, T.

    1987-06-01

    These studies have addressed firstly the effect of single small doses of X-rays upon murine hematopoietic stem cells to obtain a better estimate of the D/sub q/. It is small, of the order of 20 rad. Secondly, a dose fractionation schedule that does not kill or perturb the kinetcs of hemopoietic cell proliferation was sought in order to investigate the leukemogenic potential of low level radiation upon an unperturbed hemopoietic system. Doses used by others in past radiation leukemogenesis studies clearly perturb hemopoiesis and kill a detectable fraction of stem cells. The studies reported herein show that 1.25 rad every day decrease the CFU-S content of bone marrow by the time 80 rads are accumulated. Higher daily doses as used in published studies on radiation leukemogenesis produce greater effects. Studies on the effect of 0.5, 1.0, 2.0, and 3.0 rad 3 times per week are under way. Two rad 3 times per week produced a modest decrease in CFU-S content of bone marrow after an accumulation of 68 rad. With 3.0 rad 3 times per week an accumulation of 102 rad produced a significant decrease in CFU-S content of bone marrow. Dose fractionation at 0.5 and 1.0 rad 3 times per week has not produced a CFU-S depression after accumulation of 17 and 34 rad. Radiation leukemogenesis studies published to date have utilized single doses and chronic exposure schedules that probably have significantly perturbed the kinetcs of hematopoietic stem cells. Whether radiation will produce leukemia in animal models with dose schedules that do not perturb kinetics of hematopoietic stem cells remains to be seen.

  15. Effects of low-level radiation upon the haematopoietic stem cell. Implications for leukaemogenesis

    International Nuclear Information System (INIS)

    These studies address first the effect of single small doses of X-rays upon murine haematopoietic stem cells to obtain a better estimate of the Dsub(q). It is small, of the order of 20 rad. Second, a dose fractionation schedule that does not kill or perturb the kinetics of haematopoietic cell proliferation was sought to investigate the leukaemogenic potential of low-level radiation upon an unperturbed haematopoietic system. Doses used by others in past radiation leukaemogenesis studies clearly perturb haematopoiesis and kill a detectable fraction of stem cells. The studies reported in the paper show that 1.25 rad every day decreases the CFU-S content of bone marrow by the time 80 rad are accumulated. Higher daily doses as used in published studies on radiation leukaemogenesis produce greater effects. Studies of the effect of 0.5, 1.0, 2.0 and 3.0 rad three times per week are under way. Two rad three times per week produce a modest decrease in CFU-S content of bone marrow after an accumulation of 68 rad. With 3.0 rad three times per week, an accumulation of 102 rad produces a significant decrease in CFU-S content of bone marrow. Dose fractionation at 0.5 and 1.0 rad three times per week has not produced a CFU-S depression after accumulation of 17 and 34 rad. Radiation leukaemogenesis studies published to date have used single doses and chronic exposure schedules that probably have significantly perturbed the kinetics of haematopoietic stem cells. Whether radiation will produce leukaemia in animal models with dose schedules that do not perturb the kinetics of haematopoietic stem cells remains to be seen. (author)

  16. Synergism between arsenite and proteasome inhibitor MG132 over cell death in myeloid leukaemic cells U937 and the induction of low levels of intracellular superoxide anion

    Energy Technology Data Exchange (ETDEWEB)

    Lombardo, Tomás [Laboratorio de Immunotoxicologia (LaITO), IDEHU-CONICET, Hospital de Clínicas, José de San Martín, Universidad de Buenos Aires (UBA), Buenos Aires (Argentina); Cavaliere, Victoria; Costantino, Susana N. [Laboratorio de Inmunología Tumoral (LIT), IDEHU-CONICET, Facultad de Farmacia y Bioquímica, UBA, Buenos Aires (Argentina); Kornblihtt, Laura [Servicio de Hematología, Hospital de Clínicas, José de San Martín (UBA), Buenos Aires (Argentina); Alvarez, Elida M. [Laboratorio de Inmunología Tumoral (LIT), IDEHU-CONICET, Facultad de Farmacia y Bioquímica, UBA, Buenos Aires (Argentina); Blanco, Guillermo A., E-mail: gblanco@ffyb.uba.ar [Laboratorio de Immunotoxicologia (LaITO), IDEHU-CONICET, Hospital de Clínicas, José de San Martín, Universidad de Buenos Aires (UBA), Buenos Aires (Argentina)

    2012-02-01

    Increased oxygen species production has often been cited as a mechanism determining synergism on cell death and growth inhibition effects of arsenic-combined drugs. However the net effect of drug combination may not be easily anticipated solely from available knowledge of drug-induced death mechanisms. We evaluated the combined effect of sodium arsenite with the proteasome inhibitor MG132, and the anti-leukaemic agent CAPE, on growth-inhibition and cell death effect in acute myeloid leukaemic cells U937 and Burkitt's lymphoma-derived Raji cells, by the Chou–Talalay method. In addition we explored the association of cytotoxic effect of drugs with changes in intracellular superoxide anion (O{sub 2}{sup −}) levels. Our results showed that combined arsenite + MG132 produced low levels of O{sub 2}{sup −} at 6 h and 24 h after exposure and were synergic on cell death induction in U937 cells over the whole dose range, although the combination was antagonistic on growth inhibition effect. Exposure to a constant non-cytotoxic dose of 80 μM hydrogen peroxide together with arsenite + MG132 changed synergism on cell death to antagonism at all effect levels while increasing O{sub 2}{sup −} levels. Arsenite + hydrogen peroxide also resulted in antagonism with increased O{sub 2}{sup −} levels in U937 cells. In Raji cells, arsenite + MG132 also produced low levels of O{sub 2}{sup −} at 6 h and 24 h but resulted in antagonism on cell death and growth inhibition. By contrast, the combination arsenite + CAPE showed high levels of O{sub 2}{sup −} production at 6 h and 24 h post exposure but resulted in antagonism over cell death and growth inhibition effects in U937 and Raji cells. We conclude that synergism between arsenite and MG132 in U937 cells is negatively associated to O{sub 2}{sup −} levels at early time points after exposure. -- Highlights: ► Arsenic combined cytotoxic and anti-proliferative effects by Chou–Talalay method. ► Cytotoxic effect

  17. Synergism between arsenite and proteasome inhibitor MG132 over cell death in myeloid leukaemic cells U937 and the induction of low levels of intracellular superoxide anion

    International Nuclear Information System (INIS)

    Increased oxygen species production has often been cited as a mechanism determining synergism on cell death and growth inhibition effects of arsenic-combined drugs. However the net effect of drug combination may not be easily anticipated solely from available knowledge of drug-induced death mechanisms. We evaluated the combined effect of sodium arsenite with the proteasome inhibitor MG132, and the anti-leukaemic agent CAPE, on growth-inhibition and cell death effect in acute myeloid leukaemic cells U937 and Burkitt's lymphoma-derived Raji cells, by the Chou–Talalay method. In addition we explored the association of cytotoxic effect of drugs with changes in intracellular superoxide anion (O2−) levels. Our results showed that combined arsenite + MG132 produced low levels of O2− at 6 h and 24 h after exposure and were synergic on cell death induction in U937 cells over the whole dose range, although the combination was antagonistic on growth inhibition effect. Exposure to a constant non-cytotoxic dose of 80 μM hydrogen peroxide together with arsenite + MG132 changed synergism on cell death to antagonism at all effect levels while increasing O2− levels. Arsenite + hydrogen peroxide also resulted in antagonism with increased O2− levels in U937 cells. In Raji cells, arsenite + MG132 also produced low levels of O2− at 6 h and 24 h but resulted in antagonism on cell death and growth inhibition. By contrast, the combination arsenite + CAPE showed high levels of O2− production at 6 h and 24 h post exposure but resulted in antagonism over cell death and growth inhibition effects in U937 and Raji cells. We conclude that synergism between arsenite and MG132 in U937 cells is negatively associated to O2− levels at early time points after exposure. -- Highlights: ► Arsenic combined cytotoxic and anti-proliferative effects by Chou–Talalay method. ► Cytotoxic effect associated with superoxide levels as assessed by flow cytometry. ► Synergism between arsenite

  18. A low-level diode laser therapy reduces the lipopolysaccharide (LPS)-induced periodontal ligament cell inflammation

    Science.gov (United States)

    Huang, T. H.; Chen, C. C.; Liu, S. L.; Lu, Y. C.; Kao, C. T.

    2014-07-01

    The purpose of this study was to investigate the cytologic effects of inflammatory periodontal ligament cells in vitro after low-level laser therapy. Human periodontal ligament cells were cultured, exposed to lipopolysaccharide and subjected to low-level laser treatment of 5 J cm-2 or 10 J cm-2 using a 920 nm diode laser. A periodontal ligament cell attachment was observed under a microscope, and the cell viability was quantified by a mitochondrial colorimetric assay. Lipopolysaccharide-treated periodontal ligament cells were irradiated with the low-level laser, and the expression levels of several inflammatory markers, iNOS, TNF-α and IL-1, and pErk kinase, were analyzed by reverse transcription polymerase chain reaction and western blot. The data were collected and analyzed by one-way analysis of variance; p periodontal ligament cells increased their ability to attach and survive. After irradiation, the expression levels of iNOS, TNF-α and IL-1 in lipopolysaccharide-exposed periodontal ligament cells decreased over time (p periodontal ligament cells, low-level diode laser treatment increased the cells’ proliferative ability and decreased the expression of the examined inflammatory mediators.

  19. Multi-level behaviours in agent-based simulation: colonic crypt cell populations

    Science.gov (United States)

    Chen, Chih-Chun; Nagl, Sylvia B.; Clack, Christopher D.

    Agent-based modelling and simulation is now beginning to establish itself as a suitable technique for studying biological systems. However, a major issue in using agent-based simulations to study complex systems such as those in Systems Biology is the fact that simulations are `opaque'. While we have knowledge of individuals' behaviour through agent rules and have techniques for evaluating global behaviour by aggregating the states of individuals, methods for identifying the interactive mechanisms giving rise to this global behaviour are lacking. Formulating precise hypotheses about these multi-level behaviours is also difficult without an established formalism for describing them. The complex event formalism allows relationships between agent-rule-generated events to be defined so that behaviours at different levels of abstraction to be described. Complex event types define categories of these behaviours, which can then be detected in simulation, giving us computational method for distinguishing between alternative interactive mechanisms underlying a higher level behaviour. We apply the complex event formalism to an agent-based model of cell populations in the colonic crypt and demonstrate how competition and selection events can be identified in simulation at both the individual and clonal level, allowing us to computationally test hypotheses about the interactive mechanisms underlying a clone's success.

  20. Inhibition of hyaluronan synthesis reduces versican and fibronectin levels in trabecular meshwork cells.

    Directory of Open Access Journals (Sweden)

    Kate E Keller

    Full Text Available Hyaluronan (HA is a major component of the extracellular matrix (ECM and is synthesized by three HA synthases (HAS. Similarities between the HAS2 knockout mouse and the hdf mutant mouse, which has a mutation in the versican gene, suggest that HA and versican expression may be linked. In this study, the relationship between HA synthesis and levels of versican, fibronectin and several other ECM components in trabecular meshwork cells from the anterior segment of the eye was investigated. HA synthesis was inhibited using 4-methylumbelliferone (4MU, or reduced by RNAi silencing of each individual HAS gene. Quantitative RT-PCR and immunoblotting demonstrated a reduction in mRNA and protein levels of versican and fibronectin. Hyaluronidase treatment also reduced versican and fibronectin levels. These effects could not be reversed by addition of excess glucose or glucosamine or exogenous HA to the culture medium. CD44, tenascin C and fibrillin-1 mRNA levels were reduced by 4MU treatment, but SPARC and CSPG6 mRNA levels were unaffected. Immunostaining of trabecular meshwork tissue after exposure to 4MU showed an altered localization pattern of HA-binding protein, versican and fibronectin. Reduction of versican by RNAi silencing did not affect HA concentration as assessed by ELISA. Together, these data imply that HA concentration affects synthesis of certain ECM components. Since precise regulation of the trabecular meshwork ECM composition and organization is required to maintain the aqueous humor outflow resistance and intraocular pressure homeostasis in the eye, coordinated coupling of HA levels and several of its ECM binding partners should facilitate this process.

  1. The level of circulating endothelial progenitor cells may be associated with the occurrence and recurrence of chronic subdural hematoma

    Directory of Open Access Journals (Sweden)

    Yan Song

    2013-01-01

    Full Text Available OBJECTIVES: The onset of chronic subdural hematoma may be associated with direct or indirect minor injuries to the head or a poorly repaired vascular injury. Endothelial progenitor cells happen to be one of the key factors involved in hemostasis and vascular repair. This study was designed to observe the levels of endothelial progenitor cells, white blood cells, platelets, and other indicators in the peripheral blood of patients diagnosed with chronic subdural hematoma to determine the possible relationship between the endothelial progenitor cells and the occurrence, development, and outcomes of chronic subdural hematoma. METHOD: We enrolled 30 patients with diagnosed chronic subdural hematoma by computer tomography scanning and operating procedure at Tianjin Medical University General Hospital from July 2009 to July 2011. Meanwhile, we collected 30 cases of peripheral blood samples from healthy volunteers over the age of 50. Approximately 2 ml of blood was taken from veins of the elbow to test the peripheral blood routine and coagulation function. The content of endothelial progenitor cells in peripheral blood mononuclear cells was determined by flow cytometry. RESULTS: The level of endothelial progenitor cells in peripheral blood was significantly lower in preoperational patients with chronic subdural hematomas than in controls. There were no significant differences between the two groups regarding the blood routine and coagulation function. However, the levels of circulating endothelial progenitor cells were significantly different between the recurrent group and the non-recurrent group. CONCLUSIONS: The level of circulating endothelial progenitor cells in chronic subdural hematoma patients was significantly lower than the level in healthy controls. Meanwhile, the level of endothelial progenitor cells in recurrent patients was significantly lower than the level in patients without recurrence. Endothelial progenitor cells may be related to the

  2. Membrane estrogen receptor-α levels predict estrogen-induced ERK1/2 activation in MCF-7 cells

    International Nuclear Information System (INIS)

    We examined the participation of a membrane form of estrogen receptor (mER)-α in the activation of mitogen-activated protein kinases (extracellular signal-regulated kinase [ERK]1 and ERK2) related to cell growth responses in MCF-7 cells. We immunopanned and subsequently separated MCF-7 cells (using fluorescence-activated cell sorting) into mER-α-enriched (mERhigh) and mER-α-depleted (mERlow) populations. We then measured the expression levels of mER-α on the surface of these separated cell populations by immunocytochemical analysis and by a quantitative 96-well plate immunoassay that distinguished between mER-α and intracellular ER-α. Western analysis was used to determine colocalized estrogen receptor (ER)-α and caveolins in membrane subfractions. The levels of activated ERK1 and ERK2 were determined using a fixed cell-based enzyme-linked immunosorbent assay developed in our laboratory. Immunocytochemical studies revealed punctate ER-α antibody staining of the surface of nonpermeabilized mERhigh cells, whereas the majority of mERlow cells exhibited little or no staining. Western analysis demonstrated that mERhigh cells expressed caveolin-1 and caveolin-2, and that ER-α was contained in the same gradient-separated membrane fractions. The quantitative immunoassay for ER-α detected a significant difference in mER-α levels between mERhigh and mERlow cells when cells were grown at a sufficiently low cell density, but equivalent levels of total ER-α (membrane plus intracellular receptors). These two separated cell subpopulations also exhibited different kinetics of ERK1/2 activation with 1 pmol/l 17β-estradiol (E2), as well as different patterns of E2 dose-dependent responsiveness. The maximal kinase activation was achieved after 10 min versus 6 min in mERhigh versus mERlow cells, respectively. After a decline in the level of phosphorylated ERKs, a reactivation was seen at 60 min in mERhigh cells but not in mERlow cells. Both 1A and 2B protein phosphatases

  3. Tumor-promoting phorbol ester transiently down-modulates the p53 level and blocks the cell cycle

    DEFF Research Database (Denmark)

    Skouv, J; Jensen, P O; Forchhammer, J;

    1994-01-01

    Activation of the protein kinase C signaling pathway by tumor-promoting phorbol esters, such as 4 beta-phorbol 12-myristate 13-acetate (PMA), induced a decrease in the level of p53 mRNA in several serum-starved human cell lines. Also, the tumor-promoting phosphatase inhibitor okadaic acid induced a...... decrease in the p53 mRNA level in the cell lines. Normal diploid as well as various tumor cell lines were tested. Two tumor cell lines, HeLa and A549, both containing the wild-type p53 gene, but very different levels of p53 protein, were studied in detail. In both cell lines, the level of p53 mRNA was...... minimal after 9 h of exposure to PMA. After approximately 120 h, the p53 mRNA level was similar to the pretreatment level. PMA induced a similar transient decrease in the level of p53 protein in the A549 cell line. The decrease in the p53 mRNA level could not be explained by changes in the transcriptional...

  4. Changes in metabolic proteins in ex vivo rat retina during glutamate-induced neural progenitor cell induction.

    Science.gov (United States)

    Tokuda, Kazuhiro; Kuramitsu, Yasuhiro; Baron, Byron; Kitagawa, Takao; Tokuda, Nobuko; Kobayashi, Masaaki; Kimura, Kazuhiro; Sonoda, Koh-Hei; Nakamura, Kazuyuki

    2016-08-01

    Understanding how energy metabolism and related proteins influence neural progenitor cells in adult tissues is critical for developing new strategies in clinical tissue regeneration therapy. We have recently reported that a subtoxic concentration of glutamate-induced neural progenitor cells in the mature ex vivo rat retina. We herein explore changes in the metabolic pathways during the process. We firstly observed an increase in lactate and lactate dehydrogenase concentration in the glutamate-treated retina. We then investigated the levels of glycolytic enzymes and confirmed significant upregulation of pyruvate kinase M type (PKM), especially PKM2, enolase, phosphoglycerate mutase 1 (PGAM1), and inosine-5'-monophosphate dehydrogenase (IMPDH1) in the glutamate-treated retina compared to the untreated retina. An analysis of the subcellular localization of PKM2 revealed nuclear translocation in the treated retina, which has been reported to regulate cell cycle proliferation and glycolytic enzymes. Our findings indicate that the mature rat retina undergoes an increase in aerobic glycolysis. PKM2, both in the cytoplasm and in the nucleus, may thus play an important role during neural progenitor cell induction, as it does in other proliferating cells. PMID:27421851

  5. The relationships between serum cytokine levels and tumor infiltrating immune cells and their clinical significance in colorectal cancer.

    Science.gov (United States)

    Väyrynen, Juha P; Kantola, Tiina; Väyrynen, Sara A; Klintrup, Kai; Bloigu, Risto; Karhu, Toni; Mäkelä, Jyrki; Herzig, Karl-Heinz; Karttunen, Tuomo J; Tuomisto, Anne; Mäkinen, Markus J

    2016-07-01

    Increased inflammatory cell infiltration correlates to improved survival in colorectal cancer (CRC). Development and progression of CRC is associated with alterations in serum cytokine levels but their significance is not well defined. In this study, we investigated the relationships between the serum levels of 13 cytokines and the densities of eight types of tumor infiltrating inflammatory cells and their impact on disease-free survival (DFS), cancer-specific survival (CSS) and overall survival (OS) in a prospectively recruited group of 147 CRC patients. There were strong positive correlations between the serum concentrations of different cytokines, as well as between the different types of tumor infiltrating immune cells, whereas the associations between serum cytokines and tumor infiltrating immune cells were generally weak. High serum IL-12 levels associated with increased densities of peritumoral CD8(+) T cells, intraepithelial CD3(+) T cells and intratumoral neutrophils, while high serum CCL4 levels associated with increased densities of peritumoral CD68(+) cells. In multivariate survival models, increased infiltration of intraepithelial CD3(+) T cells and increased serum CCL4 associated with improved DFS, whereas higher intratumoral CD83(+) dendritic cell density and increased serum interferon gamma levels associated with improved CSS and OS. Also high density of peritumoral CD3(+) T cells associated with improved CSS. In conclusion, serum cytokines and tumor infiltrating immune cells in CRC represent entities with high intragroup correlations but relatively weak intergroup correlations. The results suggest that tumor infiltrating CD3(+) T cells, CD83(+) dendritic cells, serum CCL4 and serum interferon gamma represent relevant markers of disease outcome. PMID:26874795

  6. Global analysis of protein expression and phosphorylation levels in nicotine-treated pancreatic stellate cells

    Science.gov (United States)

    Paulo, Joao A.; Gaun, Aleksandr; Gygi, Steven P.

    2016-01-01

    Smoking is a risk factor in pancreatic disease, however, the biochemical mechanisms correlating smoking with pancreatic dysfunction remain poorly understood. Strategies using multiplexed isobaric tag-based mass spectrometry facilitate the study of drug-induced perturbations on biological systems. Here, we present the first large scale analysis of the proteomic and phosphoproteomic alterations in pancreatic stellate cells following treatment with two nicotinic acetylcholine receptor (nAChR) ligands: nicotine and α-bungarotoxin. We treated cells with nicotine or α-bungarotoxin for 12hr in triplicate and compared alterations in protein expression and phosphorylation levels to mock treated cells using a tandem mass tag (TMT9plex)-based approach. Over 8,100 proteins were quantified across all nine samples of which 46 were altered in abundance upon treatment with nicotine. Proteins with increased abundance included those associated with neurons, defense mechanisms, indicators of pancreatic disease and lysosomal proteins. In addition, we measured differences for ∼16,000 phosphorylation sites across all nine samples using a titanium dioxide-based strategy, of which 132 sites were altered with nicotine and 451 with α-bungarotoxin treatment. Many altered phosphorylation sites were involved in nuclear function and transcriptional events. This study supports the development of future targeted investigations to establish a better understanding for the role of nicotine and associated receptors in pancreatic disease. PMID:26265067

  7. Levels of plasma circulating cell free nuclear and mitochondrial DNA as potential biomarkers for breast tumors

    Directory of Open Access Journals (Sweden)

    Diesch Claude

    2009-11-01

    Full Text Available Abstract Background With the aim to simplify cancer management, cancer research lately dedicated itself more and more to discover and develop non-invasive biomarkers. In this connection, circulating cell-free DNA (ccf DNA seems to be a promising candidate. Altered levels of ccf nuclear DNA (nDNA and mitochondrial DNA (mtDNA have been found in several cancer types and might have a diagnostic value. Methods Using multiplex real-time PCR we investigated the levels of ccf nDNA and mtDNA in plasma samples from patients with malignant and benign breast tumors, and from healthy controls. To evaluate the applicability of plasma ccf nDNA and mtDNA as a biomarker for distinguishing between the three study-groups we performed ROC (Receiver Operating Characteristic curve analysis. We also compared the levels of both species in the cancer group with clinicopathological parameters. Results While the levels of ccf nDNA in the cancer group were significantly higher in comparison with the benign tumor group (P P P P = 0.022. The level of ccf nDNA was also associated with tumor-size (2 cmP = 0.034. Using ROC curve analysis, we were able to distinguish between the breast cancer cases and the healthy controls using ccf nDNA as marker (cut-off: 1866 GE/ml; sensitivity: 81%; specificity: 69%; P P Conclusion Our data suggests that nuclear and mitochondrial ccf DNA have potential as biomarkers in breast tumor management. However, ccf nDNA shows greater promise regarding sensitivity and specificity.

  8. Cytochrome P450 levels are altered in patients with esophageal squamous-cell carcinoma

    Institute of Scientific and Technical Information of China (English)

    I Bergheim; E Wolfgarten; E Bollschweiler; AH H(o)lscher; C Bode; A Parlesak

    2007-01-01

    AIM:To investigate the role of cytochrome P450(CYP)in the carcinogenesis of squamous-cell carcinoma(SCC)in human esophagus by determining expression patterns and protein levels of representative CYPs in esophageal tissue of patients with SCC and controls.METHODS:mRNA expression of CYP2E1,CYP2C,CYP3A4,and CYP3A5 was determined using RT-PCR in both normal and malignant esophageal tissues of patients with untreated esophageal SCC(n = 21)and in controls(n = 10).Protein levels of CYP2E1,CYP2C8,CYP3A4,and CYP3A5 were measured by Western blot.RESULTS:Within the group of SCC patients,mRNA expression of CYP 3A4 and CYP2C was significantly lower in malignant tissue(-39% and -74%,respectively,P < 0.05)than in normal tissue.Similar results were found in CYP3A4 protein levels.Between groups,CYP3A4,CYP3A5,and CYP2C8 protein concentration was significantly higher in non-malignant tissue of SCC patients(4.8-,2.9-,and 1.9-fold elevation,P < 0.05)than in controls.In contrast,CYP2E1 protein levels were significantly higher in controls than in SCC patients (+46%,P < 0.05).CONCLUSION:Significant differences exist in protein levels of certain CYPs in non-malignant esophageal tissue (e.g.CYP2C8,CYP3A4,CYP3A5,and CYP2E1)between SCC patients and healthy subjects and may contribute to the development of SCC in the esophagus.

  9. Nickel toxicity in Pseudomonas tabaci: single cell and bulk sample analysis of bacteria cultured at high cation levels

    International Nuclear Information System (INIS)

    The growth of Pseudomonas tabaci in nutrient medium is partially inhibited in the presence of 10-3 M added nickel (threshold toxic concentration), with complete inhibition at 10-2 M nickel - but no effect at 10sup-4 and 10-5 M. Toxic levels of nickel affect both cell division and cell viability. Spectrophotometric determination of intracellular levels of nickel at different external concentrations showed that the highest internal values occured with cells cultured in 10-4 M (non-toxic) nickel medium rather than in 10-3 (toxic) medium - suggesting that nickel toxicity does not primarily relate to internal concentration. X-ray microanalysis, carried out on whole bacterial cells, showed that toxic levels of nickel in the external medium resulted in a range of ionic changes in the cell, including a decrease in the level of K (K efflux) and an increase in the levels Mn, Fe, Ni, and Cu (transition metal cation influx). Other changes induced by nickel toxicity included an increase in the level of soluble S (with a decrease in insoluble S), an increased cell dry mass, and a conspicuous plasmolysis - which was observed both in whole and in ultrathin sections. The results obtained support a primary toxic effect of nickel at the cell surface - possibly directly affecting the transport activity of the plasmalemma. The resulting changes, particularly involving the influx of a range of cations, may lead to secondary toxic activities affecting the whole metabolism, leading to plasmolysis and inhibition of division. (Author)

  10. Effects of low-level radiation upon the hematopoietic steam cell: implications for leukemogenesis

    International Nuclear Information System (INIS)

    These studies have addressed firstly the effect of single small doses of x-ray upon murine hematopoietic stem cells to obtain a better estimate of the D/sub q/. It is small, of the order of 20 rads. Secondly, a dose fractionation schedule tht does not kill or perturb the kinetics of hemopoietic cell proliferation was sought in order to investigate the leukemogenic potential of low level radiation upon an unperturbed hemopoietic system. The studies reported herein show tht 1.25 rads every other day decrease the CFU-S content of bone marrow by the time 40 rads are accumulated. Studies on the effect of 0.5, 1.0, 2.0, and 3.0 rads 3 times per week are under way. Two rads 3 times per week produced a modest decrease in CFU-S content of bone marrow after an accumulation of 68 rads. With 3.0 rads 3 times per week an accumulation of 102 rads produces a significant decrease in CFU-S content of bone marrow. Dose fractionation at 0.5 and 1.0 rad 3 times per week has not produced a CFU-S depression after accumulation of 17 and 34 rads. Radiation leukemogenesis studies published to date have utilized single doses and chronic exposure schedules that probably have significantly perturbed the kinetics of hematopoietic stem cells. Whether radiation will produce leukemia in animal models with dose schedules that do not perturb kinetics of hematopoietic stem cells remains to be seen

  11. Effects of low-level radiation upon the hematopoietic steam cell: implications for leukemogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Cronkite, E.P.; Bond, V.P.; Carsten, A.L.; Miller, M.E.; Bullis, J.E.

    1983-01-01

    These studies have addressed firstly the effect of single small doses of x-ray upon murine hematopoietic stem cells to obtain a better estimate of the D/sub q/. It is small, of the order of 20 rads. Secondly, a dose fractionation schedule tht does not kill or perturb the kinetics of hemopoietic cell proliferation was sought in order to investigate the leukemogenic potential of low level radiation upon an unperturbed hemopoietic system. The studies reported herein show tht 1.25 rads every other day decrease the CFU-S content of bone marrow by the time 40 rads are accumulated. Studies on the effect of 0.5, 1.0, 2.0, and 3.0 rads 3 times per week are under way. Two rads 3 times per week produced a modest decrease in CFU-S content of bone marrow after an accumulation of 68 rads. With 3.0 rads 3 times per week an accumulation of 102 rads produces a significant decrease in CFU-S content of bone marrow. Dose fractionation at 0.5 and 1.0 rad 3 times per week has not produced a CFU-S depression after accumulation of 17 and 34 rads. Radiation leukemogenesis studies published to date have utilized single doses and chronic exposure schedules that probably have significantly perturbed the kinetics of hematopoietic stem cells. Whether radiation will produce leukemia in animal models with dose schedules that do not perturb kinetics of hematopoietic stem cells remains to be seen.

  12. Functionalized silica nanoparticles: a platform for fluorescence imaging at the cell and small animal levels.

    Science.gov (United States)

    Wang, Kemin; He, Xiaoxiao; Yang, XiaoHai; Shi, Hui

    2013-07-16

    Going in vivo, including living cells and the whole body, is very important for gaining a better understanding of the mystery of life and requires specialized imaging techniques. The diversity, composition, and temporal-spatial variation of life activities from cells to the whole body require the analysis techniques to be fast-response, noninvasive, highly sensitive, and stable, in situ and in real-time. Functionalized nanoparticle-based fluorescence imaging techniques have the potential to meet such needs through real-time and noninvasive visualization of biological events in vivo. Functionalized silica nanoparticles (SiNPs) doped with fluorescent dyes appear to be an ideal and flexible platform for developing fluorescence imaging techniques used in living cells and the whole body. We can select and incorporate different dyes inside the silica matrix either noncovalently or covalently. These form the functionalized hybrid SiNPs, which support multiplex labeling and ratiometric sensing in living systems. Since the silica matrix protects dyes from outside quenching and degrading factors, this enhances the photostability and biocompatibility of the SiNP-based probes. This makes them ideal for real-time and long-time tracking. One nanoparticle can encapsulate large numbers of dye molecules, which amplifies their optical signal and temporal-spatial resolution response. Integrating fluorescent dye-doped SiNPs with targeting ligands using various surface modification techniques can greatly improve selective recognition. Along with the endocytosis, functionalized SiNPs can be efficiently internalized into cells for noninvasive localization, assessment, and monitoring. These unique characteristics of functionalized SiNPs substantially support their applications in fluorescence imaging in vivo. In this Account, we summarize our efforts to develop functionalized dye-doped SiNPs for fluorescence imaging at the cell and small animal levels. We first discuss how to design and

  13. Plasma IFN-γ and IL-6 levels correlate with peripheral T-cell numbers but not toxicity in RCC patients treated with CAR T-cells.

    Science.gov (United States)

    Klaver, Yarne; van Steenbergen, Sabine C L; Sleijfer, Stefan; Debets, Reno; Lamers, Cor H J

    2016-08-01

    Autologous T-cells genetically modified to express a chimeric antigen receptor (CAR) against carboxy-anhydrase-IX (CAIX) were administered to twelve patients with CAIX-positive metastatic renal cell carcinoma. Here, we questioned whether plasma cytokine levels following treatment or in vitro cytokine production from the T-cell infusion products could serve as predictors for peripheral T-cell persistence or in vivo T-cell activity. We demonstrated that CAR surface as well as gene expression are down-regulated following T-cell infusion, and that peripheral numbers of CAR T-cells are best captured by flow cytometry and not by qPCR. Numbers of CAR T-cells in blood correlated with plasma levels of IFN-γ and IL-6, but not with any of the other cytokines tested. Plasma IFN-γ or IL-6 levels did not correlate with liver enzyme values. Thus, out of 27 cytokines tested, IFN-γ and IL-6 levels in plasma are potential surrogate markers for CAR T-cell persistence in solid tumors. PMID:27377533

  14. 氯胺酮对缺氧诱导胎鼠大脑皮层神经元特异性烯醇化酶的影响%Effects of ketamine on anoxia induced neurone specific enolase release from cerebral cortex neurons of fetal rats

    Institute of Scientific and Technical Information of China (English)

    王惠军; 薛玉良; 李兰英; 高春霖

    2011-01-01

    目的 探讨氯胺酮对神经元缺氧损伤的防护作用及机制.方法 应用无血清方法原代培养胎鼠大脑神经细胞,进行缺氧结合无糖处理,应用酶联免疫吸附试验测定缺氧前后细胞内外神经元特异性烯醇化酶(NSE)浓度,并应用逆转录聚合酶链反应观察缺氧前后细胞内NSE mRNA表达量的变化.结果 随着缺氧时间的延长,细胞外NSE浓度明显升高、细胞内降低(P<0.01),至缺氧5 h达高峰.缺氧1 h组缺氧前给予氯胺酮,随药物浓度的增加,细胞外NSE浓度逐渐降低[缺氧1 h组(4.95±0.41)μg/L,缺氧1 h+氯胺酮1μmol/L组(4.43±0.36)μg/L,缺氧I h+氯胺酮20 μmol/L组(3.53±0.46)μg/L,缺氧1 h+氯胺酮100 μmol/L组(3.17±0.23)μg/L]、细胞内逐渐升高[缺氧1 h组(22.0 ±0.76)μg/L,缺氧1 h+氯胺酮1 μmol/L组(24.7±0.84)μg/L,缺氧1 h+氯胺酮20μmoL/L组(28.8±0.91)μg/L,缺氧1 h+氯胺酮100 μmol/L(33.7±0.92)μg/L],呈剂量依赖性,氯胺酮100 μmol/L时接近对照组水平.随着缺氧时间延长,NSE mRNA相对表达量明显增高(对照组NSE相对表达量0.153±0.007,缺氧0.5 h组0.654±0.012.P<0.01),至1 h达高峰(NES相对表达量0.923±0.015);缺氧0.5 h组缺氧前给予患者不同浓度(1、20 μmol/L)氯胺酮后,NSE的表达量明显降低(NSE相对表达量分别为0.531±0.011、0.283±0.009,P<0.01).结论 氯胺酮可通过防止细胞膜的损伤,减少细胞内酶的外漏,并在基因表达和蛋白合成两个阶段对NSE进行调节,发挥对体外培养神经元缺氧性损伤的防护作用.%Objective To evaluate the effects of ketamine on neurone specific enolase (NSE) in combined oxygen-glucose deprivation of embryonic rat cortical neurons. Methods The mixed cell culture riching in neurons were deprived both oxygen and glucose. Using ELISA and RT-PCR method to assay the changes of intracellular and extracellular concentrations of NSE and the expression of NSE mRNA in hypoxic neurons treated with different

  15. Live-Cell Imaging Tool Optimization To Study Gene Expression Levels and Dynamics in Single Cells of Bacillus cereus

    NARCIS (Netherlands)

    Eijlander, Robyn T.; Kuipers, Oscar P.

    2013-01-01

    Single-cell methods are a powerful application in microbial research to study the molecular mechanism underlying phenotypic heterogeneity and cell-to-cell variability. Here, we describe the optimization and application of single-cell time-lapse fluorescence microscopy for the food spoilage bacterium

  16. Influence of βS-globin haplotypes and hydroxyurea on tumor necrosis factor-alpha levels in sickle cell anemia

    OpenAIRE

    Marília Rocha Laurentino; Pedro Aurio Maia Filho; Maritza Cavalcante Barbosa; Izabel Cristina Justino Bandeira; Lilianne Brito da Silva Rocha; Romelia Pinheiro Gonçalves

    2014-01-01

    Background: Sickle cell anemia is a chronic inflammatory disease characterized by an increased production of proinflammatory cytokines including tumor necrosis factor-alpha. Hydroxyurea, by decreasing the polymerization of hemoglobin, reduces inflammatory states. The effect of the genetic polymorphisms of sickle cell patients on tumor necrosis factor-alpha levels remains unknown. Objective: The aim of this study was to investigate the association of tumor necrosis factor-alpha levels with β...

  17. Patients with pancreatic adenocarcinoma exhibit elevated levels of myeloid-derived suppressor cells upon progression of disease

    OpenAIRE

    Markowitz, Joseph; Brooks, Taylor R.; Duggan, Megan C.; Paul, Bonnie K.; Pan, Xueliang; Wei, Lai; Abrams, Zachary; Luedke, Eric; Lesinski, Gregory B; Mundy-Bosse, Bethany; Bekaii-Saab, Tanios; Carson, William E

    2014-01-01

    Elevated levels of myeloid-derived suppressor cells (MDSCs) induced by tumor-derived factors are associated with inhibition of immune responses in patients with gastrointestinal malignancies. We hypothesized that pro-MDSC cytokines and levels of MDSC in the peripheral blood would be elevated in pancreatic adenocarcinoma patients with progressive disease. Peripheral blood mononuclear cells (PBMCs) were isolated from 16 pancreatic cancer patients undergoing chemotherapy and phenotyped for MDSC ...

  18. Microsphere-Incorporated Hybrid Thermogel for Neuronal Differentiation of Tonsil Derived Mesenchymal Stem Cells.

    Science.gov (United States)

    Patel, Madhumita; Moon, Hyo Jung; Jung, Bo Kyung; Jeong, Byeongmoon

    2015-07-15

    Neuronal differentiation of tonsil-derived mesenchymal stem cells (TMSCs) is investigated in a 3D hybrid system. The hybrid system is prepared by increasing the temperature of poly(ethylene glycol)-poly(l-alanine) aqueous solution to 37 °C through the heat-induced sol-to-gel transition, in which TMSCs and growth factor releasing microspheres are suspended. The in situ formed gel exhibits a modulus of 800 Pa at 37 °C, similar to that of brain tissue, and it is robust enough to hold the microspheres and cells during the 3D culture of TMSCs. The neuronal growth factors are released over 12-18 d, and the TMSCs in a spherical shape initially undergo multipolar elongation during the 3D culture. Significantly higher expressions of the neuronal biomarkers such as nuclear receptor related protein (Nurr-1), neuron specific enolase, microtubule associated protein-2, neurofilament-M, and glial fibrillary acidic protein are observed in both mRNA level and protein level in the hybrid systems than in the control experiments. This study proves the significance of a controlled drug delivery concept in tissue engineering or regenerative medicine, and a 3D hybrid system with controlled release of growth factors from microspheres in a thermogel can be a very promising tool. PMID:26033880

  19. Effects of Salvianolic Acid B on Protein Expression in Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Tsong-Min Chang

    2011-01-01

    Full Text Available Salvianolic acid B (Sal B, a pure water-soluble compound extracted from Radix Salviae miltiorrhizae, has been reported to possess potential cardioprotective efficacy. To identify proteins or pathways by which Sal B might exert its protective activities on the cardiovascular system, two-dimensional gel electrophoresis-based comparative proteomics was performed, and proteins altered in their expression level after Sal B treatment were identified by MALDI-TOF MS/MS. Human umbilical vein endothelial cells (HUVECs were incubated at Sal B concentrations that can be reached in human plasma by pharmacological intervention. Results indicated that caldesmon, an actin-stabilizing protein, was downregulated in Sal B-exposed HUVECs. Proteins that showed increased expression levels upon Sal B treatment were vimentin, T-complex protein 1, protein disulfide isomerase, tropomyosin alpha, heat shock protein beta-1, UBX domain-containing protein 1, alpha enolase, and peroxiredoxin-2. Additionally, Sal B leads to increased phosphorylation of nucleophosmin in a dose-dependent manner and promotes proliferation of HUVECs. We found that Sal B exhibited a coordinated regulation of enzymes and proteins involved in cytoskeletal reorganization, oxidative stress, and cell growth. Our investigation would provide understanding to the endothelium protection information of Sal B.

  20. Decreased Circulating Interleukin-35 Levels Are Related to Interleukin-4-Producing CD8+ T Cells in Patients with Allergic Asthma.

    Science.gov (United States)

    Wang, Wei; Li, Ping; Yang, Jiong

    2015-08-01

    Interleukin (IL)-35 is a newly discovered suppressive cytokine and has been shown to alleviate inflammatory and autoimmune diseases. The purpose of this study was to investigate immunomodulatory capacity of IL-35 in patients with allergic asthma. IL-35 mRNA expression levels in peripheral blood mononuclear cells (PBMCs) were detected by quantitative real-time PCR (qPCR). The frequencies of cytotoxic T cells (Tc)1, Tc2 and Tc17 cells were measured by flow cytometry. Plasma levels of IL-35, interferon (IFN)-γ, IL-4, and IL-17 were examined by enzyme-linked immunosorbent assay (ELISA). The correlations between plasma IL-35 levels and Tc1, Tc2, and Tc17 cytokine production in allergic asthmatics (n = 25) and healthy controls (n = 12) were analyzed by Pearson's test. IL-35 protein and mRNA expression levels were down-regulated in allergic asthmatics compared with healthy controls. The frequencies of Tc2 and Tc17 cells were significantly increased in patients with asthma, and the frequency of Tc1 cells did not differ between asthmatic patients and healthy controls. Similarly, plasma levels of IL-4 and IL-17 were significantly increased in asthmatic patients, while there was no difference in IFN-γ levels between allergic asthma patients and healthy controls. More importantly, plasma IL-35 protein levels were negatively correlated with the frequency of IL-4-producing CD8+ T (Tc2) cells and with the IL-4 level in patients with allergic asthma. Our results suggest that decreased circulating IL-35 levels could contribute to the pathogenesis of allergic asthma by regulating CD8+ T cells. PMID:26547705

  1. Ethacrynic acid inhibition of histamine release from rat mast cells: effect on cellular ATP levels and thiol groups

    DEFF Research Database (Denmark)

    Johansen, Torben

    1983-01-01

    The experiments concerned the effect of ethacrynic acid (0.5 mM) on the adenosine triphosphate (ATP) content of rat mast cells and the effect on histamine release induced by the ionophore A23187 (10 microM). Ethacrynic acid decreased the ATP level of the cells in presence of antimycin A and glucose...

  2. The production of VEGF involving MAP kinase activation by low level laser therapy in human granulosa cells

    OpenAIRE

    Kawano, Yasushi; Utsunomiya-Kai, Yufuko; Kai, Kentaro; Miyakawa, Isao; Ohshiro, Toshio; Narahara, Hisashi

    2012-01-01

    Objective: The function of granulosa cells is regulated by various hormones and growth factors. Our aim is to clarify the regulation of vascular endothelial growth factor (VEGF) production via mitogen-activated protein kinase (MAPK) induced by low level laser therapy (LLLT) in human granulosa cells.

  3. Levels of synthesis of primate-specific nuclear proteins differ between growth-arrested and proliferating cells

    International Nuclear Information System (INIS)

    A monoclonal antibody that reacts specifically with the proliferation-sensitive nuclear proteins, isoelectric focusing (IEF) 8Z31 (molecular weight (MW), 76,000 charge variants, HeLa protein catalogue number) has been characterized. As determined by indirect immunofluorescence, the antibody stains the nucleolus and nucleoplasm of interphase-cultured cells of primate origin, but does not react with cells of other species. Proteins having similar MWs and isoelectric points as the human or monkey (primates) proteins were not observed in cultured cells of the following species: aves, bat, dog, dolphin, goat, hamster, mink, mouse, pisces, potoroo, rabbit and rat. Quantitative two-dimensional (2D) gel electrophoretic analysis of [35S]methionine-labelled proteins synthesized by normal (quiescent, proliferating) and SV40-transformed human MRC-5 fibroblasts revealed significant differences in the levels of synthesis of both IEF 8Z30 and 8Z31. In quiescent cells the main labelled product corresponded to IEF 8Z31 (ratio IEF 8Z31/8Z30, 2.3), while in the transformed cells the major product was IEF 8Z30 (ratio, 0.62). Normal proliferating fibroblasts exhibited similar levels of both proteins (ratio, 1.21). Combined levels of synthesis of both proteins were 1.50 and 1.20 times as high in the transformed cells as in the quiescent and proliferating cells, respectively. Modulation of the levels of synthesis of these proteins may play a role in cell proliferation

  4. Endothelial adhesion of synchronized gastric tumor cells changes during cell cycle transit and correlates with the expression level of CD44 splice variants

    Institute of Scientific and Technical Information of China (English)

    Anton Oertl; Jens Castein; Tobias Engl; Wolf-Dietrich Beecken; Dietger Jonas; Richard Melamed; Roman A. Blaheta

    2005-01-01

    AIM: To study adhesion capacity and CD44 expression of human gastric adenocarcinoma MKN45 cells at different stages of a first cell cycle.METHODS: MKN45 cells were synchronized by aphidicolin and assayed for adhesion to an endothelial cell (HUVEC)monolayer. Surface expression of CD44 and CD44 splice variants on MKN45 cells was evaluated by flow cytometry.Functional relevance of CD44 adhesion receptors was investigated by blocking studies using anti CD44 monoclonal antibodies or by hyaluronan digestion.RESULTS: Adhesion of MKN45 to HUVEC was increased during G2/M transit, after which adhesion returned to baseline levels with cell cycle completion. In parallel, CD44splice variants CD44v4, CD44v5, and CD44v7 were all upregulated on MKN45 during cell cycle progression with a maximum effect in G2/M. The function of CD44 surface receptors was assessed with specific receptor blocking monodonal antibodies or removal of hyaluronan by digestion with hyaluronidase. Both strategies inhibited tumor cell adhesion to HUVEC by nearly 50%, which indicates that MKN45-HUVEC-interaction is CD44 dependent.CONCLUSION: CD44 expression level is linked to the cell cycle in gastrointestinal tumor cells, which in turn leads to cell cyde dependent alterations of their adhesion behaviour to endothelium.

  5. A low threshold level of expression of mutant-template telomerase RNA inhibits human tumor cell proliferation

    OpenAIRE

    Kim, Moses M.; Rivera, Melissa A.; Botchkina, Inna L.; Shalaby, Refaat; Thor, Ann D; Elizabeth H. Blackburn

    2001-01-01

    The ribonucleoprotein telomerase synthesizes telomeric DNA by copying an intrinsic RNA template. In most cancer cells, telomerase is highly activated. Here we report a telomerase-based antitumor strategy: expression of mutant-template telomerase RNAs in human cancer cells. We expressed mutant-template human telomerase RNAs in prostate (LNCaP) and breast (MCF-7) cancer cell lines. Even a low threshold level of expression of telomerase RNA gene constructs containing various mutant templates, bu...

  6. Analysis of Thalassiosira pseudonana Silicon Transporters Indicates Distinct Regulatory Levels and Transport Activity through the Cell Cycle▿

    OpenAIRE

    Thamatrakoln, Kimberlee; Hildebrand, Mark

    2006-01-01

    An analysis of the expression and activity of silicon transporters (SITs) was done on synchronously growing cultures of the diatom Thalassiosira pseudonana to provide insight into the role these proteins play in cellular silicon metabolism during the cell cycle. The first SIT-specific polyclonal peptide antibody was generated and used in the immunoblot analysis of whole-cell protein lysates to monitor SIT protein levels during synchronized progression through the cell cycle. Peaks in SIT prot...

  7. Endothelin and a Ca2+ ionophore raise cyclic GMP levels in a neuronal cell line via formation of nitric oxide.

    OpenAIRE

    Reiser, G.

    1990-01-01

    1. The vasoconstrictor peptide endothelin-1 caused a fast, transient rise in guanosine 3':5'-cyclic monophosphate (cyclic GMP) levels in a neuronal cell line (mouse neuroblastoma x rat glioma hybrid cells 108CC15). The mechanism of activation of guanylate cyclase by endothelin-1 was investigated. The endothelin-1-induced rise depended on the release of internal Ca2+. 2. The stimulation of cyclic GMP synthesis induced by endothelin-1 was suppressed after preincubating the cells in medium conta...

  8. ATP consumption of eukaryotic flagella measured at a single-cell level

    CERN Document Server

    Chen, Daniel T N; Fraden, Seth; Nicastro, Daniela; Dogic, Zvonimir

    2015-01-01

    The motility of cilia and flagella is driven by thousands of dynein motors that hydrolyze adenosine triphosphate (ATP). Despite decades of genetic, biochemical, structural and biophysical studies, some aspects of ciliary motility remain elusive, such as the regulation of beating patterns and the energetic efficiency of these nanomachines. Here, we introduce an experimental method to measure ATP consumption of actively beating axonemes on a single-cell level. We encapsulated individual sea urchin sperm with demembranated flagellum inside water-in-oil emulsion droplets and measured the axonemes ATP consumption by monitoring fluorescence intensity of a fluorophore-coupled reporter system for ATP turnover in the droplet. Concomitant phase contrast imaging allowed us to extract a linear dependence between the ATP consumption rate and the flagellar beating frequency, with ~2.3e5 ATP molecules consumed per beat of a demembranated flagellum. Increasing the viscosity of the aqueous medium led to modified beating wavef...

  9. Preconditioning Serum Levels of Endothelial Cell-Derived Molecules and the Risk of Posttransplant Complications in Patients Treated with Allogeneic Stem Cell Transplantation

    Directory of Open Access Journals (Sweden)

    Roald Lindås

    2014-01-01

    Full Text Available Endothelial cells are involved in the pathogenesis of acute graft-versus-host disease (GVHD after allogeneic stem cell transplantation. These cells express several molecules that can be detected as biologically active soluble forms; serum levels of these molecules may thereby reflect the functional status of endothelial cells. Furthermore, acute GVHD is an inflammatory reaction and endothelial cells function as local regulators of inflammation. We therefore investigated whether differences in preconditioning/pretransplant serum levels of endothelium-expressed molecules (i.e., endocan, vascular cell adhesion molecule 1 (VCAM-1, and E-selectin were associated with a risk of posttransplant GVHD. Our study should be regarded as a population-based study of consecutive and thereby unselected patients (n=56. Analysis of this pretreatment endothelium biomarker profile by unsupervised hierarchical clustering identified a subset of patients with increased early nonrelapse mortality. Furthermore, low endocan levels were significantly associated with acute GVHD in the liver and gastrointestinal tract, whereas high VCAM-1 levels were associated with acute GVHD in the skin only. Our study suggests that the preconditioning/pretransplant status of endothelial cells (possibly through altered trafficking of immunocompetent cells is important for the risk and the organ involvement of later acute GVHD.

  10. Elevated lead levels and adverse effects on natural killer cells in children from an electronic waste recycling area.

    Science.gov (United States)

    Zhang, Yu; Huo, Xia; Cao, Junjun; Yang, Tian; Xu, Long; Xu, Xijin

    2016-06-01

    Lead (Pb) has been proved to exert immunotoxicity to influence immune homeostasis in humans. To monitor the internal Pb level and evaluate its effect on natural killer (NK) cells and cytokine/chemokine concentrations, we recruited 285 preschool children from Guiyu, one of the largest electronic waste (e-waste) destinations and recycling areas in the world, and known to have high concentrations of Pb in the air, soil, water, sediment and plants. A total of 126 preschool children were selected from Haojiang as a reference group. Results showed that children in Guiyu, the exposed area, had higher blood Pb levels and lower percentages of NK cells than children from the reference area. A significantly negative association was found between the percentage of NK cells and increasing Pb levels. Moreover, children in Guiyu area had higher platelet counts and IL-1β concentrations, and lower levels of IL-2, IL-27, MIP-1α and MIP-1β were observed in the exposed children. These changes might not be conducive to the development and differentiation of NK cells. Taken together, the elevated Pb levels result in the lower percentages of NK cells, but also alter the levels of platelets, IL-1β and IL-27, which might be unconducive to the activity and function of NK cells. PMID:26895538

  11. High level of chromosomal instability in circulating tumor cells of ROS1-rearranged non-small-cell lung cancer

    Science.gov (United States)

    Pailler, E.; Auger, N.; Lindsay, C. R.; Vielh, P.; Islas-Morris-Hernandez, A.; Borget, I.; Ngo-Camus, M.; Planchard, D.; Soria, J.-C.; Besse, B.; Farace, F.

    2015-01-01

    Background Genetic aberrations affecting the c-ros oncogene 1 (ROS1) tyrosine kinase gene have been reported in a small subset of patients with non-small-cell lung cancer (NSCLC). We evaluated whether ROS1-chromosomal rearrangements could be detected in circulating tumor cells (CTCs) and examined tumor heterogeneity of CTCs and tumor biopsies in ROS1-rearranged NSCLC patients. Patients and methods Using isolation by size of epithelial tumor cells (ISET) filtration and filter-adapted-fluorescence in situ hybridization (FA-FISH), ROS1 rearrangement was examined in CTCs from four ROS1-rearranged patients treated with the ROS1-inhibitor, crizotinib, and four ROS1-negative patients. ROS1-gene alterations observed in CTCs at baseline from ROS1-rearranged patients were compared with those present in tumor biopsies and in CTCs during crizotinib treatment. Numerical chromosomal instability (CIN) of CTCs was assessed by DNA content quantification and chromosome enumeration. Results ROS1 rearrangement was detected in the CTCs of all four patients with ROS1 rearrangement previously confirmed by tumor biopsy. In ROS1-rearranged patients, median number of ROS1-rearranged CTCs at baseline was 34.5 per 3 ml blood (range, 24–55). In ROS1-negative patients, median background hybridization of ROS1-rearranged CTCs was 7.5 per 3 ml blood (range, 7–11). Tumor heterogeneity, assessed by ROS1 copy number, was significantly higher in baseline CTCs compared with paired tumor biopsies in the three patients experiencing PR or SD (P < 0.0001). Copy number in ROS1-rearranged CTCs increased significantly in two patients who progressed during crizotinib treatment (P < 0.02). CTCs from ROS1-rearranged patients had a high DNA content and gain of chromosomes, indicating high levels of aneuploidy and numerical CIN. Conclusion We provide the first proof-of-concept that CTCs can be used for noninvasive and sensitive detection of ROS1 rearrangement in NSCLC patients. CTCs from ROS1-rearranged

  12. Cholinergic neuronal differentiation of bone marrow mesenchymal stem cells in rhesus monkeys

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    The purpose of the present study was to determine the best cholinergic neuronal differentiation method of rhesus monkey bone marrow mesenchymal stem cells(BMSCs).Four methods were used to induce differentiation,and the groups were assigned accordingly:basal inducing group(culture media,bFGF,and forskolin);SHH inducing group(SHH,inducing group);RA inducing group(RA,basal inducing group);and SHH+RA inducing group(SHH,RA,and basal inducing group).All groups displayed neuronal morphology and increased expression of nestin and neuron-specific enolase.The basal inducing group did not express synapsin,and cells from the SHH inducing group did not exhibit neuronal resting membrane potential.In contrast,results demonstrated that BMSCs from the RA and SHH+RA inducing groups exhibited neuronal resting membrane potential,and cells from the SHH+RA inducing group expressed higher levels of synapsin and acetylcholine.In conclusion,the induction of cholinergic differentiation through SHH+RA was determined to be superior to the other methods.

  13. Auxins differentially regulate root system architecture and cell cycle protein levels in maize seedlings.

    Science.gov (United States)

    Martínez-de la Cruz, Enrique; García-Ramírez, Elpidio; Vázquez-Ramos, Jorge M; Reyes de la Cruz, Homero; López-Bucio, José

    2015-03-15

    Maize (Zea mays) root system architecture has a complex organization, with adventitious and lateral roots determining its overall absorptive capacity. To generate basic information about the earlier stages of root development, we compared the post-embryonic growth of maize seedlings germinated in water-embedded cotton beds with that of plants obtained from embryonic axes cultivated in liquid medium. In addition, the effect of four different auxins, namely indole-3-acetic acid (IAA), 1-naphthaleneacetic acid (NAA), indole-3-butyric acid (IBA) and 2,4-dichlorophenoxyacetic acid (2,4-D) on root architecture and levels of the heat shock protein HSP101 and the cell cycle proteins CKS1, CYCA1 and CDKA1 were analyzed. Our data show that during the first days after germination, maize seedlings develop several root types with a simultaneous and/or continuous growth. The post-embryonic root development started with the formation of the primary root (PR) and seminal scutellar roots (SSR) and then continued with the formation of adventitious crown roots (CR), brace roots (BR) and lateral roots (LR). Auxins affected root architecture in a dose-response fashion; whereas NAA and IBA mostly stimulated crown root formation, 2,4-D showed a strong repressing effect on growth. The levels of HSP101, CKS1, CYCA1 and CDKA in root and leaf tissues were differentially affected by auxins and interestingly, HSP101 registered an auxin-inducible and root specific expression pattern. Taken together, our results show the timing of early branching patterns of maize and indicate that auxins regulate root development likely through modulation of the HSP101 and cell cycle proteins. PMID:25615607

  14. ATP Consumption of Eukaryotic Flagella Measured at a Single-Cell Level.

    Science.gov (United States)

    Chen, Daniel T N; Heymann, Michael; Fraden, Seth; Nicastro, Daniela; Dogic, Zvonimir

    2015-12-15

    The motility of cilia and flagella is driven by thousands of dynein motors that hydrolyze adenosine triphosphate (ATP). Despite decades of genetic, biochemical, structural, and biophysical studies, some aspects of ciliary motility remain elusive, such as the regulation of beating patterns and the energetic efficiency of these nanomachines. In this study, we introduce an experimental method to measure ATP consumption of actively beating axonemes on a single-cell level. We encapsulated individual sea urchin sperm with demembranated flagellum inside water-in-oil emulsion droplets and measured the axoneme's ATP consumption by monitoring fluorescence intensity of a fluorophore-coupled reporter system for ATP turnover in the droplet. Concomitant phase contrast imaging allowed us to extract a linear dependence between the ATP consumption rate and the flagellar beating frequency, with ∼2.3 × 10(5) ATP molecules consumed per beat of a demembranated flagellum. Increasing the viscosity of the aqueous medium led to modified beating waveforms of the axonemes and to higher energy consumption per beat cycle. Our single-cell experimental platform provides both new insights, to our knowledge, into the beating mechanism of flagella and a powerful tool for future studies. PMID:26682814

  15. Emerging methods to study bacteriophage infection at the single-cell level.

    Science.gov (United States)

    Dang, Vinh T; Sullivan, Matthew B

    2014-01-01

    Bacteria and their viruses (phages) are abundant across diverse ecosystems and their interactions influence global biogeochemical cycles and incidence of disease. Problematically, both classical and metagenomic methods insufficiently assess the host specificity of phages and phage-host infection dynamics in nature. Here we review emerging methods to study phage-host interaction and infection dynamics with a focus on those that offer resolution at the single-cell level. These methods leverage ever-increasing sequence data to identify virus signals from single-cell amplified genome datasets or to produce primers/probes to target particular phage-bacteria pairs (digital PCR and phageFISH), even in complex communities. All three methods enable study of phage infection of uncultured bacteria from environmental samples, while the latter also discriminates between phage-host interaction outcomes (e.g., lytic, chronic, lysogenic) in model systems. Together these techniques enable quantitative, spatiotemporal studies of phage-bacteria interactions from environmental samples of any ecosystem, which will help elucidate and predict the ecological and evolutionary impacts of specific phage-host pairings in nature. PMID:25566233

  16. Emerging methods to study bacteriophage infection at the single-cell level

    Directory of Open Access Journals (Sweden)

    Vinh Toan Dang

    2014-12-01

    Full Text Available Bacteria and their viruses (phages are abundant across diverse ecosystems and their interactions influence global biogeochemical cycles and incidence of disease. Problematically, both classical and metagenomic methods insufficiently assess the host specificity of phages and phage–host infection dynamics in nature. Here we review emerging methods to study phage–host interaction and infection dynamics with a focus on those that offer resolution at the single-cell level. These methods leverage ever-increasing sequence data to identify virus signals from single-cell amplified genome (SAG datasets or to produce primers/probes to target particular phage– bacteria pairs (digital PCR and phageFISH, even in complex communities. All three methods enable study of phage infection of uncultured bacteria from environmental samples, while the latter also discriminates between phage–host interaction outcomes (e.g., lytic, chronic, lysogenic in model systems. Together these techniques enable quantitative, spatiotemporal studies of phage–bacteria interactions from environmental samples of any ecosystem, which will help elucidate and predict the ecological and evolutionary impacts of specific phage–host pairings in nature.

  17. Effects of temperature and humidification levels on the performance of a proton exchange membrane fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Chiang, M-S. [Nan Kai Institute of Technology, Nantou, Taiwan (China). Department of Mechanical Engineering; Chu, H-S. [National Chiao Tung University, Hsinchu, Taiwan (China). Department of Mechanical Engineering

    2006-07-01

    Numerical investigation of a proton exchange membrane fuel cell performance subjected to various humidification and thermal conditions is the focus of this study. Governing equations describing species, mass, momentum, and enthalpy conservation are employed and solved by a computational fluid dynamic algorithm to obtain domain's physical properties and cell performance. The model accounts for electrochemical kinetic as well as two-phase flow with phase change and water transport. Numerical prediction results use polarization curves and contour plots to illustrate the effects of various humidification schemes and temperature gradient scenarios. Findings show that humidification-level perturbation on the anode or cathode side creates different effects. Mechanisms influencing performance-variation tendencies are interpreted. In addition modelling results with existing temperature gradient exhibit different trends on the overpotentials depending on the slope and magnitude. At higher cathode temperature, it is shown that polarization curves are dominated by reaction kinetics and membrane water content at medium and high reaction rates, respectively. (author)

  18. ATP Consumption of Eukaryotic Flagella Measured at a Single-Cell Level

    Science.gov (United States)

    Chen, Daniel T. N.; Heymann, Michael; Fraden, Seth; Nicastro, Daniela; Dogic, Zvonimir

    2015-12-01

    The motility of cilia and flagella is driven by thousands of dynein motors that hydrolyze adenosine triphosphate (ATP). Despite decades of genetic, biochemical, structural and biophysical studies, some aspects of ciliary motility remain elusive, such as the regulation of beating patterns and the energetic efficiency of these nanomachines. Here, we introduce an experimental method to measure ATP consumption of actively beating axonemes on a single-cell level. We encapsulated individual sea urchin sperm with demembranated flagellum inside water-in-oil emulsion droplets and measured the axonemes ATP consumption by monitoring fluorescence intensity of a fluorophore-coupled reporter system for ATP turnover in the droplet. Concomitant phase contrast imaging allowed us to extract a linear dependence between the ATP consumption rate and the flagellar beating frequency, with ~2.3e5 ATP molecules consumed per beat of a demembranated flagellum. Increasing the viscosity of the aqueous medium led to modified beating waveforms of the axonemes and to higher energy consumption per beat cycle. Our single-cell experimental platform provides both new insights into the beating mechanism of flagella and a powerful tool for future studies.

  19. Optimization of an HPLC Method for Determining the Genomic Methylation Levels of Taxus Cells.

    Science.gov (United States)

    Li, Xiao-li; Yuan, Jie; Dong, Yan-shan; Fu, Chun-hua; Li, Mao-Teng; Yu, Long-jiang

    2016-02-01

    An HPLC method for quantifying total DNA methylation in Taxus chinensis cells is described. Optimal conditions for the method were established as follows: DNA was hydrolyzed with DNA degradase at 37°C for 3 h. The mobile phase was a mixture of Solvent A [50 mM potassium dihydrogen phosphate/triethylamine (100:0.2, v/v)] and Solvent B (methanol); the gradient was 10% (v/v) solvent B. The calibration curves for deoxycytidine monophosphate (dCMP) and methylated dCMP were linear within 1.0-160.0 µg mL(-1), with correlation coefficients of 0.9996 and 0.9998. The limits of detection for dCMP and 5-mdCMP were 0.482 and 0.301 ng mL(-1), respectively, and the limits of quantification were 1.6 and 1.0 ng mL(-1), respectively. The method has been validated according to the current International Conference Harmonization guidelines. The method was able to quantify the content of dCMP and methylated dCMP specifically, accurately and precisely. The global DNA methylation level in different Taxus cells was measured using as little as 3 µg of DNA according to the optimized procedure. In addition, degradation of 5-methylcytosine was prevented. PMID:26341490

  20. Tetracycline-inducible protein expression in pancreatic cancer cells: Effects of CapG overexpression

    Institute of Scientific and Technical Information of China (English)

    Sarah Tonack; Sabina Patel; Mehdi Jalali; Taoufik Nedjadi; Rosalind E Jenkins; Christopher Goldring; John Neoptolemos; Eithne Costello

    2011-01-01

    AIM: To establish stable tetracycline-inducible pancre-atic cancer cell lines.METHODS: Suit-2, MiaPaca-2, and Panc-1 cells were transfected with a second generation reverse tetra-cycline-controlled transactivator protein (rtTA2S-M2), under the control of either a cytomegalovirus (CMV) or a chicken β-actin promoter, and the resulting clones were characterised.RESULTS: Use of the chicken (β-actin) promoter proved superior for both the production and mainte-nance of doxycycline-inducible cell lines. The system proved versatile, enabling transient inducible expression of a variety of genes, including GST-P, CYP2E1, S100A6, and the actin capping protein, CapG. To determine the physiological utility of this system in pancreatic cancer cells, stable inducible CapG expressors were established. Overexpressed CapG was localised to the cytoplasm and the nuclear membrane, but was not observed in the nu-cleus. High CapG levels were associated with enhanced motility, but not with changes to the cell cycle, or cellu-lar proliferation. In CapG-overexpressing cells, the levels and phosphorylation status of other actin-moduating proteins (Cofilin and Ezrin/Radixin) were not altered. However, preliminary analyses suggest that the levels of other cellular proteins, such as ornithine aminotransfer-ase and enolase, are altered upon CapG induction. CONCLUSION: We have generated pancreatic-cancer derived cell lines in which gene expression is fully con-trollable.

  1. Melanogenesis stimulation in B16-F10 melanoma cells induces cell cycle alterations, increased ROS levels and a differential expression of proteins as revealed by proteomic analysis

    International Nuclear Information System (INIS)

    Considering that stimulation of melanogenesis may lead to alterations of cellular responses, besides melanin production, our main goal was to study the cellular effects of melanogenesis stimulation of B16-F10 melanoma cells. Our results show increased levels of the reactive oxygen species after 15 h of melanogenesis stimulation. Following 48 h of melanogenesis stimulation, proliferation was inhibited (by induction of cell cycle arrest in the G1 phase) and the expression levels of p21 mRNA were increased. In addition, melanogenesis stimulation did not induce cellular senescence. Proteomic analysis demonstrated the involvement of proteins from other pathways besides those related to the cell cycle, including protein disulfide isomerase A3, heat-shock protein 70, and fructose biphosphate aldolase A (all up-regulated), and lactate dehydrogenase (down-regulated). In RT-qPCR experiments, the levels of pyruvate kinase M2 mRNA dropped, whereas the levels of ATP synthase (beta-F1) mRNA increased. These data indicate that melanogenesis stimulation of B16-F10 cells leads to alterations in metabolism and cell cycle progression that may contribute to an induction of cell quiescence, which may provide a mechanism of resistance against cellular injury promoted by melanin synthesis. -- Highlights: ► Melanogenesis stimulation by L-tyrosine+NH4Cl in B16-F10 melanoma cells increases ROS levels. ► Melanogenesis inhibits cell proliferation, and induced cell cycle arrest in the G1 phase. ► Proteomic analysis showed alterations in proteins of the cell cycle and glucose metabolism. ► RT-qPCR analysis confirmed alterations of metabolic targets after melanogenesis stimulation.

  2. Melanogenesis stimulation in B16-F10 melanoma cells induces cell cycle alterations, increased ROS levels and a differential expression of proteins as revealed by proteomic analysis

    Energy Technology Data Exchange (ETDEWEB)

    Cunha, Elizabeth S.; Kawahara, Rebeca [Departamento de Bioquimica e Biologia Molecular, Setor de Ciencias Biologicas, Universidade Federal do Parana, P.O. Box 19046, CEP 81531-990, Curitiba, PR (Brazil); Kadowaki, Marina K. [Universidade Estadual do Oeste do Parana, Cascavel, PR (Brazil); Amstalden, Hudson G.; Noleto, Guilhermina R.; Cadena, Silvia Maria S.C.; Winnischofer, Sheila M.B. [Departamento de Bioquimica e Biologia Molecular, Setor de Ciencias Biologicas, Universidade Federal do Parana, P.O. Box 19046, CEP 81531-990, Curitiba, PR (Brazil); Martinez, Glaucia R., E-mail: grmartinez@ufpr.br [Departamento de Bioquimica e Biologia Molecular, Setor de Ciencias Biologicas, Universidade Federal do Parana, P.O. Box 19046, CEP 81531-990, Curitiba, PR (Brazil)

    2012-09-10

    Considering that stimulation of melanogenesis may lead to alterations of cellular responses, besides melanin production, our main goal was to study the cellular effects of melanogenesis stimulation of B16-F10 melanoma cells. Our results show increased levels of the reactive oxygen species after 15 h of melanogenesis stimulation. Following 48 h of melanogenesis stimulation, proliferation was inhibited (by induction of cell cycle arrest in the G1 phase) and the expression levels of p21 mRNA were increased. In addition, melanogenesis stimulation did not induce cellular senescence. Proteomic analysis demonstrated the involvement of proteins from other pathways besides those related to the cell cycle, including protein disulfide isomerase A3, heat-shock protein 70, and fructose biphosphate aldolase A (all up-regulated), and lactate dehydrogenase (down-regulated). In RT-qPCR experiments, the levels of pyruvate kinase M2 mRNA dropped, whereas the levels of ATP synthase (beta-F1) mRNA increased. These data indicate that melanogenesis stimulation of B16-F10 cells leads to alterations in metabolism and cell cycle progression that may contribute to an induction of cell quiescence, which may provide a mechanism of resistance against cellular injury promoted by melanin synthesis. -- Highlights: Black-Right-Pointing-Pointer Melanogenesis stimulation by L-tyrosine+NH{sub 4}Cl in B16-F10 melanoma cells increases ROS levels. Black-Right-Pointing-Pointer Melanogenesis inhibits cell proliferation, and induced cell cycle arrest in the G1 phase. Black-Right-Pointing-Pointer Proteomic analysis showed alterations in proteins of the cell cycle and glucose metabolism. Black-Right-Pointing-Pointer RT-qPCR analysis confirmed alterations of metabolic targets after melanogenesis stimulation.

  3. Ultrastructure of neuronal-like cells differentiated from adult adipose-derived stromal cells

    Institute of Scientific and Technical Information of China (English)

    Changqing Ye; Xiaodong Yuan; Hui Liu; Yanan Cai; Ya Ou

    2010-01-01

    β-mercaptoethanol induces in vitro adult adipose-derived stromal cells (ADSCs) to differentiate into neurons. However, the ultrastructural features of the differentiated neuronal-like cells remain unknown. In the present study, inverted phase contrast microscopy was utilized to observe β-mercaptcethanol-induced differentiation of neuronal-like cells from human ADSCs, and immunocytochemistry and real-time polymerase chain reaction were employed to detect expression of a neural stem cells marker (nestin), a neuronal marker (neuron-specific enolase), and a glial marker (glial fibrillary acidic protein). In addition, ultrastructure of neuronal-like cells was observed by transmission election microscopy. Results revealed highest expression rate of nestin and neuron-specific enolase at 3 and 5 hours following induced differentiation; cells in the 5-hour induction group exhibited a neuronal-specific structure, i.e., Nissl bodies. However, when induction solution was replaced by complete culture medium after 8-hour induction, the differentiated cells reverted to the fibroblast-like morphology from day 1. These results demonstrate that β-mercaptoethanol-induced ADSCs induced differentiation into neural stem cells, followed by morphology of neuronal-like cells. However, this differentiation state was not stable.

  4. Aircraft noise exposure affects rat behavior, plasma norepinephrine levels, and cell morphology of the temporal lobe.

    Science.gov (United States)

    Di, Guo-Qing; Zhou, Bing; Li, Zheng-Guang; Lin, Qi-Li

    2011-12-01

    In order to investigate the physiological effects of airport noise exposure on organisms, in this study, we exposed Sprague-Dawley rats in soundproof chambers to previously recorded aircraft-related noise for 65 d. For comparison, we also used unexposed control rats. Noise was arranged according to aircraft flight schedules and was adjusted to its weighted equivalent continuous perceived noise levels (L(WECPN)) of 75 and 80 dB for the two experimental groups. We examined rat behaviors through an open field test and measured the concentrations of plasma norepinephrine (NE) by high performance liquid chromatography-fluorimetric detection (HPLC-FLD). We also examined the morphologies of neurons and synapses in the temporal lobe by transmission electron microscopy (TEM). Our results showed that rats exposed to airport noise of 80 dB had significantly lower line crossing number (Paircraft noise of 80 dB exposure for 65 d. In conclusion, exposing rats to long-term aircraft noise affects their behaviors, plasma NE levels, and cell morphology of the temporal lobe. PMID:22135145

  5. Aircraft noise exposure affects rat behavior, plasma norepinephrine levels, and cell morphology of the temporal lobe

    Institute of Scientific and Technical Information of China (English)

    Guo-qing DI; Bing ZHOU; Zheng-guang; LI, Qi-li LIN

    2011-01-01

    In order to investigate the physiological effects of airport noise exposure on organisms,in this study,we exposed Sprague-Dawley rats in soundproof chambers to previously recorded aircraft-related noise for 65 d.For comparison,we also used unexposed control rats.Noise was arranged according to aircraft flight schedules and was adjusted to its weighted equivalent continuous perceived noise levels (LwEcPN) of 75 and 80 dB for the two experimental groups.We examined rat behaviors through an open field test and measured the concentrations of plasma norepinephrine (NE) by high performance liquid chromatography-fluorimetric detection (HPLC-FLD).We also examined the morphologies of neurons and synapses in the temporal lobe by transmission electron microscopy (TEM).Our results showed that rats exposed to airport noise of 80 dB had significantly lower line crossing number (P<0.05) and significantly longer center area duration (P<0.05) than control animals.After 29 d of airport noise exposure,the concentration of plasma NE of exposed rats was significantly higher than that of the control group (P<0.05).We also determined that the neuron and synapsis of the temporal lobe of rats showed signs of damage after aircraft noise of 80 dB exposure for 65 d.In conclusion,exposing rats to long-term aircraft noise affects their behaviors,plasma NE levels,and cell morphology of the temporal lobe.

  6. ULK1 regulates melanin levels in MNT-1 cells independently of mTORC1.

    Directory of Open Access Journals (Sweden)

    Eyal Kalie

    Full Text Available Melanosomes are lysosome-related organelles that serve as specialized sites of melanin synthesis and storage in melanocytes. The progression of melanosomes through the different stages of their formation requires trafficking of specific proteins and membrane constituents in a sequential manner, which is likely to deploy ubiquitous cellular machinery along with melanocyte-specific proteins. Recent evidence revealed a connection between melanogenesis and the autophagy machinery, suggesting a novel role for members of the latter in melanocytes. Here we focused on ULK1, a key autophagy protein which is negatively regulated by mTORC1, to assess its potential role in melanogenesis in MNT-1 cells. We found that ULK1 depletion causes an increase in melanin levels, suggesting an inhibitory function for this protein in melanogenesis. Furthermore, this increase was accompanied by increased transcription of MITF (microphthalmia-associated transcription factor and tyrosinase and by elevated protein levels of tyrosinase, the rate-limiting factor in melanin biogenesis. We also provide evidence to show that ULK1 function in this context is independent of the canonical ULK1 autophagy partners, ATG13 and FIP200. Furthermore we show that regulation of melanogenesis by ULK1 is independent of mTORC1 inhibition. Our data thus provide intriguing insights regarding the involvement of the key regulatory autophagy machinery in melanogenesis.

  7. Effect of hypoxic cell radiosensitizers on glutathione level and related enzyme activities in isolated rat hepatocytes

    International Nuclear Information System (INIS)

    A comparative study of the effect of misonidazole and novel radiosensitizers on glutathione (GSH) levels and related enzyme activities in isolated rat hepatocytes was performed. Incubation of hepatocytes with 5 mM radiosensitizers led to a decrease in the intracellular GSH level. The most pronounced decrease in cellular GSH was evoked by 2,4-dinitromidazole-1-ethanol (DNIE); after incubation for only 15 min, GSH was hardly detected. DNIE-mediated GSH loss was dependent upon its concentration. DNIE reacted with GSH nonenzymatically as well as with diethylmaleate, while misonidazole and 1-methyl-2-methyl-sulfinyl-5-methoxycarbonylimidazole (KIH-3) did not. Addition of partially purified glutathione S-transferase (GST) did not enhance DNIE-mediated GSH loss in a cell-free system. DNIE inhibited glutathione peroxidase (GSH-Px), GST, and glutathione reductase (GSSG-R) activities in hepatocytes, while misonidazole and KIH-3 did not. GSH-Px activity assayed with H2O2 as substrate was the most inhibited. Inhibition of GSH-Px activity assayed with cumene hydroperoxide as substrate and GST was less than that of GSH-Px assayed with H2O2 as substrate. GSSG-R activity was decreased by DNIE, but not significantly. Incubation of purified GSH-Px with DNIE resulted in a little change in the activity when assayed with H2O2 as substrate. 26 references, 2 figures, 4 tables

  8. Obesity suppresses circulating level and function of endothelial progenitor cells and heart function

    Directory of Open Access Journals (Sweden)

    Tsai Tzu-Hsien

    2012-07-01

    Full Text Available Abstract Background and aim This study tested the hypothesis that obesity suppresses circulating number as well as the function of endothelial progenitor cells (EPCs and left ventricular ejection fraction (LVEF. Methods High fat diet (45 Kcal% fat was given to 8-week-old C57BL/6 J mice (n = 8 for 20 weeks to induce obesity (group 1. Another age-matched group (n = 8 were fed with control diet for 20 weeks as controls (group 2. The animals were sacrificed at the end of 20 weeks after obesity induction. Results By the end of study period, the heart weight, body weight, abdominal fat weight, serum levels of total cholesterol and fasting blood sugar were remarkably higher in group 1 than in group 2 (all p Conclusions Obesity diminished circulating EPC level, impaired the recovery of damaged endothelium, suppressed EPC angiogenesis ability and LVEF, and increased LV remodeling.

  9. Influence of Hydration Level on Polymer and Water Dynamics in Alkaline Anion Exchange Fuel Cell Membranes

    Science.gov (United States)

    Tarver, Jacob; Kim, Jenny; Tyagi, Madhu; Soles, Christopher; Tsai, Tsung-Han; Coughlin, Bryan

    2015-03-01

    Triblock copolymers based on poly(chloromethylstyrene)-b-poly(ethylene)-b-poly(chloromethylstyrene) can be quaternized to different extents to yield anion exchange membranes for alkaline fuel cells. In the absence of moisture, these membranes demonstrate bilayer lamellar morphology. Upon high levels of hydration, however, in-situ small angle neutron scattering reveals the emergence of higher-order diffraction peaks. This phenomena has previously been observed in analogous diblock copolymer-based membranes and has been attributed to the induction of a multilayer lamellar morphology in which selective striping of water occurs in the center of the ion-rich domain. By conducting humidity-resolved quasielastic neutron scattering (QENS) measurements using deuterated water, we are able to isolate differences in the pico- to nanosecond timescale dynamics of the hydrogenated membrane upon hydration. QENS measurements in the presence of a hydrogenated water source subsequently permit deconvolution and isolation of the translational and rotational dynamics of water as a function of relative humidity, revealing spatial and temporal changes in polymer and water motion at high levels of hydration.

  10. Na+ and K+ levels in living cells: do they depend on the rate of outward transport of Na+?

    Science.gov (United States)

    Ling, G N; Ochsenfeld, M M

    1976-01-01

    At 25 degrees C, frog sartorius muslces rapidly gained Na+ and lost K+ in iodoacetamide and pure nitrogen. Beginning at normal levels, the concentrations of these ions in the cells reached those in the surrounding Ringer solution in 140 min. Yet during that time the Na+ efflux rate showed no sign of the slowing down demanded by Na-pump theory. The data support the view that maintenance and alterations of N1+ levels in frog muslce cells reflect adsorption on protein sites and the solubility property of bulk phase water and are independent of the rate at which Na+ leaves the cell surface. PMID:1088477

  11. The level of expression of the rat growth hormone gene in liver tumor cells is at least eight orders of magnitude less than that in anterior pituitary cells.

    OpenAIRE

    Ivarie, R D; Schacter, B S; O'Farrell, P H

    1983-01-01

    Rat liver hepatoma cells (HTC) which express liver-specific gene products were assayed for the expression of the rat growth hormone (rGH) gene, which is normally expressed in anterior pituitary somatotrophs. The combination of immunoprecipitation and two-dimensional gel electrophoresis provided a highly sensitive assay for rGH synthesis at levels as low as one part in 10(9) of cell protein synthesis (or four molecules of rGH per cell). No rGH expression was detected at this level. The lack of...

  12. High normal fasting glucose level in obese youth: a marker for insulin resistance and beta cell dysregulation.

    LENUS (Irish Health Repository)

    O'Malley, G

    2010-06-01

    A high but normal fasting plasma glucose level in adults is a risk factor for future development of type 2 diabetes mellitus and cardiovascular disease. We investigated whether normal fasting plasma glucose levels (<5.60 mmol\\/l) are associated with decreases in insulin sensitivity and beta cell function, as well as an adverse cardiovascular profile in obese youth.

  13. A medaka model of cancer allowing direct observation of transplanted tumor cells in vivo at a cellular-level resolution.

    Science.gov (United States)

    Hasegawa, Sumitaka; Maruyama, Kouichi; Takenaka, Hikaru; Furukawa, Takako; Saga, Tsuneo

    2009-08-18

    The recent success with small fish as an animal model of cancer with the aid of fluorescence technique has attracted cancer modelers' attention because it would be possible to directly visualize tumor cells in vivo in real time. Here, we report a medaka model capable of allowing the observation of various cell behaviors of transplanted tumor cells, such as cell proliferation and metastasis, which were visualized easily in vivo. We established medaka melanoma (MM) cells stably expressing GFP and transplanted them into nonirradiated and irradiated medaka. The tumor cells were grown at the injection sites in medaka, and the spatiotemporal changes were visualized under a fluorescence stereoscopic microscope at a cellular-level resolution, and even at a single-cell level. Tumor dormancy and metastasis were also observed. Interestingly, in irradiated medaka, accelerated tumor growth and metastasis of the transplanted tumor cells were directly visualized. Our medaka model provides an opportunity to visualize in vivo tumor cells "as seen in a culture dish" and would be useful for in vivo tumor cell biology. PMID:19666513

  14. Theobroma cacao increases cells viability and reduces IL-6 and sVCAM-1 level in endothelial cells induced by plasma from preeclamptic patients.

    Science.gov (United States)

    Rahayu, Budi; Baktiyani, Siti Candra Windu; Nurdiana, Nurdiana

    2016-01-01

    This study aims to investigate whether an ethanolic extract of Theobroma cacao bean is able to increase cell viability and decrease IL-6 and sVCAM-1 in endothelial cells induced by plasma from preeclamptic patients. Endothelial cells were obtained from human umbilical vascular endothelial cells. At confluency, endothelial cells were divided into six groups, which included control (untreated), endothelial cells exposed to plasma from normal pregnancy, endothelial cells exposed to 2% plasma from preeclamptic patients (PP), endothelial cells exposed to PP in the presence of ethanolic extract of T. cacao (PP+TC) at the following three doses: 25, 50, and 100ppm. The analysis was performed in silico using the Hex 8.0, LigPlus and LigandScout 3.1 software. Analysis on IL-6 and sVCAM-1 levels were done by enzyme linked immunosorbent assay (ELISA). We found that seven of them could bind to the protein NFκB (catechin, leucoanthocyanidin, niacin, phenylethylamine, theobromine, theophylline, and thiamin). This increase in IL-6 was significantly (Pcacao extract. Plasma from PP significantly increased sVCAM-1 levels compared to untreated cells. This increase in sVCAM-1 was significantly attenuated by all doses of the extract. In conclusion, T. cacao extract prohibits the increase in IL-6 and sVCAM-1 in endothelial cells induced by plasma from preeclamptic patients. Therefore this may provide a herbal therapy for attenuating the endothelial dysfunction found in preeclampsia. PMID:26955771

  15. The influence of 60Co gamma rays to cell reproduction (An experiment using low dose levels on vero and primary monkey kidney cells)

    International Nuclear Information System (INIS)

    Vero and primary monkey kidney cells in culture were gamma irradiated with doses of 0, 0.4 and 0.8 Gy at a dose-rate of 1.30-1.45x103Gy/hour. At harvest time 3 days post irradiation, 0.4 Gy proved to be able to lower the number of vero cells in such a degree that it became significantly different from the control, whereas 0.8 Gy could not suppress the number of primary cells to a level that differed significantly from its control. At harvest time of 7 days post irradiation, 0.4 Gy was found effective in lowering both vero and primary cells so that the number of the harvested cells were significantly different from the controls. At harvest time of 3 days post irradiation, 0.8 Gy caused both cell types reached levels that were not significantly different from 0.4 Gy, but at 7 days post irradiation the number of vero cells was very significantly different from that of 0.4 Gy, while the number of primary cells remained equal to that of 0.4 Gy. This phenomenon showed that irradiation could cause greater injurious effect at more advanced post irradiation times, while the more proliferative vero cells proved to be more susceptible to irradiation than primary cells, but at the same time more potential in performing repair. (author)

  16. Cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser.

    Science.gov (United States)

    Alexsandra da Silva Neto Trajano, Larissa; da Silva, Camila Luna; de Carvalho, Simone Nunes; Cortez, Erika; Mencalha, André Luiz; de Souza da Fonseca, Adenilson; Stumbo, Ana Carolina

    2016-07-01

    Low-level infrared laser is considered safe and effective for treatment of muscle injuries. However, the mechanism involved on beneficial effects of laser therapy are not understood. The aim was to evaluate cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser at therapeutic fluences. C2C12 myoblast cultures at different (2 and 10 %) fetal bovine serum (FBS) concentrations were exposed to low-level infrared laser (808 nm, 100 mW) at different fluences (10, 35, and 70 J/cm(2)) and evaluated after 24, 48, and 72 h. Cell viability was evaluated by WST-1 assay; reactive oxygen species (ROS), apoptosis, and necrosis were evaluated by flow cytometry. Cell viability was decreased atthe lowest FBS concentration. Laser exposure increased the cell viability in myoblast cultures at 2 % FBS after 48 and 72 h, but no significant increase in ROS was observed. Apoptosis was decreased at the higher fluence and necrosis was increased at lower fluence in myoblast cultures after 24 h of laser exposure at 2 % FBS. No laser-induced alterations were obtained at 10 % FBS. Results show that level of reactive oxygen species is not altered, at least to those evaluated in this study, but low-level infrared laser exposure affects cell viability, apoptosis, and necrosis in myoblast cultures depending on laser fluence and physiologic conditions of cells. PMID:26886589

  17. Quasi-Fermi level splitting evaluation based on electroluminescence analysis in multiple quantum-well solar cells for investigating cell performance under concentrated light

    Science.gov (United States)

    Inoue, Tomoyuki; Toprasertpong, Kasidit; Delamarre, Amaury; Watanabe, Kentaroh; Paire, Myriam; Lombez, Laurent; Guillemoles, Jean-François; Sugiyama, Masakazu; Nakano, Yoshiaki

    2016-03-01

    Insertion of InGaAs/GaAsP strain-balanced multiple quantum wells (MQWs) into i-regions of GaAs p-i-n solar cells show several advantages against GaAs bulk p-i-n solar cells. Particularly under high-concentration sunlight condition, enhancement of the open-circuit voltage with increasing concentration ratio in thin-barrier MQW cells has been reported to be more apparent than that in GaAs bulk cells. However, investigation of the MQW cell mechanisms in terms of I-V characteristics under high-concentration sunlight suffers from the increase in cell temperature and series resistance. In order to investigate the mechanism of the steep enhancement of open-circuit voltage in MQW cells under high-concentration sunlight without affected by temperature, the quasi-Fermi level splitting was evaluated by analyzing electroluminescence (EL) from a cell. Since a cell under current injection with a density Jinjhas similar excess carrier density to a cell under concentrated sunlight with an equivalent short-circuit current Jsc = Jinj, EL measurement with varied Jinj can approximately evaluate a cell performance under a variety of concentration ratio. In addition to the evaluation of quasi-Fermi level splitting, the external luminescence efficiency was also investigated with the EL measurement. The MQW cells showed higher external luminescence efficiency than the GaAs reference cells especially under high-concentration condition. The results suggest that since the MQW region can trap and confine carriers, the localized excess carriers inside the cells make radiative recombination more dominant.

  18. Analysis of Protein Levels of 24 Cytokines in Scrapie Agent-Infected Brain and Glial Cell Cultures from Mice Differing in Prion Protein Expression Levels

    OpenAIRE

    Tribouillard-Tanvier, Déborah; Striebel, James F; Peterson, Karin E.; Chesebro, Bruce

    2009-01-01

    Activation of microglia and astroglia is seen in many neurodegenerative diseases including prion diseases. Activated glial cells produce cytokines as a protective response against certain pathogens and as part of the host inflammatory response to brain damage. In addition, cytokines might also exacerbate tissue damage initiated by other processes. In the present work using multiplex assays to analyze protein levels of 24 cytokines in scrapie agent-infected C57BL/10 mouse brains, we observed e...

  19. β-asarone and levodopa co-administration increase striatal dopamine level in 6-hydroxydopamine induced rats by modulating P-glycoprotein and tight junction proteins at the blood-brain barrier and promoting levodopa into the brain.

    Science.gov (United States)

    Huang, Liping; Deng, Minzhen; He, Yuping; Lu, Shiyao; Ma, Ruanxin; Fang, Yongqi

    2016-06-01

    Levodopa (L-dopa) is widely considered as one of the most effective drug constituents in the treatment of Parkinson's disease (PD), but the blood-brain barrier (BBB) permeability of L-dopa is dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), S100β and neuron-specific enolase (NSE) levels were subsequently determined. The P-glycoprotein (P-gp), zonula occludens-1 (ZO-1), claudin-5, occludin and actin expression was also assessed in cortex. Changes in BBB ultrastructure were observed using transmission electron microscopy. Our results showed that the co-administered treatment increased levels of L-dopa, DA, DOPAC and HVA in striatum, and S100β in plasma, but down-regulated NSE, P-gp, ZO-1, occludin, actin and claudin-5 in cortex. Crevices were observed between capillary endothelial cells at intercellular tight junction of the striatum in co-administered-treated group, while the endothelial cells in untreated group were tightly jointing each other. In addition, the correlations of L-dopa or DA and P-gp or tight junction proteins respectively were significantly negative in co-administered- and β-asarone-treated groups. These findings suggest that co-administered treatment may enhance the L-dopa BBB permeability and attenuate brain injury, which may be beneficial to PD treatment. PMID:26991136

  20. The ATM and ATR inhibitors CGK733 and caffeine suppress cyclin D1 levels and inhibit cell proliferation

    International Nuclear Information System (INIS)

    The ataxia telangiectasia mutated (ATM) and the ATM- related (ATR) kinases play a central role in facilitating the resistance of cancer cells to genotoxic treatment regimens. The components of the ATM and ATR regulated signaling pathways thus provide attractive pharmacological targets, since their inhibition enhances cellular sensitivity to chemo- and radiotherapy. Caffeine as well as more specific inhibitors of ATM (KU55933) or ATM and ATR (CGK733) have recently been shown to induce cell death in drug-induced senescent tumor cells. Addition of these agents to cancer cells previously rendered senescent by exposure to genotoxins suppressed the ATM mediated p21 expression required for the survival of these cells. The precise molecular pharmacology of these agents however, is not well characterized. Herein, we report that caffeine, CGK733, and to a lesser extent KU55933, inhibit the proliferation of otherwise untreated human cancer and non-transformed mouse fibroblast cell lines. Exposure of human cancer cell lines to caffeine and CGK733 was associated with a rapid decline in cyclin D1 protein levels and a reduction in the levels of both phosphorylated and total retinoblastoma protein (RB). Our studies suggest that observations based on the effects of these compounds on cell proliferation and survival must be interpreted with caution. The differential effects of caffeine/CGK733 and KU55933 on cyclin D1 protein levels suggest that these agents will exhibit dissimilar molecular pharmacological profiles

  1. Increased serum cell-free DNA levels in relation to inflammation are predictive of distant metastasis of esophageal squamous cell carcinoma

    OpenAIRE

    TOMOCHIKA, SHINOBU; Iizuka, Norio; Watanabe, Yusaku; TSUTSUI, MASAHITO; Takeda, Shigeru; Yoshino, Shigefumi; ICHIHARA, KIYOSHI; Oka, Masaaki

    2010-01-01

    Distant metastasis hinders a favorable outcome for patients with esophageal squamous cell carcinoma (ESCC) by limiting the surgical cure. The levels of cell-free DNA (cfDNA) in the blood have served as a predictor for metastasis and recurrence in distant organs in liver cancer. Thus, this study tested the clinical efficacy of serum cfDNA levels as a predictive marker for distant metastasis of ESCC. We investigated cfDNA levels in a cohort of 101 ESCC patients and 46 age- and gender-matched co...

  2. High level PHGDH expression in breast is predominantly associated with keratin 5-positive cell lineage independently of malignancy

    DEFF Research Database (Denmark)

    Gromova, Irina; Gromov, Pavel; Honma, Naoko; Kumar, Sudha; Rimm, David; Talman, Maj-Lis Møller; Wielenga, Vera Timmermans; Moreira, José

    2015-01-01

    showed high-level expression of Phgdh in normal CK5-positive mammary epithelial cells, indicating that expression of this protein was not associated with malignancy, but rather with cell lineage. However, proteomic profiling of Phgdh showed it to be expressed in two major protein forms, and that the...... in TNBC samples. One such protein was D-3-phosphoglycerate dehydrogenase (Phgdh), a candidate oncogene. We analysed expression of Phgdh in normal and TNBC mammary tissue samples by 2D gel-based proteomics and immunohistochemistry (IHC), and show here that high-level expression of Phgdh in mammary...... epithelial cells is primarily associated with cell lineage, as we found that Phgdh expression was predominant in CK5-positive cells, normal as well as malignant, thus identifying an association of this protein with the basal phenotype. Quantitative IHC analysis of Phgdh expression in normal breast tissue...

  3. Genes adopt non-optimal codon usage to generate cell cycle-dependent oscillations in protein levels

    DEFF Research Database (Denmark)

    Frenkel-Morgenstern, Milana; Danon, Tamar; Christian, Thomas;

    2012-01-01

    The cell cycle is a temporal program that regulates DNA synthesis and cell division. When we compared the codon usage of cell cycle-regulated genes with that of other genes, we discovered that there is a significant preference for non-optimal codons. Moreover, genes encoding proteins that cycle at...... the protein level exhibit non-optimal codon preferences. Remarkably, cell cycle-regulated genes expressed in different phases display different codon preferences. Here, we show empirically that transfer RNA (tRNA) expression is indeed highest in the G2 phase of the cell cycle, consistent with the non......-optimal codon usage of genes expressed at this time, and lowest toward the end of G1, reflecting the optimal codon usage of G1 genes. Accordingly, protein levels of human glycyl-, threonyl-, and glutamyl-prolyl tRNA synthetases were found to oscillate, peaking in G2/M phase. In light of our findings, we propose...

  4. NMR spectroscopy analysis of phosphorus metabolites and the effect of adriamycin on these metabolite levels in an adriamycin-sensitive and -resistant human small cell lung carcinoma cell line

    NARCIS (Netherlands)

    de Jong, Steven; Mulder, N H; de Vries, Liesbeth; Robillard, G T

    1991-01-01

    P-31 nuclear magnetic resonance (NMR) spectra of cells and of cell extracts revealed high levels of phosphorylcholine (PC) and phosphocreatine (PCr) in an adriamycin-resistant human small cell lung carcinoma cell line (GLC4/ADR) and the adriamycin-sensitive parental cell line (GLC4). PCr levels in e

  5. Grape pomace extract exerts antioxidant effects through an increase in GCS levels and GST activity in muscle and endothelial cells.

    Science.gov (United States)

    Goutzourelas, Nikolaos; Stagos, Dimitrios; Housmekeridou, Anastasia; Karapouliou, Christina; Kerasioti, Efthalia; Aligiannis, Nektarios; Skaltsounis, Alexios L; Spandidos, Demetrios A; Tsatsakis, Aristidis M; Kouretas, Demetrios

    2015-08-01

    In a previous study, we demonstrated that a grape pomace extract (GPE) exerted antioxidant activity in endothelial (EA.hy926) and muscle (C2C12) cells through an increase in glutathione (GSH) levels. In the present study, in order to elucidate the mechanisms responsible for the antioxidant activity of GPE, its effects on the expression of critical antioxidant enzymes, such as catalase (CAT), superoxide dismutase (SOD)1, heme oxygenase 1 (HO-1) and gamma-glutamylcysteine synthetase (GCS) were assessed in EA.hy926 and C2C12 cells. Moreover, the effects of GPE on CAT, SOD and glutathione S-transferase (GST) enzymatic activity were evaluated. For this purpose, the C2C12 and EA.hy926 cells were treated with GPE at low and non-cytotoxic concentrations (2.5 and 10 µg/ml for the C2C12 cells; 0.068 and 0.250 µg/ml for the EA.hy926 cells) for 3, 6, 12, 18 and 24 h. Following incubation, enzymatic expression and activity were assessed. The results revealed that treatment with GPE significantly increased GCS levels and GST activity in both the C2C12 and EA.hy926 cells. However, GPE significantly decreased CAT levels and activity, but only in the muscle cells, while it had no effect on CAT levels and activity in the endothelial cells. Moreover, treatment with GPE had no effect on HO-1 and SOD expression and activity in both cell lines. Therefore, the present results provide further evidence of the crucial role of GSH systems in the antioxidant effects exerted by GPE. Thus, GPE may prove to be effective for use as a food supplement for the treatment of oxidative stress-induced pathological conditions of the cardiovascular and skeletal muscle systems, particularly those associated with low GSH levels. PMID:26082074

  6. Low levels of Bax inhibitor-1 gene expression increase tunicamycin-induced apoptosis in human neuroblastoma SY5Y cells

    Institute of Scientific and Technical Information of China (English)

    Dan Wu; Peirong Wang; Shiyao Wang

    2012-01-01

    A human SH-SY5Y neuroblastoma cell line with a low level of Bax inhibitor-1 expression was established by lentivirus-mediated RNA interference and fluorescence-activated cell sorting. In control SH-SY5Y cells, tunicamycin treatment induced endoplasmic reticulum stress-mediated apoptosis; however, after Bax inhibitor-1 gene knockdown, cell survival rates were significantly decreased and the degree of apoptosis was significantly increased following tunicamycin treatment. In addition, chromatin condensation and apparent apoptotic phenomena, such as marginalization and cytoplasmic vesicles, were observed. Our findings indicate that Bax inhibitor-1 can delay apoptosis induced by endoplasmic reticulum stress.

  7. Acute plasma biomarkers of T cell activation set-point levels and of disease progression in HIV-1 infection.

    Directory of Open Access Journals (Sweden)

    Anne-Sophie Liovat

    Full Text Available T cell activation levels, viral load and CD4(+ T cell counts at early stages of HIV-1 infection are predictive of the rate of progression towards AIDS. We evaluated whether the inflammatory profile during primary HIV-1 infection is predictive of the virological and immunological set-points and of disease progression. We quantified 28 plasma proteins during acute and post-acute HIV-1 infection in individuals with known disease progression profiles. Forty-six untreated patients, enrolled during primary HIV-1 infection, were categorized into rapid progressors, progressors and slow progressors according to their spontaneous progression profile over 42 months of follow-up. Already during primary infection, rapid progressors showed a higher number of increased plasma proteins than progressors or slow progressors. The plasma levels of TGF-β1 and IL-18 in primary HIV-1 infection were both positively associated with T cell activation level at set-point (6 months after acute infection and together able to predict 74% of the T cell activation variation at set-point. Plasma IP-10 was positively and negatively associated with, respectively, T cell activation and CD4(+ T cell counts at set-point and capable to predict 30% of the CD4(+ T cell count variation at set-point. Moreover, plasma IP-10 levels during primary infection were predictive of rapid progression. In primary infection, IP-10 was an even better predictor of rapid disease progression than viremia or CD4(+ T cell levels at this time point. The superior predictive capacity of IP-10 was confirmed in an independent group of 88 HIV-1 infected individuals. Altogether, this study shows that the inflammatory profile in primary HIV-1 infection is associated with T cell activation levels and CD4(+ T cell counts at set-point. Plasma IP-10 levels were of strong predictive value for rapid disease progression. The data suggest IP-10 being an earlier marker of disease progression than CD4(+ T cell counts or

  8. Raloxifene induces cell death and inhibits proliferation through multiple signaling pathways in prostate cancer cells expressing different levels of estrogen receptor α and β.

    Science.gov (United States)

    Rossi, V; Bellastella, G; De Rosa, C; Abbondanza, C; Visconti, D; Maione, L; Chieffi, P; Della Ragione, F; Prezioso, D; De Bellis, A; Bellastella, A; Sinisi, A A

    2011-05-01

    Raloxifene (RAL), a selective estrogen receptor (ER) modulator (SERM) seems to induce apoptosis in both androgen-dependent and -independent prostate cell (PC) lines via activation of ERβ and an antagonistic effect on ERα. In this study, we evaluated the effects of RAL on epithelial PC growth using the two following in vitro models: the androgen-dependent cell line EPN which expressed both ERs; and a stabilized epithelial cell line derived from a prostate cancer specimen (CPEC), which expressed low levels of ERβ and lacked ERα. In EPN cells, there was an increase in the pre-G1 apoptotic peak and a reduction in the S phase of the cell cycle with G0/G1 arrest after E2 or RAL treatment; bcl-2 mRNA and Bcl-2 protein levels were significantly reduced, while activated caspase-3 and Par-4 levels increased significantly after either E2 or RAL treatment; in addition, c-myc transcript was inhibited after 10(-6)  M RAL treatment. A dose-dependent increase of metallothionein II gene RNA level was also induced by RAL in EPN. In CPEC, there was only a weak apoptotic peak associated with caspase-3 activation and Par-4 increase after either E2 or RAL treatment; while c-myc transcript level increased. RAL induced a rapid but transient phosphorylation of ERK 1/2 in EPN cells but generated a sustained effect in CPEC. These findings suggest that RAL effects on PC growth control in vitro are cell-specific, depending on ERβ or ERβ/ERα relative expression levels. Moreover, this study demonstrated that RAL affected both transcriptional regulation and non-genomic signals, which resulted in the modulation of multiple signaling pathways of apoptosis and of cell cycle progression. PMID:20945400

  9. Differential regulation of polysome mRNA levels in mouse Hepa-1C1C7 cells exposed to dioxin.

    Science.gov (United States)

    Thornley, Jessica A; Trask, Heidi W; Ridley, Christian J A; Korc, Murray; Gui, Jiang; Ringelberg, Carol S; Wang, Sinny; Tomlinson, Craig R

    2011-10-01

    The environmental agent 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) causes a multitude of human illnesses. In order to more fully understand the underlying biology of TCDD toxicity, we tested the hypothesis that new candidate genes could be identified using polysome RNA from TCDD-treated mouse Hepa-1c1c7 cells. We found that (i) differentially expressed whole cell and cytoplasm RNA levels are both poor predictors of polysome RNA levels; (ii) for a majority of RNAs, differential RNA levels are regulated independently in the nucleus, cytoplasm, and polysomes; (iii) for the remaining polysome RNAs, levels are regulated via several different mechanisms, including a "tagging" of mRNAs in the nucleus for immediate polysome entry; and (iv) most importantly, a gene list derived from differentially expressed polysome RNA generated new genes and cell pathways potentially related to TCDD biology. PMID:21570461

  10. Study on the red blood cell immuno-function and relevant cytokines levels in elderly patients with chronic bronchitis

    International Nuclear Information System (INIS)

    Objective: To explore the changes of red blood cell immuno-function and serum IL-4, IL-5, IL-8, IL-10 levels in elderly patients with chronic bronchitis. Methods: The red cell C3b receptor (RBC-C3bR) and red blood cell immune complex rosette (RBC-ICR) (with immune methods), IL-4, IL-8 levels (with RIA) and IL-10, IL-5 levels (with ELISA) were measured were measured in 48 elderly patients with chronic bronchitis and 35 controls. Results: RBC-C3bRR percentage and IL-10 levels were significantly lower in patients with chronic bronchitis than those in controls (P<0.01) while IL-4, IL-5, IL-8 levels were significantly higher (P<0.01), RBC-C3bRR was significantly negatively correlated to IL-4, IL-5 and IL-8 (r=-0.3112, -0.3415, -0. 3718, P<0.05) and RBC-ICRRR was positiviely correlated to IL-10 level (r=0.3715,P<0.05). Conclusion: The lower red cell immuno-function is closely associated with alteration of cytokines levels in elderly patients with chronic bronchitis. (authors)

  11. Plasma cell-free DNA levels are elevated in acute Puumala hantavirus infection.

    Directory of Open Access Journals (Sweden)

    Tuula K Outinen

    Full Text Available INTRODUCTION: Puumala hantavirus (PUUV causes a hemorrhagic fever with renal syndrome called nephropathia epidemica (NE. The aim of the present study was to evaluate plasma cell-free DNA (cf-DNA levels and urinary cf-DNA excretion in acute NE as well as their associations with the severity of the disease. METHODS: Total plasma cf-DNA was quantified directly in plasma of 61 patients and urine of 20 patients with acute NE. We also carried out a qualitative high-sensitivity lab-on-a-chip DNA assay in 20 patients to elucidate the appearance of cf-DNA in plasma and urine. RESULTS: The maximum plasma cf-DNA values taken during acute NE were significantly higher than the control values taken after the hospitalization period (median 1.33 µg/ml, range 0.94-3.29 µg/ml vs. median 0.77 µg/ml, range 0.55-0.99 µg/ml, P<0.001. The maximum plasma cf-DNA levels correlated positively with maximum blood leukocyte count (r = 0.388, P = 0.002 and the length of hospital stay (r = 0.376, P = 0.003, and inversely with minimum blood platelet count (r = -0.297, P = 0.020. Qualitative analysis of plasma cf-DNA revealed that in most of the patients cf-DNA displayed a low-molecular weight appearance, corresponding to the size of apoptotic DNA (150-200 bp. The visually graded maximum cf-DNA band intensity correlated positively with the maximum quantity of total plasma cf-DNA (r = 0.513, P = 0.021. Maximum urinary excretion of cf-DNA in turn was not markedly increased during the acute phase of NE and did not correlate with any of the variables reflecting severity of the disease or with the maximum plasma cf-DNA level. CONCLUSIONS: The plasma levels of cf-DNA are elevated during acute PUUV infection and correlate with the apoptotic cf-DNA-band intensity. The plasma cf-DNA concentration correlates with some variables reflecting the severity of the disease. The urinary excretion of cf-DNA does not reflect the degree of inflammation in the kidney.

  12. Determination of gamma radiation lethal dose (LD{sub 50}) and resveratrol cytotoxicity level in tumor cells line

    Energy Technology Data Exchange (ETDEWEB)

    Magalhaes, Vanessa D.; Rogero, Sizue O.; Rogero, Jose R. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Cruz, Aurea S. [Instituto Adolfo Lutz (IAL-SP) Secao de Culturas Celulares, SP (Brazil)

    2011-07-01

    Cancer is a disease with high incidence and it is considered a worldwide public health problem. Resveratrol is a polyphenol occurring naturally in a wide variety of plants according to response of ultraviolet radiation (UV) exposition or according to mechanical stress resulting of pathogens or chemical and physical agents. This polyphenol possesses a pharmacological activity of carcinogenesis inhibition in multiple levels. It also protects cells by scavenging the free radicals which are considered toxic products. These free radicals are formed of natural process of cell aging and also by incidence of ionizing radiation in the organism. Thus, resveratrol is considered as a cell radioprotector. On the other hand, in some elevated concentrations resveratrol may be considered as a radiosensitizing. The aim of this work was the determination of radiation lethal dose (LD{sub 50}) and also verifies the cytotoxicity level of resveratrol in tumor cells line: muco epidermoid pulmonary carcinoma cells (NCI-H292) and rhabdomyosarcoma cells (RD). The cytotoxicity test was performed by neutral red uptake assay. The results of resveratrol IC{sub 50%} in NCI-H292 cells was 192{mu}M and in RD cells was 128{mu}M; and RD cells gamma radiation LD{sub 50} was 435Gy. (author)

  13. High-level expression of human insulin receptor cDNA in mouse NIH 3T3 cells

    International Nuclear Information System (INIS)

    In order to develop a simple, efficient system for the high-level expression of human insulin receptors in eukaryotic cells, a full-length human kidney insulin receptor cDNA was inserted into a bovine papilloma virus vector under the control of the mouse metallothionein promoter. After transfection of mouse NIH 3T3 cells with this construct, seven cell lines expressing insulin receptors were isolated; two cell lines had more than 106 receptors per cell. The cell line with the highest 125I-insulin binding (NIH 3T3 HIR3.5) had 6 x 106 receptors with a K/sub d/ of 10-9 M. This level was not dependent on exposure to metals but could be increased further to 2 x 107 receptors per cell by addition of sodium butyrate to the culture medium. The α and β subunits had apparent molecular weights of 147,000 and 105,000, respectively (compared to 135,000 and 95,000 in IM-9 human lymphocytes), values identical to those of the α and β subunits of the insulin receptors of nontransformed NIH 3T3 cells. This size difference was due to altered carbohydrate composition, as N-glycanase digestion reduced the apparent receptor subunit size of the transfected cells and IM-9 lymphocytes to identical values. The alteration in N-linked oligosaccharide composition could not be ascribed to differences in the kinetics of posttranslational processing of the insulin receptors, which was comparable to that of other cells studied. The basal rate of glycogen synthesis in the cells overexpressing insulin receptors was increased 4- to 5-fold compared with controls. Low levels of added insulin (0.1 nM) caused a 50% increase in the rate of glycogen synthesis

  14. High-level expression of human insulin receptor cDNA in mouse NIH 3T3 cells

    Energy Technology Data Exchange (ETDEWEB)

    Whittaker, J.; Okamoto, A.K.; Thys, R.; Bell, G.I.; Steiner, D.F.; Hofmann, C.A.

    1987-08-01

    In order to develop a simple, efficient system for the high-level expression of human insulin receptors in eukaryotic cells, a full-length human kidney insulin receptor cDNA was inserted into a bovine papilloma virus vector under the control of the mouse metallothionein promoter. After transfection of mouse NIH 3T3 cells with this construct, seven cell lines expressing insulin receptors were isolated; two cell lines had more than 10/sup 6/ receptors per cell. The cell line with the highest /sup 125/I-insulin binding (NIH 3T3 HIR3.5) had 6 x 10/sup 6/ receptors with a K/sub d/ of 10/sup -9/ M. This level was not dependent on exposure to metals but could be increased further to 2 x 10/sup 7/ receptors per cell by addition of sodium butyrate to the culture medium. The ..cap alpha.. and ..beta.. subunits had apparent molecular weights of 147,000 and 105,000, respectively (compared to 135,000 and 95,000 in IM-9 human lymphocytes), values identical to those of the ..cap alpha.. and ..beta.. subunits of the insulin receptors of nontransformed NIH 3T3 cells. This size difference was due to altered carbohydrate composition, as N-glycanase digestion reduced the apparent receptor subunit size of the transfected cells and IM-9 lymphocytes to identical values. The alteration in N-linked oligosaccharide composition could not be ascribed to differences in the kinetics of posttranslational processing of the insulin receptors, which was comparable to that of other cells studied. The basal rate of glycogen synthesis in the cells overexpressing insulin receptors was increased 4- to 5-fold compared with controls. Low levels of added insulin (0.1 nM) caused a 50% increase in the rate of glycogen synthesis

  15. Varying Dietary Levels of Molybdenum Inducing Cell Apoptosis of Spleen Under Cadmium Stress in Caprine.

    Science.gov (United States)

    Xiao, Qingyang; Zhang, Caiying; Gu, Xiaolong; Zhuang, Yu; Luo, Junrong; Liu, Ping; Guo, Xiaoquan; Hu, Guoliang; Cao, Huabin

    2016-07-01

    The present experiment aims at evaluating chronic toxic effects of the combination of cadmium (Cd) and molybdenum (Mo) according to residual element contents, apoptosis gene expression, and ultrastructure and histopathology changes of caprine spleen. In total, 36 Boer goats were randomly divided into four groups with the equal number in each group. The control group was orally administered with deionized water while the experimental groups I, II, and III were administered with the equal quantity of CdCl2 (1 mg kg(-1) BW) and (NH4)6·Mo7O24·4H2O including 15, 30, and 45 mg·Mo kg(-1) BW, respectively. Three individuals from each group were treated with euthanasia on days 0, 25, and 50. The data showed that the content of splenic residual Mo and Cd increased (P < 0.05) in the experimental groups on days 25 and 50, while no significant difference was observed in the content of Cu. The apoptosis-related gene expression levels including Bcl-2, Bax, Caspase-3, Smac, and ceruloplasmin (CP) were also determined. Results showed that significant reductions were observed in Bcl-2 and CP expressions (P < 0.01), while Caspase-3 gene was up-regulated (P < 0.05). However, no significant difference was observed in Smac and Bax expressions. Furthermore, on day 50, spleen tissues were presented to observe ultrastructural changes in lesions by means of transmission electron microscopy, with fragmentized nucleus, vesiculation of cytoplasm, mitochondria hyperplasia, and increasing lysosomes included. In addition, histopathology results corroborated the toxicity by showing cell hemorrhage, thickening central arteries, and enhanced capsule thickness. To sum up, our study revealed that the combination of Cd and Mo could induce remarkable damage to the spleen of goats by promoting cell apoptosis in the mitochondrial pathway and affecting the deposition of Mo and Cd. PMID:26585322

  16. Plumbagin reduces chronic lymphocytic leukemia cell survival by downregulation of Bcl-2 but upregulation of the Bax protein level.

    Science.gov (United States)

    Fu, Chunling; Gong, Yanqing; Shi, Xuanxuan; Sun, Zengtian; Niu, Mingshan; Sang, Wei; Xu, Linyan; Zhu, Feng; Wang, Ying; Xu, Kailin

    2016-09-01

    Chronic lymphocytic leukemia (CLL) is the most common leukemia in Western countries, and mainly originates from an accumulation of abnormal B cells caused by the dysregulation of cell proliferation and apoptosis rates. The aberration of apoptosis-related genes in CLL cells results in defective apoptosis of CLL cells in response to traditional therapeutic medicine. Plumbagin (5-hydroxy-2-methyl-1, 4-naphthoquinone), a natural compound from Plumbago zeylinica, has been shown to exhibit pro-apoptotic activities in tumor cells. In the present study, we report that plumbagin effectively inhibited CLL cell viability with a lower dose compared to fludarabine, and inhibited cell proliferation in a dose-dependent manner. In addition, plumbagin promoted accumulation of MEC-1 cells in the S phase, and blocked cell cycle transition of HG3 cells from G0/G1 to S phase. Molecularly, plumbagin markedly induced CLL cell apoptosis through reduction of Bcl-2, but through an increase in the Bax protein level. These results suggest that plumbagin may be considered as a potential anticancer agent for CLL therapy. PMID:27461100

  17. Intestinal crypt homeostasis revealed at single stem cell level by in vivo live-imaging

    Science.gov (United States)

    Zomer, Anoek; Snippert, Hugo J.; de Sauvage, Frederic J.; Simons, Benjamin D.; Clevers, Hans; van Rheenen, Jacco

    2014-01-01

    Summary The rapid turnover of the mammalian intestinal epithelium is supported by stem cells located around the base of the crypt1. Alongside Lgr5, intestinal stem cells have been associated with various markers, which are expressed heterogeneously within the crypt base region1-6. Previous quantitative clonal fate analyses have proposed that homeostasis occurs as the consequence of neutral competition between dividing stem cells7-9. However, the short-term behaviour of individual Lgr5+ cells positioned at different locations within the crypt base compartment has not been resolved. Here, we established the short-term dynamics of intestinal stem cells using a novel approach of continuous intravital imaging of Lgr5-Confetti mice. We find that Lgr5+ cells in the upper part of the niche (termed ‘border cells’) can be passively displaced into the transit-amplifying (TA) domain, following division of proximate cells, implying that determination of stem cell fate can be uncoupled from division. Through the quantitative analysis of individual clonal lineages, we show that stem cells at the crypt base, termed ‘central cells’, experience a survival advantage over border stem cells. However, through the transfer of stem cells between the border and central regions, all Lgr5+ cells are endowed with long-term self-renewal potential. These findings establish a novel paradigm for stem cell maintenance in which a dynamically heterogeneous cell population is able to function long-term as a single stem cell pool. PMID:24531760

  18. Low-level laser therapy: Effects on human face aged skin and cell viability of HeLa cells exposed to UV radiation

    Directory of Open Access Journals (Sweden)

    Mezghani Sana

    2015-01-01

    Full Text Available Chronic and excessive exposure to UV radiation leads to photoaging and photocarcinogenesis. Adequate protection of the skin against the deleterious effects of UV irradiation is essential. Low-level laser therapy (LLLT is a light source in the red to near-infrared range that has been accepted in a variety of medical applications. In this study, we explored the effect of LLLT in human face aged skin and the cell viability of HeLa cells exposed to UV radiation. We found that LLLT significantly reduced visible wrinkles and the loss of firmness of facial skin in aging subjects. Additionally, treatment of cultured HeLa cells with LLLT prior to or post UVA or UVB exposure significantly protected cells from UV-mediated cell death. All results showed the beneficial effects of LLLT on relieving signs of skin aging and its prevention and protection of the cell viability against UV-induced damage.

  19. Cancer Stem Cells in Small Cell Lung Cancer Cell Line H446: Higher Dependency on Oxidative Phosphorylation and Mitochondrial Substrate-Level Phosphorylation than Non-Stem Cancer Cells.

    Science.gov (United States)

    Gao, Cuicui; Shen, Yao; Jin, Fang; Miao, Yajing; Qiu, Xiaofei

    2016-01-01

    Recently, targeting cancer stem cells (CSCs) metabolism is becoming a promising therapeutic approach to improve cancer treatment outcomes. However, knowledge of the metabolic state of CSCs in small cell lung cancer is still lacking. In this study, we found that CSCs had significantly lower oxygen consumption rate and extracellular acidification rate than non-stem cancer cells. Meanwhile, this subpopulation of cells consumed less glucose, produced less lactate and maintained lower ATP levels. We also revealed that CSCs could produce more ATP through mitochondrial substrate-level phosphorylation during respiratory inhibition compared with non-stem cancer cells. Furthermore, they were more sensitive to suppression of oxidative phosphorylation. Therefore, oligomycin (inhibitor of oxidative phosphorylation) could severely impair sphere-forming and tumor-initiating abilities of CSCs. Our work suggests that CSCs represent metabolically inactive tumor subpopulations which sustain in a state showing low metabolic activity. However, mitochondrial substrate-level phosphorylation of CSCs may be more active than that of non-stem cancer cells. Moreover, CSCs showed preferential use of oxidative phosphorylation over glycolysis to meet their energy demand. These results extend our understanding of CSCs metabolism, potentially providing novel treatment strategies targeting metabolic pathways in small cell lung cancer. PMID:27167619

  20. Cancer Stem Cells in Small Cell Lung Cancer Cell Line H446: Higher Dependency on Oxidative Phosphorylation and Mitochondrial Substrate-Level Phosphorylation than Non-Stem Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Cuicui Gao

    Full Text Available Recently, targeting cancer stem cells (CSCs metabolism is becoming a promising therapeutic approach to improve cancer treatment outcomes. However, knowledge of the metabolic state of CSCs in small cell lung cancer is still lacking. In this study, we found that CSCs had significantly lower oxygen consumption rate and extracellular acidification rate than non-stem cancer cells. Meanwhile, this subpopulation of cells consumed less glucose, produced less lactate and maintained lower ATP levels. We also revealed that CSCs could produce more ATP through mitochondrial substrate-level phosphorylation during respiratory inhibition compared with non-stem cancer cells. Furthermore, they were more sensitive to suppression of oxidative phosphorylation. Therefore, oligomycin (inhibitor of oxidative phosphorylation could severely impair sphere-forming and tumor-initiating abilities of CSCs. Our work suggests that CSCs represent metabolically inactive tumor subpopulations which sustain in a state showing low metabolic activity. However, mitochondrial substrate-level phosphorylation of CSCs may be more active than that of non-stem cancer cells. Moreover, CSCs showed preferential use of oxidative phosphorylation over glycolysis to meet their energy demand. These results extend our understanding of CSCs metabolism, potentially providing novel treatment strategies targeting metabolic pathways in small cell lung cancer.

  1. Cytokine levels correlate with immune cell infiltration after anti-VEGF therapy in preclinical mouse models of breast cancer.

    Directory of Open Access Journals (Sweden)

    Christina L Roland

    Full Text Available The effect of blocking VEGF activity in solid tumors extends beyond inhibition of angiogenesis. However, no studies have compared the effectiveness of mechanistically different anti-VEGF inhibitors with respect to changes in tumor growth and alterations in the tumor microenvironment. In this study we use three distinct breast cancer models, a MDA-MB-231 xenograft model, a 4T1 syngenic model, and a transgenic model using MMTV-PyMT mice, to explore the effects of various anti-VEGF therapies on tumor vasculature, immune cell infiltration, and cytokine levels. Tumor vasculature and immune cell infiltration were evaluated using immunohistochemistry. Cytokine levels were evaluated using ELISA and electrochemiluminescence. We found that blocking the activation of VEGF receptor resulted in changes in intra-tumoral cytokine levels, specifically IL-1beta, IL-6 and CXCL1. Modulation of the level these cytokines is important for controlling immune cell infiltration and ultimately tumor growth. Furthermore, we demonstrate that selective inhibition of VEGF binding to VEGFR2 with r84 is more effective at controlling tumor growth and inhibiting the infiltration of suppressive immune cells (MDSC, Treg, macrophages while increasing the mature dendritic cell fraction than other anti-VEGF strategies. In addition, we found that changes in serum IL-1beta and IL-6 levels correlated with response to therapy, identifying two possible biomarkers for assessing the effectiveness of anti-VEGF therapy in breast cancer patients.

  2. Distinct expression levels and patterns of stem cell marker, aldehyde dehydrogenase isoform 1 (ALDH1, in human epithelial cancers.

    Directory of Open Access Journals (Sweden)

    Shan Deng

    Full Text Available Aldehyde dehydrogenase isoform 1 (ALDH1 has been proved useful for the identification of cancer stem cells. However, our knowledge of the expression and activity of ALDH1 in common epithelial cancers and their corresponding normal tissues is still largely absent. Therefore, we characterized ALDH1 expression in 24 types of normal tissues and a large collection of epithelial tumor specimens (six cancer types, n = 792 by immunohistochemical staining. Using the ALDEFUOR assay, ALDH1 activity was also examined in 16 primary tumor specimens and 43 established epithelial cancer cell lines. In addition, an ovarian cancer transgenic mouse model and 7 murine ovarian cancer cell lines were analyzed. We found that the expression levels and patterns of ALDH1 in epithelial cancers are remarkably distinct, and they correlate with their corresponding normal tissues. ALDH1 protein expression levels are positively correlated with ALDH1 enzymatic activity measured by ALDEFLUOR assay. Long-term in vitro culture doesn't significantly affect ALDH1 activity in epithelial tumor cells. Consistent with research on other cancers, we found that high ALDH1 expression is significantly associated with poor clinical outcomes in serous ovarian cancer patients (n = 439, p = 0.0036. Finally, ALDH(br tumor cells exhibit cancer stem cell properties and are resistant to chemotherapy. As a novel cancer stem cell marker, ALDH1 can be used for tumors whose corresponding normal tissues express ALDH1 in relatively restricted or limited levels such as breast, lung, ovarian or colon cancer.

  3. Discrete levels of Twist activity are required to direct distinct cell functions during gastrulation and somatic myogenesis.

    Directory of Open Access Journals (Sweden)

    Ming-Ching Wong

    Full Text Available Twist (Twi, a conserved basic helix-loop-helix transcriptional regulator, directs the epithelial-to-mesenchymal transition (EMT, and regulates changes in cell fate, cell polarity, cell division and cell migration in organisms from flies to humans. Analogous to its role in EMT, Twist has been implicated in metastasis in numerous cancer types, including breast, pancreatic and prostate. In the Drosophila embryo, Twist is essential for discrete events in gastrulation and mesodermal patterning. In this study, we derive a twi allelic series by examining the various cellular events required for gastrulation in Drosophila. By genetically manipulating the levels of Twi activity during gastrulation, we find that coordination of cell division is the most sensitive cellular event, whereas changes in cell shape are the least sensitive. Strikingly, we show that by increasing levels of Snail expression in a severe twi hypomorphic allelic background, but not a twi null background, we can reconstitute gastrulation and produce viable adult flies. Our results demonstrate that the level of Twi activity determines whether the cellular events of ventral furrow formation, EMT, cell division and mesodermal migration occur.

  4. High level of chromosomal instability in circulating tumor cells of ROS1-rearranged non-small-cell lung cancer

    OpenAIRE

    Pailler, E.; Auger, N.; Lindsay, C. R.; Vielh, P; Islas-Morris-Hernandez, A.; Borget, I; Ngo-Camus, M.; Planchard, D.; Soria, J.-C.; Besse, B.; Farace, F.

    2015-01-01

    Background Genetic aberrations affecting the c-ros oncogene 1 (ROS1) tyrosine kinase gene have been reported in a small subset of patients with non-small-cell lung cancer (NSCLC). We evaluated whether ROS1-chromosomal rearrangements could be detected in circulating tumor cells (CTCs) and examined tumor heterogeneity of CTCs and tumor biopsies in ROS1-rearranged NSCLC patients. Patients and methods Using isolation by size of epithelial tumor cells (ISET) filtration and filter-adapted-fluoresce...

  5. Does MW Radiation Affect Gene Expression, Apoptotic Level, and Cell Cycle Progression of Human SH-SY5Y Neuroblastoma Cells?

    Science.gov (United States)

    Kayhan, Handan; Esmekaya, Meric Arda; Saglam, Atiye Seda Yar; Tuysuz, Mehmed Zahid; Canseven, Ayşe Gulnihal; Yagci, Abdullah Munci; Seyhan, Nesrin

    2016-06-01

    Neuroblastoma (NB) is a cancer that occurs in sympathetic nervous system arising from neuroblasts and nerve tissue of the adrenal gland, neck, chest, or spinal cord. It is an embryonal malignancy and affects infants and children. In this study, we investigated the effects of microwave (MW) radiation on apoptotic activity, cell viability, and cell cycle progression in human SH-SY5Y NB cells which can give information about MW radiation effects on neural cells covering the period from the embryonic stages to infants. SH-SY5Y NB cells were exposed to 2.1 GHz W-CDMA modulated MW radiation for 24 h at a specific absorption rate of 0.491 W/kg. Control samples were in the same conditions with MW-exposed samples but they were not exposed to MW radiation. The apoptotic activity of cells was measured by Annexin-V-FITC and propidium iodide staining. Moreover, mRNA levels of proliferative and cell cycle proteins were determined by real-time RT-PCR. The change in cell cycle progression was observed by using CycleTest-Plus DNA reagent. No significant change was observed in apoptotic activity of MW-exposed cells compared to control cells. The mRNA levels of c-myc and cyclin D1 were significantly reduced in MW group (p CDMA modulated MW radiation did not cause apoptotic cell death but changed cell cycle progression. PMID:27260669

  6. Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels.

    Science.gov (United States)

    Smith, M Ryan; Vayalil, Praveen K; Zhou, Fen; Benavides, Gloria A; Beggs, Reena R; Golzarian, Hafez; Nijampatnam, Bhavitavya; Oliver, Patsy G; Smith, Robin A J; Murphy, Michael P; Velu, Sadanandan E; Landar, Aimee

    2016-08-01

    Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study

  7. Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels

    Directory of Open Access Journals (Sweden)

    M. Ryan Smith

    2016-08-01

    Full Text Available Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP, decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231 breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC protein levels, although other protein levels were

  8. Analyzing the Expression Level and Cellular Location of the Tip-1 Protein in Oral Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    N Mansoursamaei

    2005-10-01

    Full Text Available Oral squamous cell carcinoma (OSCC is the sixth most common cancer in the world and accounts for approximately 4% of all cancers and 2% of all cancer death.The single most important factor in the prevention of the disease is early detection although, due to increased risk of secondary malignancy, survival remains poor with only a 25% 5 years survival. Both hereditary and environmental factors have been shown to have a productive role in this disease. For example, although chronic exposure of oral epithelium to tobacco smoke and alcohol are amongst the most important aetiological factors, it is now becoming realized that infection with high risk types of human papilloma virus (HPV are also involved as causative agent in a subset of this disease. All of these OSCC associated factors are known to promote genetic instability in the target oral epithelial cells. Work in our laboratories has indicated that the Tax interacting protein 1 (Tip-1 is also a target for the HPV 16 E6 protein may play an important role in controlling genetic instability during the oncogenic process (Hampson L., et al unpublished. So far 14 oral cancer cell lines have been grown in cell culture and RNA extracted from these. Tip-1 transcript levels were analyzed in this material by Northern blotting and competitive template quantitative PCR, which showed that Tip-1 levels were higher in some, cell lines than others (4 high, 6 moderate, 4 low level. Cell ploidy was determined by FACS analysis of propidium iodide stained cells, which showed that out of all the OSCC cell lines tested the cell line (BICR 68 had the greatest numbers of polyploid cells and also had the highest expression of Tip-1 RNA.

  9. Microenvironmental geometry guides platelet adhesion and spreading: a quantitative analysis at the single cell level.

    Directory of Open Access Journals (Sweden)

    Ashley Kita

    Full Text Available To activate clot formation and maintain hemostasis, platelets adhere and spread onto sites of vascular injury. Although this process is well-characterized biochemically, how the physical and spatial cues in the microenvironment affect platelet adhesion and spreading remain unclear. In this study, we applied deep UV photolithography and protein micro/nanostamping to quantitatively investigate and characterize the spatial guidance of platelet spreading at the single cell level and with nanoscale resolution. Platelets adhered to and spread only onto micropatterned collagen or fibrinogen surfaces and followed the microenvironmental geometry with high fidelity and with single micron precision. Using micropatterned lines of different widths, we determined that platelets are able to conform to micropatterned stripes as thin as 0.6 µm and adopt a maximum aspect ratio of 19 on those protein patterns. Interestingly, platelets were also able to span and spread over non-patterned regions of up to 5 µm, a length consistent with that of maximally extended filopodia. This process appears to be mediated by platelet filopodia that are sensitive to spatial cues. Finally, we observed that microenvironmental geometry directly affects platelet biology, such as the spatial organization and distribution of the platelet actin cytoskeleton. Our data demonstrate that platelet spreading is a finely-tuned and spatially-guided process in which spatial cues directly influence the biological aspects of how clot formation is regulated.

  10. Significance of salivary phosphodiesterase level in oral squamous cell carcinoma patients

    Directory of Open Access Journals (Sweden)

    Yousef Rezaei Chianeh

    2013-08-01

    Full Text Available Oral cancer, more specifically oral squamous cell carcinoma (OSCC consider as common cancer that 300,000 people diagnosed per year worldwide. The only effective treatment for OSCC is surgical intervention. Over the past two decades, overall disease condition has not improved although advancement of treatment has considerably increased. The phosphodiesterase (PDEs are responsible for the hydrolysis of the second messengers with a fundamental role in the transduction of the intracellular signals. In numerous pathological conditions such as cellular differentiation, apoptosis, and tumor invasivity the different PDF activity has been observed that shown role in pathophysiological mechanism. The role of PDEs as an intervention factor for activation of angiogenesis by influencing a tumor growth has been shown. The objective of this study was to estimate and compare salivary PDEs levels in healthy controls and biopsy-proven oral cancer patients before definitive therapy. Study was done in patients age between 25-65 years biopsy proven oral cancer patients and control group. After obtaining prior consent from biopsy-proven oral cancer patients (n=26 (before onset of any definitive treatment and age- and sex-matched healthy controls (n=29, salivary sample was collected for estimation of the activity of phosphodiesterases (PDEs. [Int J Res Med Sci 2013; 1(4.000: 417-420

  11. Educational level of patients with germ cell tumor radiated in childhood

    International Nuclear Information System (INIS)

    In order to estimate the influence of radiotherapy on the intellectual development of children with brain tumor, we investigated the educational level of 21 patients with germ cell tumor who had undergone radiotherapy. They were divided into three groups in accordance with their age at the time of radiation; under school age group (under 6 years of age), elementary school age group (from 7 to 12 years of age), and junior high and high school age group (from 13 to 18 years of age). There were 2 cases in the under school age group, one of them graduated from high school and the other is presently a junior high school student. There were 5 cases in the elementary school age group. Three of these graduated from university, 1 is presently a university student and 1 is a high school student. There were 14 cases in the junior high and high school age group. Two of these are university students, 7 graduated from high school, 1 is presently a junior high school student, and 4 died because of tumor progression. The mean period of hospitalization of the patients who have been admitted to university was 63.0 days, and that of patients who have not been admitted university was 135 days. There is a statistical difference (p<0.05). It could be concluded that the period of hospitalization rather than radiotherapy seemed to influence the educational status of children with brain tumor. (author)

  12. Increased interferon alpha receptor 2 mRNA levels is associated with renal cell carcinoma metastasis

    Directory of Open Access Journals (Sweden)

    Yamanishi Tomonori

    2007-08-01

    Full Text Available Abstract Background Interferon-α (IFN-α is one of the central agents in immunotherapy for renal cell carcinoma (RCC and binds to the IFN-α receptor (IFNAR. We investigated the role of IFNAR in RCC. Methods We quantified IFNAR mRNA expression in paired tumor and non-tumor samples from the surgical specimens of 103 consecutive patients with RCC using a real-time reverse transcription polymerase chain reaction (RT-PCR, and IFNAR2 protein using Western blotting. Results The absolute level of IFNAR1 and IFNAR2 mRNAs in tumor and non-tumor tissues did not correlate with the malignant and metastatic profiles. The relative yields of the PCR product from the tumor tissue to that from the corresponding non-tumor tissue (T/N for the expression of IFNAR mRNAs were calculated. While the T/N ratio of IFNAR1 did not correlate with any factor, a high T/N ratio of IFNAR2 correlated with poor differentiation (P P P P P Conclusion IFNAR2 is associated with the progression of RCC.

  13. A microbial fuel cell driven capacitive deionization technology for removal of low level dissolved ions.

    Science.gov (United States)

    Feng, Cuijie; Hou, Chia-Hung; Chen, Shaohua; Yu, Chang-Ping

    2013-04-01

    The microbial fuel cell (MFC) is an emerging technology, which uses exoelectrogenic microorganisms to oxidize organic matter in the wastewater to produce electricity. However, the low energy output limits its application in practice. Capacitive deionization (CDI), an electrochemically controlled method for deionization by the adsorption of ions in the electrical double layer region at an electrode-solution interface, requires a low external power supply. Therefore, in this study, we investigated the MFC driven CDI (MFC-CDI) technology to integrate deionization with wastewater treatment and electricity production. Taking advantage of the low potential requirement of CDI, voltage generated from a continuous flow MFC could be used to drive the CDI to achieve removal of the electrolyte to a stable status. The results indicated that among the three connection types of MFCs including single-, series-, and parallel-configuration, the parallel connection of two MFCs resulted in the highest potential (0.63V) applied to CDI and the conductivity removal of NaCl solution was more than 60%. The electrosorption capacities under different electrolyte concentrations of 50, 100 and 150 mg L(-1) were 150, 346 and 295 μg g(-1), respectively. These results suggest that the new MFC-CDI technology, which utilizes energy recovery from the wastewater, has great potential to be an energy saving technology to remove low level dissolved ions from aqueous solutions for the water and wastewater treatment processes. PMID:23375820

  14. Deciphering Mineral Homeostasis in Barley Seed Transfer Cells at Transcriptional Level.

    Directory of Open Access Journals (Sweden)

    Behrooz Darbani

    Full Text Available In addition to the micronutrient inadequacy of staple crops for optimal human nutrition, a global downtrend in crop-quality has emerged from intensive breeding for yield. This trend will be aggravated by elevated levels of the greenhouse gas carbon dioxide. Therefore, crop biofortification is inevitable to ensure a sustainable supply of minerals to the large part of human population who is dietary dependent on staple crops. This requires a thorough understanding of plant-mineral interactions due to the complexity of mineral homeostasis. Employing RNA sequencing, we here communicate transfer cell specific effects of excess iron and zinc during grain filling in our model crop plant barley. Responding to alterations in mineral contents, we found a long range of different genes and transcripts. Among them, it is worth to highlight the auxin and ethylene signaling factors Arfs, Abcbs, Cand1, Hps4, Hac1, Ecr1, and Ctr1, diurnal fluctuation components Sdg2, Imb1, Lip1, and PhyC, retroelements, sulfur homeostasis components Amp1, Hmt3, Eil3, and Vip1, mineral trafficking components Med16, Cnnm4, Aha2, Clpc1, and Pcbps, and vacuole organization factors Ymr155W, RabG3F, Vps4, and Cbl3. Our analysis introduces new interactors and signifies a broad spectrum of regulatory levels from chromatin remodeling to intracellular protein sorting mechanisms active in the plant mineral homeostasis. The results highlight the importance of storage proteins in metal ion toxicity-resistance and chelation. Interestingly, the protein sorting and recycling factors Exoc7, Cdc1, Sec23A, and Rab11A contributed to the response as well as the polar distributors of metal-transporters ensuring the directional flow of minerals. Alternative isoform switching was found important for plant adaptation and occurred among transcripts coding for identical proteins as well as transcripts coding for protein isoforms. We also identified differences in the alternative-isoform preference between

  15. Inverse associations between obesity indicators and thymic T-cell production levels in aging atomic-bomb survivors.

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    Kengo Yoshida

    Full Text Available Reduction of the naive T-cell population represents a deteriorating state in the immune system that occurs with advancing age. In animal model studies, obesity compromises the T-cell immune system as a result of enhanced adipogenesis in primary lymphoid organs and systemic inflammation. In this study, to test the hypothesis that obesity may contribute to the aging of human T-cell immunity, a thousand atomic-bomb survivors were examined for obesity status and ability to produce naive T cells, i.e., T-cell receptor excision circle (TREC numbers in CD4 and CD8 T cells. The number of TRECs showed a strong positive correlation with naive T cell numbers, and lower TREC numbers were associated with higher age. We found that the TREC number was inversely associated with levels of obesity indicators (BMI, hemoglobin A1c and serum CRP levels. Development of type-2 diabetes and fatty liver was also associated with lower TREC numbers. This population study suggests that obesity with enhanced inflammation is involved in aging of the human T-cell immune system. Given the fact that obesity increases the risk of numerous age-related diseases, attenuated immune competence is a possible mechanistic link between obesity and disease development among the elderly.

  16. Inverse associations between obesity indicators and thymic T-cell production levels in aging atomic-bomb survivors.

    Science.gov (United States)

    Yoshida, Kengo; Nakashima, Eiji; Kubo, Yoshiko; Yamaoka, Mika; Kajimura, Junko; Kyoizumi, Seishi; Hayashi, Tomonori; Ohishi, Waka; Kusunoki, Yoichiro

    2014-01-01

    Reduction of the naive T-cell population represents a deteriorating state in the immune system that occurs with advancing age. In animal model studies, obesity compromises the T-cell immune system as a result of enhanced adipogenesis in primary lymphoid organs and systemic inflammation. In this study, to test the hypothesis that obesity may contribute to the aging of human T-cell immunity, a thousand atomic-bomb survivors were examined for obesity status and ability to produce naive T cells, i.e., T-cell receptor excision circle (TREC) numbers in CD4 and CD8 T cells. The number of TRECs showed a strong positive correlation with naive T cell numbers, and lower TREC numbers were associated with higher age. We found that the TREC number was inversely associated with levels of obesity indicators (BMI, hemoglobin A1c) and serum CRP levels. Development of type-2 diabetes and fatty liver was also associated with lower TREC numbers. This population study suggests that obesity with enhanced inflammation is involved in aging of the human T-cell immune system. Given the fact that obesity increases the risk of numerous age-related diseases, attenuated immune competence is a possible mechanistic link between obesity and disease development among the elderly. PMID:24651652

  17. Levels of circulating CD45dimCD34+VEGFR2+ progenitor cells correlate with outcome in metastatic renal cell carcinoma patients treated with tyrosine kinase inhibitors

    Science.gov (United States)

    Farace, F; Gross-Goupil, M; Tournay, E; Taylor, M; Vimond, N; Jacques, N; Billiot, F; Mauguen, A; Hill, C; Escudier, B

    2011-01-01

    Background: Predicting the efficacy of antiangiogenic therapy would be of clinical value in patients (pts) with metastatic renal cell carcinoma (mRCC). We tested the hypothesis that circulating endothelial cell (CEC), bone marrow-derived CD45dimCD34+VEGFR2+ progenitor cell or plasma angiogenic factor levels are associated with clinical outcome in mRCC pts undergoing treatment with tyrosine kinase inhibitors (TKI). Methods: Fifty-five mRCC pts were prospectively monitored at baseline (day 1) and day 14 during treatment (46 pts received sunitinib and 9 pts received sorafenib). Circulating endothelial cells (CD45−CD31+CD146+7-amino-actinomycin (7AAD)− cells) were measured in 1 ml whole blood using four-color flow cytometry (FCM). Circulating CD45dimCD34+VEGFR2+7AAD− progenitor cells were measured in progenitor-enriched fractions by four-color FCM. Plasma VEGF, sVEGFR2, SDF-1α and sVCAM-1 levels were determined by ELISA. Correlations between baseline CEC, CD45dimCD34+VEGFR2+7AAD− progenitor cells, plasma factors, as well as day 1–day 14 changes in CEC, CD45dimCD34+VEGFR2+7AAD− progenitor, plasma factor levels, and response to TKI, progression-free survival (PFS) and overall survival (OS) were examined. Results: No significant correlation between markers and response to TKI was observed. No association between baseline CEC, plasma VEGF, sVEGFR-2, SDF-1α, sVCAM-1 levels with PFS and OS was observed. However, baseline CD45dimCD34+VEGFR2+7AAD− progenitor cell levels were associated with PFS (P=0.01) and OS (P=0.006). Changes in this population and in SDF-1α levels between day 1 and day 14 were associated with PFS (P=0.03, P=0.002). Changes in VEGF and SDF-1α levels were associated with OS (P=0.02, P=0.007). Conclusion: Monitoring CD45dimCD34+VEGFR2+ progenitor cells, plasma VEGF and SDF-1α levels could be of clinical interest in TKI-treated mRCC pts to predict outcome. PMID:21386843

  18. Smoking decreases the level of circulating CD34+ progenitor cells in young healthy women - a pilot study

    Directory of Open Access Journals (Sweden)

    Baumann Gert

    2010-05-01

    Full Text Available Abstract Background Decreased levels of circulating bone marrow-derived progenitor cells have been associated with risk factors and cardiovascular diseases. Smoking is the most important modifiable risk factor for atherosclerosis in young women. The aim of this pilot study was to assess in healthy premenopausal women without other risk factors for cardiovascular disease the influence of nicotine abuse on the number of circulating progenitor cells in relation to endothelial function. Methods The number of endothelial progenitor cells, measured as colony-forming units in a cell-culture assay (EPC-CFU and the number of circulating CD34 + and CD34 + /CD133 + cells, measured by flow cytometry, was estimated in 32 women at the menstrual phase of the menstrual cycle. In addition, flow-mediated dilation (FMD was assessed as a marker for vascular function. In a subgroup of these women (n = 20, progenitor cells were also investigated at the mid-follicular and luteal phases of the menstrual cycle. Results Compared to non-smokers, the abundance of circulating CD34 + cells was significantly lower in smoking women in the menstrual, mid-luteal, and mid-follicular phases of the menstrual cycle. The number of CD34 + progenitor cells was revealed to have significant positive correlation with FMD in young healthy women, whereas CD34 + /CD133 + progenitor cells and EPC-CFU showed no significant correlation. Conclusion The number of CD34 + progenitor cells positively correlates with FMD in young healthy women and is decreased by smoking.

  19. Effect of Low Level Laser Therapy on Proliferation and Differentiation of the Cells Contributing in Bone Regeneration

    OpenAIRE

    AMID, Reza; Kadkhodazadeh, Mahdi; Ahsaie, Mitra Ghazizadeh; Hakakzadeh, Arian

    2014-01-01

    Introduction: Low level laser therapy (LLLT) also known as photobiomodulation, is a treatment that uses low-level lasers or light-emitting diodes (LEDs) to change cellular function and is a clinically well accepted tool in regenerative medicine and dentistry. Considering the variety of laser, exposure, cells and study types, the exact effects of low level laser therapy seems to be unclear. The aim of this study was to review the data published in the field of the effects of low level laser th...

  20. Elevated fluoride levels and periostitis in pediatric hematopoietic stem cell transplant recipients receiving long-term voriconazole.

    Science.gov (United States)

    Tarlock, Katherine; Johnson, Darren; Cornell, Cathy; Parnell, Shawn; Meshinchi, Soheil; Baker, K Scott; Englund, Janet A

    2015-05-01

    Azole therapy is widely utilized in hematopoietic stem cell transplant (HCT) recipients for the treatment of aspergillus. Complications of voriconazole treatment related to its elevated fluoride content have been described in adults, including reports of symptomatic skeletal fluorosis. We review fluoride levels, clinical, and laboratory data in five pediatric HCT recipients on long-term voriconazole therapy, all found to have elevated serum fluoride levels. Two patients had toxic fluoride levels, one infant had symptoms of significant pain with movement and radiographs confirmed skeletal fluorosis. Monitoring fluoride levels in children, especially with skeletal symptoms, should be considered in patients on long-term voriconazole. PMID:25327935

  1. GnRH receptor activation competes at a low level with growth signaling in stably transfected human breast cell lines

    International Nuclear Information System (INIS)

    Gonadotrophin releasing hormone (GnRH) analogs lower estrogen levels in pre-menopausal breast cancer patients. GnRH receptor (GnRH-R) activation also directly inhibits the growth of certain cells. The applicability of GnRH anti-proliferation to breast cancer was therefore analyzed. GnRH-R expression in 298 primary breast cancer samples was measured by quantitative immunofluorescence. Levels of functional GnRH-R in breast-derived cell lines were assessed using 125I-ligand binding and stimulation of 3H-inositol phosphate production. Elevated levels of GnRH-R were stably expressed in cells by transfection. Effects of receptor activation on in vitro cell growth were investigated in comparison with IGF-I and EGF receptor inhibition, and correlated with intracellular signaling using western blotting. GnRH-R immunoscoring was highest in hormone receptor (triple) negative and grade 3 breast tumors. However prior to transfection, functional endogenous GnRH-R were undetectable in four commonly studied breast cancer cell lines (MCF-7, ZR-75-1, T47D and MDA-MB-231). After transfection with GnRH-R, high levels of cell surface GnRH-R were detected in SVCT and MDA-MB-231 clones while low-moderate levels of GnRH-R occurred in MCF-7 clones and ZR-75-1 clones. MCF-7 sub-clones with high levels of GnRH-R were isolated following hygromycin phosphotransferase transfection. High level cell surface GnRH-R enabled induction of high levels of 3H-inositol phosphate and modest growth-inhibition in SVCT cells. In contrast, growth of MCF-7, ZR-75-1 or MDA-MB-231 clones was unaffected by GnRH-R activation. Cell growth was inhibited by IGF-I or EGF receptor inhibitors. IGF-I receptor inhibitor lowered levels of p-ERK1/2 in MCF-7 clones. Washout of IGF-I receptor inhibitor resulted in transient hyper-elevation of p-ERK1/2, but co-addition of GnRH-R agonist did not alter the dynamics of ERK1/2 re-phosphorylation. Breast cancers exhibit a range of GnRH-R immunostaining, with higher levels of

  2. Study on Fuel Cell Network System Considering Reduction in Fuel Cell Capacity Using Load Leveling and Heat Release Loss

    Science.gov (United States)

    Obara, Shin'ya; Kudo, Kazuhiko

    Reduction in fuel cell capacity linked to a fuel cell network system is considered. When the power demand of the whole network is small, some of the electric power generated by the fuel cell is supplied to a water electrolysis device, and hydrogen and oxygen gases are generated. Both gases are compressed with each compressor and they are stored in cylinders. When the electric demand of the whole network is large, both gases are supplied to the network, and fuel cells are operated by these hydrogen and oxygen gases. Furthermore, an optimization plan is made to minimize the quantity of heat release of the hot water piping that connects each building. Such an energy network is analyzed assuming connection of individual houses, a hospital, a hotel, a convenience store, an office building, and a factory. Consequently, compared with the conventional system, a reduction of 46% of fuel cell capacity is expected.

  3. Transient recombinant protein expression in mammalian cells: the role of mRNA level and stability

    OpenAIRE

    Wulhfard, Sarah

    2009-01-01

    Transient gene expression (TGE) is a rapid method for generating recombinant proteins in mammalian cells, but the volumetric productivities for secreted proteins in transiently transfected CHO DG44 cells are typically more than an order of magnitude lower than the yields achieved with recombinant CHO-derived cell lines. The goals of the thesis are to identify the limitations to higher TGE yields in CHO DG44 cells and to find possible solutions to overcome the problems. Initially an attempt wa...

  4. Mast cell mediator tryptase levels after inhalation or intravenous administration of high doses pharmaceutically prepared heroin

    NARCIS (Netherlands)

    E.J. Rook; A.P. van Zanten; W. van den Brink; J.M. van Ree; J.H. Beijnen

    2006-01-01

    Background: Opioids like morphine and heroin induce mast cell degranulation in vitro. The release of mast cell mediators like histamine and tryptase may lead to allergic symptoms. In this study it was investigated whether mast cell mediator release also occurs in vivo in addicted patients who partic

  5. Gene induction during differentiation of human monocytes into dendritic cells: an integrated study at the RNA and protein levels

    OpenAIRE

    Angénieux, Catherine; Fricker, Dominique; Strub, Jean-Marc; Luche, Sylvie; Bausinger, Huguette; Cazenave, Jean-Pierre; Van Dorsselaer, Alain; Hanau, Daniel; de la Salle, Henri; Rabilloud, Thierry

    2006-01-01

    Changes in gene expression occurring during differentiation of human monoytes into dendritic cells were studied at the RNA and protein levels. These studies showed the induction of several gene classes corresponding to various biological functions. These functions encompass of course antigen processing and presentation, cytoskeleton, cell signalling and signal transduction, but also an increase of mitochondrial function and of the protein synthesis machinery, including some, but not all, chap...

  6. Heavy Water Reduces GFP Expression in Prokaryotic Cell-Free Assays at the Translation Level While Stimulating Its Transcription

    OpenAIRE

    Hohlefelder, Luisa S.; Tobias Stögbauer; Madeleine Opitz; Bayerl, Thomas M.; Joachim O. Rädler

    2013-01-01

    The in vitro proliferation of prokaryotic and eukaryotic cells is remarkably hampered in the presence of heavy water (D2O). Impairment of gene expression at the transcription or translation level can be the base for this effect. However, insights into the underlying mechanisms are lacking. Here, we employ a cell-free expression system for the quantitative analysis of the effect of increasing percentages of D2O on the kinetics of in-vitro GFP expression. Experiments are designed to discriminat...

  7. Changes in Blood B Cell-Activating Factor (BAFF Levels in Multiple Sclerosis: A Sign of Treatment Outcome.

    Directory of Open Access Journals (Sweden)

    Karin Kannel

    Full Text Available Multiple sclerosis (MS is mediated primarily by autoreactive T cells. However, evidence suggesting the involvement of humoral immunity in brain diseases has increased interest in the role of B cells and their products during MS pathogenesis. The major survival factor for B cells, BAFF has been shown to play a role in several autoimmune conditions. Elevated BAFF levels have been reported in MS animal model and during MS relapse in patients. Moreover, disease-modifying treatments (DMT reportedly influence blood BAFF levels in MS patients, but the significance of these changes remains unclear. The present study addresses how blood BAFF levels are associated with the clinical course of relapsing-remitting MS and the effectiveness of DMT and short-term steroid treatment. During a prospective longitudinal follow-up of 2.3 years, BAFF was measured in the blood of 170 MS patients in the stable phase and within 186 relapses. BAFF levels were significantly higher in MS patients compared to healthy controls. However, stable MS patients without relapses exhibited significantly higher BAFF levels than relapsing patients. Treatment with interferon-β and immunosuppressants raised BAFF blood levels. Interestingly, a similar effect was not seen in patients treated with glatiramer acetate. Short-term treatment with high doses of intravenous methylprednisolone did not significantly alter plasma BAFF levels in 65% of relapsing-remitting MS patients. BAFF were correlated weakly but significantly with monocyte and basophil counts, but not with other blood cell types (neutrophils, lymphocytes, or eosinophils or inflammatory biomarkers. To our knowledge, this is the first report demonstrating that higher blood BAFF levels may reflect a more stable and effective MS treatment outcome. These results challenge hypotheses suggesting that elevated blood BAFF levels are associated with more severe disease presentation and could explain the recent failure of pharmaceutical

  8. Low-level laser therapy: Effects on human face aged skin and cell viability of HeLa cells exposed to UV radiation

    OpenAIRE

    Mezghani Sana; Hammami Amira; Amri Mohamed

    2015-01-01

    Chronic and excessive exposure to UV radiation leads to photoaging and photocarcinogenesis. Adequate protection of the skin against the deleterious effects of UV irradiation is essential. Low-level laser therapy (LLLT) is a light source in the red to near-infrared range that has been accepted in a variety of medical applications. In this study, we explored the effect of LLLT in human face aged skin and the cell viability of HeLa cells exposed to UV radiatio...

  9. TERRA Expression Levels Do Not Correlate With Telomere Length and Radiation Sensitivity in Human Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Alexandra eSmirnova

    2013-05-01

    Full Text Available Mammalian telomeres are transcribed into long non-coding telomeric RNA molecules (TERRA that seem to play a role in the maintenance of telomere stability. In human cells, CpG island promoters drive TERRA transcription and are regulated by methylation. It was suggested that the amount of TERRA may be related to telomere length. To test this hypothesis we measured telomere length and TERRA levels in single clones isolated from five human cell lines: HeLa (cervical carcinoma, BRC-230 (breast cancer, AKG and GK2 (gastric cancers and GM847 (SV40 immortalized skin fibroblasts. We observed great clonal heterogeneity both in TRF (Terminal Restriction Fragment length and in TERRA levels. However, these two parameters did not correlate with each other. Moreover, cell survival to γ-rays did not show a significant variation among the clones, suggesting that, in this cellular system, the intra-population variability in telomere length and TERRA levels does not influence sensitivity to ionizing radiation. This conclusion was supported by the observation that in a cell line in which telomeres were greatly elongated by the ectopic expression of telomerase, TERRA expression levels and radiation sensitivity were similar to the parental HeLa cell line.

  10. Probing the energy levels of perovskite solar cells via Kelvin probe and UV ambient pressure photoemission spectroscopy.

    Science.gov (United States)

    Harwell, J R; Baikie, T K; Baikie, I D; Payne, J L; Ni, C; Irvine, J T S; Turnbull, G A; Samuel, I D W

    2016-07-20

    The field of organo-lead halide perovskite solar cells has been rapidly growing since their discovery in 2009. State of the art devices are now achieving efficiencies comparable to much older technologies like silicon, while utilising simple manufacturing processes and starting materials. A key parameter to consider when optimising solar cell devices or when designing new materials is the position and effects of the energy levels in the materials. We present here a comprehensive study of the energy levels present in a common structure of perovskite solar cell using an advanced macroscopic Kelvin probe and UV air photoemission setup. By constructing a detailed map of the energy levels in the system we are able to predict the importance of each layer to the open circuit voltage of the solar cell, which we then back up through measurements of the surface photovoltage of the cell under white illumination. Our results demonstrate the effectiveness of air photoemission and Kelvin probe contact potential difference measurements as a method of identifying the factors contributing to the open circuit voltage in a solar cell, as well as being an excellent way of probing the physics of new materials. PMID:27384817

  11. Exposure to low level GSM 935 MHz radiofrequency fields does not induce apoptosis in proliferating or differentiated murine neuroblastoma cells

    International Nuclear Information System (INIS)

    The aim of this study was to investigate whether radiofrequency (RF) fields characteristic of mobile phones at non-thermal levels can induce apoptosis in murine neuroblastoma (N2a) cells in both proliferating and differentiated states. Cells were exposed continuously for 24 h to one of the three 935-MHz RF signals: global system for mobile communication (GSM) basic, GSM talk and a continuous wave, unmodulated signal; all at a specific energy absorption rate of 2 W kg-1. The measured increase in temperature of the cells due to the RF fields was around 0.06 deg. C. At a number of time points between 0 and 48 h post-exposure, the cells were assessed for apoptosis under a fluorescence microscope using three independent assays: Annexin V, caspase activation and in situ end-labelling. No statistically significant differences in apoptosis levels were observed between the exposed and sham-exposed cells using the three assays at any time point post-exposure. These data suggest that RF exposures, characteristic of GSM mobile phones, do not significantly affect the apoptosis levels in proliferating and differentiated murine neuroblastoma cell line N2a. (authors)

  12. A systems level strategy for analyzing the cell death network: implication in exploring the apoptosis/autophagy connection.

    Science.gov (United States)

    Zalckvar, E; Yosef, N; Reef, S; Ber, Y; Rubinstein, A D; Mor, I; Sharan, R; Ruppin, E; Kimchi, A

    2010-08-01

    The mammalian cell death network comprises three distinct functional modules: apoptosis, autophagy and programmed necrosis. Currently, the field lacks systems level approaches to assess the extent to which the intermodular connectivity affects cell death performance. Here, we developed a platform that is based on single and double sets of RNAi-mediated perturbations targeting combinations of apoptotic and autophagic genes. The outcome of perturbations is measured both at the level of the overall cell death responses, using an unbiased quantitative reporter, and by assessing the molecular responses within the different functional modules. Epistatic analyses determine whether seemingly unrelated pairs of proteins are genetically linked. The initial running of this platform in etoposide-treated cells, using a few single and double perturbations, identified several levels of connectivity between apoptosis and autophagy. The knock down of caspase3 turned on a switch toward autophagic cell death, which requires Atg5 or Beclin-1. In addition, a reciprocal connection between these two autophagic genes and apoptosis was identified. By applying computational tools that are based on mining the protein-protein interaction database, a novel biochemical pathway connecting between Atg5 and caspase3 is suggested. Scaling up this platform into hundreds of perturbations potentially has a wide, general scope of applicability, and will provide the basis for future modeling of the cell death network. PMID:20150916

  13. 17β-Estradiol treatment inhibits breast cell proliferation, migration and invasion by decreasing MALAT-1 RNA level

    International Nuclear Information System (INIS)

    Highlights: • E2 affects not only estrogen-receptor α positive breast cells but also negative ones. • 100 nM E2 treatment affects breast cells proliferation, migration. • 100 nM E2 treatment functions in an estrogen-receptor α-independent way. • E2 treatment decreases MALAT-1 RNA level by post-transcriptional regulation. - Abstract: Breast cancer cells, which express estrogen receptor α (ERα), respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. But breast cancer cells without ERα show no effect on low concentration of estrogen treatment. Proliferation, migration and invasion of MCF10a, MCF7 and MB231 cells treated with low (1 nM) or high (100 nM) dose of 17β-Estradiol (E2) was performed. We identified the effects of E2 on these breast cell lines, and looked for the difference in the presence and absence of ERα. Specifically, we looked for the changes of long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 (MALAT-1), which is found extensively and highly expressed in several kinds of tumor cells, including breast carcinoma. It was observed that proliferation, migration and invasion of breast cells were greatly affected by high concentration E2 treatment and were not affected by low concentration E2 treatment in an ERα independent way. We found that the high concentration E2 treatment largely decreased MALAT-1 RNA level. Interestingly, MALAT-1 decreasing by knocking down showed similar effects on proliferation, migration and invasion. E2 treatment affects breast tumor or non-tumor cells proliferation, migration and invasion in an ERα -independent, but a dose-dependent way by decreasing the MALAT-1 RNA level

  14. 17β-Estradiol treatment inhibits breast cell proliferation, migration and invasion by decreasing MALAT-1 RNA level

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Ziyi [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China); Chen, Changjin [Department of Pediatrics, West China Second University Hospital, Sichuan University, Chengdu 610041 (China); Liu, Yu [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China); Wu, Chuanfang, E-mail: 879413966@qq.com [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China)

    2014-03-07

    Highlights: • E2 affects not only estrogen-receptor α positive breast cells but also negative ones. • 100 nM E2 treatment affects breast cells proliferation, migration. • 100 nM E2 treatment functions in an estrogen-receptor α-independent way. • E2 treatment decreases MALAT-1 RNA level by post-transcriptional regulation. - Abstract: Breast cancer cells, which express estrogen receptor α (ERα), respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. But breast cancer cells without ERα show no effect on low concentration of estrogen treatment. Proliferation, migration and invasion of MCF10a, MCF7 and MB231 cells treated with low (1 nM) or high (100 nM) dose of 17β-Estradiol (E2) was performed. We identified the effects of E2 on these breast cell lines, and looked for the difference in the presence and absence of ERα. Specifically, we looked for the changes of long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 (MALAT-1), which is found extensively and highly expressed in several kinds of tumor cells, including breast carcinoma. It was observed that proliferation, migration and invasion of breast cells were greatly affected by high concentration E2 treatment and were not affected by low concentration E2 treatment in an ERα independent way. We found that the high concentration E2 treatment largely decreased MALAT-1 RNA level. Interestingly, MALAT-1 decreasing by knocking down showed similar effects on proliferation, migration and invasion. E2 treatment affects breast tumor or non-tumor cells proliferation, migration and invasion in an ERα -independent, but a dose-dependent way by decreasing the MALAT-1 RNA level.

  15. Elevated serum IL-10 levels in diffuse large B-cell lymphoma: a mechanism of aberrant JAK2 activation

    OpenAIRE

    Gupta, Mamta; Han, Jing Jing; Stenson, Mary; Maurer, Matthew; Wellik, Linda; Hu, Guangzhen; Ziesmer, Steve; Dogan, Ahmet; Witzig, Thomas E.

    2012-01-01

    Cytokines are deregulated in cancers and can contribute to tumor growth. In patients with diffuse large-cell lymphoma (DLBCL), we observed higher levels of JAK/STAT pathway-related serum cytokines (ie, IL-6, IL-10, epidermal growth factor, and IL-2) compared with controls. Of these, only IL-10 activated the JAK2 pathway in lymphoma cells in vitro. Patients with high serum IL-10 had shorter event-free survival (EFS) than patients with low levels (P > .01) and high IL-10 was correlated with hig...

  16. Influence of ?S-globin haplotypes and hydroxyurea on tumor necrosis factor-alpha levels in sickle cell anemia

    Directory of Open Access Journals (Sweden)

    Marília Rocha Laurentino

    2014-04-01

    Full Text Available Background: Sickle cell anemia is a chronic inflammatory disease characterized by an increased production of proinflammatory cytokines including tumor necrosis factor-alpha. Hydroxyurea, by decreasing the polymerization of hemoglobin, reduces inflammatory states. The effect of the genetic polymorphisms of sickle cell patients on tumor necrosis factor-alpha levels remains unknown. Objective: The aim of this study was to investigate the association of tumor necrosis factor-alpha levels with β-globin haplotypes and the use of hydroxyurea. Methods: A cross-sectional study was performed of 67 patients with sickle cell anemia diagnosed at steady-state in a referral hospital in Fortaleza, Ceará, Brazil. A group of 26 healthy individuals was used as control. βS-haplotype analysis was performed by restriction fragment length polymorphism-polymerase chain reaction. The tumor necrosis factor-alpha levels were measured by the enzyme-linked immunosorbent assay test. Laboratory data (complete blood count and fetal hemoglobin and information regarding the use of hydroxyurea were obtained from medical records. Statistical analysis was performed using R software with the Kruskal-Wallis and Mann-Whitney tests. Statistical significance was established for p-values < 0.05 for all analyses. Results: The mean age of the participants was 35.48 years. Patients with sickle cell anemia had significantly higher tumor necrosis factor-alpha levels than controls (p-values < 0.0001. Tumor necrosis factor-alpha levels were lower in sickle cell anemia patients who were receiving hydroxyurea treatment than those who were not (p-value = 0.1249. Sickle cell anemia patients with Bantu/n genotype had significantly higher levels than patients with the Bantu/Benin genotype (p-value = 0.0021. Conclusion: In summary, βS-globin haplotypes, but not hydroxyurea therapy, have a role in modulating tumor necrosis factor-alpha levels in sickle cell anemia adults at steady-state. Many

  17. Rat ciliary neurothrophic factor (CNTF): gene structure and regulation of mRNA levels in glial cell cultures.

    OpenAIRE

    Carroll, Patrick; Sendtner, Michael; Meyer, Michael; Thoenen, Hans

    2009-01-01

    The structure of the rat ciliary neurotrophic factor (CNTF) gene and the regulation ofCNTF mRNA levels in cultured glial cells were investigated. The rat mRNA is encoded by a simple two-exon transcription unit. Sequence analysis of the region upstream of the transcription start-site did not reveal a typical TATA-box consensus sequence. Low levels of CNTF mRNA were detected in cultured Schwann cells, and CNTF mRNA was not increased by a variety of treatments. Three-week-old astrocyteenriched c...

  18. Influences of Plant Growth Regulators,Basal Media and Carbohydrate Levels on Cell Suspension Culture of Panax ginseng

    Institute of Scientific and Technical Information of China (English)

    TangWei; WuJiongyuan; 等

    1995-01-01

    A cell suspension culture of Panax ginseng which may be continuously subcultured has been established.Embryogenic callus derived from clutured young leaves was used to initiate the culture,Plant growth regulators,basal medium formula and carbohydrate levels were examined to determine their various effects on suspension culture cell growth and development ,The best selection of plant growth regulator,basal medium and carbohydrate level is 2mg/L 2,4-D:0.5mg/L KT,MS and 3% sucrose respectively.

  19. Prognostic impact of serum albumin levels on the recurrence of stage I non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Ying Jin

    2013-05-01

    Full Text Available Objective: Patients with stage I non-small cell lung cancer who have undergone complete surgical resection harbor a 30% risk for tumor recurrence. Thus, the identification of factors that are predictive for tumor recurrence is urgently needed. The aim of this study was to test the prognostic value of serum albumin levels on tumor recurrence in patients with stage I non-small cell lung cancer. Methods: Stage I non-small cell lung cancer patients who underwent complete surgical resection of the primary tumor at Zhejiang Hospital were analyzed in this study. Serum albumin levels were measured before surgery and once again after surgery in 101 histologically diagnosed non-small cell lung cancer patients. Correlations between the pre- and post-operative serum albumin levels and various clinical demographics and recurrence-free survival rates were analyzed. Results: Patients with pre-operative hypoalbuminemia (<3.5 g/dl had a significantly worse survival rate than patients with normal pre-operative serum albumin levels (≥3.5 g/dl (p=0.008. Patients with post-operative hypoalbuminemia had a worse survival rate when compared with patients with normal post-operative serum albumin levels (p=0.001. Cox multivariate analysis identified pre-operative hypoalbuminemia, post-operative hypoalbuminemia and tumor size over 3 cm as independent negative prognostic factors for recurrence. Conclusion: Serum albumin levels appear to be a significant independent prognostic factor for tumor recurrence in patients with stage I non-small cell lung cancer who have undergone complete resection. Patient pre-treatment and post-treatment serum albumin levels provide an easy and early means of discrimination between patients with a higher risk for recurrence and patients with a low risk of recurrence.

  20. How do heterogeneities in single cell rigidity influence the mechanical behavior at the tissue level?

    Science.gov (United States)

    Bi, Dapeng; Wetzel, Franziska; Fritsch, Anatol; Marchetti, M. Cristina; Manning, M. Lisa; Kaes, Josef

    It has been long recognized that solid tumor tissues are mechanically more rigid than surrounding healthy tissues. However recent experiments have shown that in primary tumor samples from patients with mammary and cervix carcinomas, cells exhibit a broad distribution of rigidities, with a higher fraction of softer and more contractile cells compared to normal tissues. This gives rise to a paradox: does softness emerge from adaptation to mechanical and chemical cues in the external microenvironment, or are soft cells already present inside a primary solid tumor? Motivated by these observations, we study a model of dense tissues that incorporates the experimental data for cell stiffness variations to reveal that, surprisingly, tumors with a significant fraction of very soft cells can still remain rigid. Moreover, in tissues with the observed distributions of cell stiffnesses, softer cells spontaneously self-organize into lines or streams, possibly facilitating cancer metastasis.

  1. Decreased Circulating Interleukin-35 Levels Are Related to Interleukin-4-Producing CD8+ T Cells in Patients with Allergic Asthma

    Directory of Open Access Journals (Sweden)

    Wei Wang

    2015-10-01

    Full Text Available  Interleukin (IL-35 is a newly discovered suppressive cytokine and has been shown to alleviate  inflammatory  and  autoimmune  diseases.  The  purpose  of  this  study  was  to investigate immunomodulatory capacity of IL-35 in patients with allergic asthma.IL-35 mRNA expression levels in peripheral blood mononuclear cells (PBMCs were detected  by  quantitative  real-time  PCR  (qPCR.  The  frequencies  of  cytotoxic  T  cells (Tc1,Tc2  and  Tc17  cells  were  measured  by  flow  cytometry.  Plasma  levels  of  IL-35, interferon (IFN-γ, IL-4, and IL-17 were examined by enzyme-linked immunosorbent assay (ELISA. The correlations between plasma IL-35 levels and Tc1, Tc2, and Tc17 cytokine production in allergic asthmatics (n = 25 and healthy controls (n = 12 were analyzed by Pearson’s test.IL-35 protein and mRNA expression levels were down-regulated in allergic asthmaticscompared with healthy controls. The frequencies of Tc2 and Tc17 cells were significantly increased in patients with asthma, and the frequency of Tc1 cells did not differ between asthmatic patients and healthy controls. Similarly, plasma levels of IL-4 and IL-17 were significantly increased in asthmatic patients, while there was no difference in IFN-γ levels between allergic asthma patients  and  healthy  controls.  More importantly,  plasma  IL-35 protein levels were negatively correlated with the frequency of IL-4-producing CD8+ T (Tc2 cells and with the IL-4 level in patients with allergic asthma.Our results suggest that decreased circulating IL-35 levels could contribute to the pathogenesis of allergic asthma by regulating CD8+ T cells

  2. Biphasic regulation by dibutyryl cyclic AMP of tubulin and actin mRNA levels in neuroblastoma cells.

    OpenAIRE

    Ginzburg, I.; Rybak, S.; Kimhi, Y; Littauer, U. Z.

    1983-01-01

    Blot hybridization analysis that used labeled tubulin cDNA probes revealed that N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate [dibutyryl cyclic AMP (Bt2cAMP)] initially increases and later decreases the level of tubulin mRNA in a neuroblastoma-glioma hybrid cell line as well as in the parent cells. A significant increase in tubulin mRNA sequences is already evident 1 hr after the addition of Bt2cAMP to the neuroblastoma cells, and a maximal induction of 2-fold is seen after 12 hr. Cont...

  3. Promoter methylation and expression levels of selected hematopoietic genes in pediatric B-cell acute lymphoblastic leukemia

    OpenAIRE

    Musialik, Ewa; Bujko, Mateusz; Kober, Paulina; Agnieszka WYPYCH; Gawle-Krawczyk, Karolina; Matysiak, Michal; Siedlecki, Janusz Aleksander

    2015-01-01

    Background Precursor B-cell acute lymphoblastic leukemia (B-cell ALL) is the most common neoplasm in children and is characterized by genetic and epigenetic aberrations in hematopoietic transcription factor (TF) genes. This study evaluated promoter DNA methylation and aberrant expression levels of early- and late-acting hematopoietic TF genes homeobox A4 and A5 (HOXA4 and HOXA5), Meis homeobox 1 (MEIS1), T-cell acute lymphocytic leukemia 1 (TAL1), and interferon regulatory factors 4 and 8 (IR...

  4. Prognostic impact of serum albumin levels on the recurrence of stage I non-small cell lung cancer

    OpenAIRE

    Ying Jin; Li Zhao; Fang Peng

    2013-01-01

    Objective: Patients with stage I non-small cell lung cancer who have undergone complete surgical resection harbor a 30% risk for tumor recurrence. Thus, the identification of factors that are predictive for tumor recurrence is urgently needed. The aim of this study was to test the prognostic value of serum albumin levels on tumor recurrence in patients with stage I non-small cell lung cancer. Methods: Stage I non-small cell lung cancer patients who underwent complete surgical resection of the...

  5. A Transformer-less Partial Power Boost Converter for PV Applications Using a Three-Level Switching Cell

    Energy Technology Data Exchange (ETDEWEB)

    Agamy, Mohammed; Harfman-Todorovic, Maja; Elasser, Ahmed; Essakiappan, Somasundaram

    2013-03-01

    Photovoltaic architectures with distributed power electronics provide many advantages in terms of energy yield as well as system level optimization. As the power level of the solar farm increases it becomes more beneficial to increase the dc collection network voltage, which requires the use of power devices with higher voltage ratings, and thus making the design of efficient, low cost, distributed power converters more challenging. In this paper a simple partial power converter topology is proposed. The topology is implemented using a three-level switching cell, which allows the use of semiconductor devices with lower voltage rating; thus improving design and performance and reducing converter cost. This makes the converters suitable for use for medium to high power applications where dc-link voltages of 600V~1kV may be needed without the need for high voltage devices. Converter operation and experimental results are presented for two partial power circuit variants using three-level switching cells.

  6. Levels of circulating CD45dimCD34+VEGFR2+ progenitor cells correlate with outcome in metastatic renal cell carcinoma patients treated with tyrosine kinase inhibitors

    OpenAIRE

    Farace, F.; Gross-Goupil, M; Tournay, E; Taylor, M; Vimond, N.; Jacques, N; Billiot, F.; Mauguen, A.; Hill, C.; Escudier, B

    2011-01-01

    Background: Predicting the efficacy of antiangiogenic therapy would be of clinical value in patients (pts) with metastatic renal cell carcinoma (mRCC). We tested the hypothesis that circulating endothelial cell (CEC), bone marrow-derived CD45dimCD34+VEGFR2+ progenitor cell or plasma angiogenic factor levels are associated with clinical outcome in mRCC pts undergoing treatment with tyrosine kinase inhibitors (TKI). Methods: Fifty-five mRCC pts were prospectively monitored at baseline (day 1) a...

  7. An internal ribosome entry site (IRES mutant library for tuning expression level of multiple genes in mammalian cells.

    Directory of Open Access Journals (Sweden)

    Esther Y C Koh

    Full Text Available A set of mutated Encephalomyocarditis virus (EMCV internal ribosome entry site (IRES elements with varying strengths is generated by mutating the translation initiation codons of 10(th, 11(th, and 12(th AUG to non-AUG triplets. They are able to control the relative expression of multiple genes over a wide range in mammalian cells in both transient and stable transfections. The relative strength of each IRES mutant remains similar in different mammalian cell lines and is not gene specific. The expressed proteins have correct molecular weights. Optimization of light chain over heavy chain expression by these IRES mutants enhances monoclonal antibody expression level and quality in stable transfections. Uses of this set of IRES mutants can be extended to other applications such as synthetic biology, investigating interactions between proteins and its complexes, cell engineering, multi-subunit protein production, gene therapy, and reprogramming of somatic cells into stem cells.

  8. Response of Listeria monocytogenes to disinfection stress at the single-cell and population levels as monitored by intracellular pH measurements and viable-cell counts

    DEFF Research Database (Denmark)

    Kastbjerg, Vicky Gaedt; Nielsen, Dennis S.; Arneborg, Nils;

    2009-01-01

    bacterium. In situ analyses of Listeria monocytogenes single cells were performed during exposure to different concentrations of the disinfectant Incimaxx DES to study a possible population subdivision. Bacterial survival was quantified with plate counting and disinfection stress at the single-cell level by.......05). The protective effect of NaCl was reflected by viable-cell counts at a higher concentration of Incimaxx (0.0031%), where the salt-grown population survived better than the population grown without NaCl (P <0.05). NaCl protected attached cells through drying but not during disinfection. This study......Listeria monocytogenes has a remarkable ability to survive and persist in food production environments. The purpose of the present study was to determine if cells in a population of L. monocytogenes differ in sensitivity to disinfection agents as this could be a factor explaining persistence of the...

  9. The levels of DNA polymerase alpha and beta during the cell cycle and their role in heat radiosensitization in CHO cells

    International Nuclear Information System (INIS)

    The levels of DNA polymerase alpha and beta were measured during the cell cycle using a whole cell assay technique. The results indicate a decrease in the levels of both enzymes during the G/sub 1/ phase and a gradual increase as cells enter the S phase. The recovery of the DNA polymerases was measured after heating for 10 minutes at 45.50C during G/sub 1/ phase or S phase. The activity of DNA polymerase beta recovers fully during 20-25 hours after heating for both G/sub 1/ phase or S phase cells. There is no recovery of the activity of the DNA polymerase alpha during this time. Survival was also measured when cells were irradiated (4 GY) at various times after hyperthermia (10 min at 45.50C), and for both G/sub 1/ and S phase the interaction between heat and x-ray disappeared fully after 20-25 hours following heating and was parallel to recovery of DNA polymerase beta. Furthermore, treatment with cyclohexamide inhibited protein synthesis and prevented recovery from heat damage assayed in terms of both cell survival and beta polymerase. These results, in addition to experiments with heat sensitization at low pH and heat protection with glycerol, indicate that beta polymerase is probably involved in repairing x-ray induced damage resulting in cell lethality

  10. Elevated Plasma Level of Interferon-λ1 in Chronic Spontaneous Urticaria: Upregulated Expression in CD8+ and Epithelial Cells and Induction of Inflammatory Cell Accumulation

    Science.gov (United States)

    Wang, S. F.; Gao, X. Q.; Xu, Y. N.; Li, D. N.; Wang, H. Y.

    2016-01-01

    Interferon- (IFN-) λ1 is regarded as a potent bio-active molecule in innate immunity. However, little is known about its role in chronic spontaneous urticaria (CSU). We therefore investigated expression of IFN-λ1 in CSU, its cellular location, and its influence on inflammatory cell accumulation by using flow cytometry analysis, skin tissue dispersion, immunohistochemical stain, and a mouse peritoneal inflammation model. The results showed that level of IFN-λ1 was 2.0-fold higher in plasma of the patients with CSU than the level in healthy control (HC) subjects. Among leukocytes examined, only CD8+ T cells expressed more IFN-λ1 in CSU blood. Double labeling immunohistochemical staining revealed that IFN-λ1+ inflammatory cells such as mast cells, eosinophils, B cells, neutrophils, and macrophages were mainly located in dermis, whereas epidermis tissue highly expressed IFN-λ1. IFN-λ1 induced a dose-dependent increase in number of eosinophils, lymphocytes, mast cells, macrophages, and neutrophils in the peritoneum of mice at 6 h following injection, which was inhibited by pretreatment of the animals with anti-intercellular adhesion molecule- (ICAM-) 1 and/or anti-L-selectin antibodies. In conclusion, IFN-λ1 is likely to play a role in the pathogenesis of CSU. Blocking IFN-λ1 production may help to reduce the accumulation of inflammatory cells in the involved CSU skin.

  11. Proteasome inhibitors alter levels of intracellular peptides in HEK293T and SH-SY5Y cells.

    Directory of Open Access Journals (Sweden)

    Sayani Dasgupta

    Full Text Available The proteasome cleaves intracellular proteins into peptides. Earlier studies found that treatment of human embryonic kidney 293T (HEK293T cells with epoxomicin (an irreversible proteasome inhibitor generally caused a decrease in levels of intracellular peptides. However, bortezomib (an antitumor drug and proteasome inhibitor caused an unexpected increase in the levels of most intracellular peptides in HEK293T and SH-SY5Y cells. To address this apparent paradox, quantitative peptidomics was used to study the effect of a variety of other proteasome inhibitors on peptide levels in HEK293T and SH-SY5Y cells. Inhibitors tested included carfilzomib, MG132, MG262, MLN2238, AM114, and clasto-Lactacystin β-lactone. Only MG262 caused a substantial elevation in peptide levels that was comparable to the effect of bortezomib, although carfilzomib and MLN2238 elevated the levels of some peptides. To explore off-target effects, the proteosome inhibitors were tested with various cellular peptidases. Bortezomib did not inhibit tripeptidyl peptidase 2 and only weakly inhibited cellular aminopeptidase activity, as did some of the other proteasome inhibitors. However, potent inhibitors of tripeptidyl peptidase 2 (butabindide and cellular aminopeptidases (bestatin did not substantially alter the peptidome, indicating that the increase in peptide levels due to proteasome inhibitors is not a result of peptidase inhibition. Although we cannot exclude other possibilities, we presume that the paradoxical increase in peptide levels upon treatment with bortezomib and other inhibitors is the result of allosteric effects of these compounds on the proteasome. Because intracellular peptides are likely to be functional, it is possible that some of the physiologic effects of bortezomib and carfilzomib arise from the perturbation of peptide levels inside the cell.

  12. DFM viewpoints of cell-level layout assessments and indications for concurrent layout optimization

    Science.gov (United States)

    Fu, Chung-Min; Yeh, Ping-Heng; Cheng, Yi-Kan; Klaver, Simon

    2008-10-01

    Design-for-manufacturing (DFM) is becoming an actual design practice among IC manufacturers, designers and EDA companies. Layout assessment by design-rule-check (DRC) using EDA tools is a common practice today to ensure well-manufactured design geometries. Standalone DFM tools, which require iteration loops of DFM analysis and fixing, do not fit well in design flows and are considered cumbersome. A better layout assessment method for DFM issues is required: one that gives actionable feedback, and that can be used with automatic optimization in early design stages. The latter is needed to avoid costly design re-spins that will consume critical time-to-market as well as use a lot of engineering resources, reticles and wafer material costs. For example, a DFM checking tool may report the hotspot types and locations, but this information is not sufficient for designers to decide tradeoffs between different fixing choices and to take care of trade-off between physical and electrical design constraints at the same time. When model-based properties are introduced such as lithographic contour, the tradeoffs between rule-based and model-based properties can only be resolved by the automatic and concurrent optimization. This work demonstrates a methodology of DFM scoring of layout based on preferred rules compliance, lithography GATE printability, as well as the layout fixing. The electrical impact on gates is analyzed and showed reduced variability (compared to nominal behavior) in gate performance. Designers can get visual feedback of the layout quality, as well as improvement suggestions. Takumi TKE software is used to demonstrate automatic and concurrent optimization. The method applies to both cell-level and custom designs.

  13. Disruption of canonical TGFβ-signaling in murine coronary progenitor cells by low level arsenic

    International Nuclear Information System (INIS)

    Exposure to arsenic results in several types of cancers as well as heart disease. A major contributor to ischemic heart pathologies is coronary artery disease, however the influences by environmental arsenic in this disease process are not known. Similarly, the impact of toxicants on blood vessel formation and function during development has not been studied. During embryogenesis, the epicardium undergoes proliferation, migration, and differentiation into several cardiac cell types including smooth muscle cells which contribute to the coronary vessels. The TGFβ family of ligands and receptors is essential for developmental cardiac epithelial to mesenchymal transition (EMT) and differentiation into coronary smooth muscle cells. In this in vitro study, 18 hour exposure to 1.34 μM arsenite disrupted developmental EMT programming in murine epicardial cells causing a deficit in cardiac mesenchyme. The expression of EMT genes including TGFβ2, TGFβ receptor-3, Snail, and Has-2 are decreased in a dose-dependent manner following exposure to arsenite. TGFβ2 cell signaling is abrogated as detected by decreases in phosphorylated Smad2/3 when cells are exposed to 1.34 μM arsenite. There is also loss of nuclear accumulation pSmad due to arsenite exposure. These observations coincide with a decrease in vimentin positive mesenchymal cells invading three-dimensional collagen gels. However, arsenite does not block TGFβ2 mediated smooth muscle cell differentiation by epicardial cells. Overall these results show that arsenic exposure blocks developmental EMT gene programming in murine coronary progenitor cells by disrupting TGFβ2 signals and Smad activation, and that smooth muscle cell differentiation is refractory to this arsenic toxicity. - Highlights: • Arsenic blocks TGFβ2 induced expression of EMT genes. • Arsenic blocks TGFβ2 triggered Smad2/3 phosphorylation and nuclear translocation. • Arsenic blocks epicardial cell differentiation into cardiac mesenchyme.

  14. Disruption of canonical TGFβ-signaling in murine coronary progenitor cells by low level arsenic

    Energy Technology Data Exchange (ETDEWEB)

    Allison, Patrick; Huang, Tianfang; Broka, Derrick; Parker, Patti [Department of Pharmacology and Toxicology College of Pharmacy, Southwest Environmental Health Sciences Center, Steele Children' s Research Center and Bio5 Institute, University of Arizona, Tucson, AZ 85721 (United States); Barnett, Joey V. [Department of Pharmacology, Vanderbilt Medical University, Nashville, TN (United States); Camenisch, Todd D., E-mail: camenisch@pharmacy.arizona.edu [Department of Pharmacology and Toxicology College of Pharmacy, Southwest Environmental Health Sciences Center, Steele Children' s Research Center and Bio5 Institute, University of Arizona, Tucson, AZ 85721 (United States)

    2013-10-01

    Exposure to arsenic results in several types of cancers as well as heart disease. A major contributor to ischemic heart pathologies is coronary artery disease, however the influences by environmental arsenic in this disease process are not known. Similarly, the impact of toxicants on blood vessel formation and function during development has not been studied. During embryogenesis, the epicardium undergoes proliferation, migration, and differentiation into several cardiac cell types including smooth muscle cells which contribute to the coronary vessels. The TGFβ family of ligands and receptors is essential for developmental cardiac epithelial to mesenchymal transition (EMT) and differentiation into coronary smooth muscle cells. In this in vitro study, 18 hour exposure to 1.34 μM arsenite disrupted developmental EMT programming in murine epicardial cells causing a deficit in cardiac mesenchyme. The expression of EMT genes including TGFβ2, TGFβ receptor-3, Snail, and Has-2 are decreased in a dose-dependent manner following exposure to arsenite. TGFβ2 cell signaling is abrogated as detected by decreases in phosphorylated Smad2/3 when cells are exposed to 1.34 μM arsenite. There is also loss of nuclear accumulation pSmad due to arsenite exposure. These observations coincide with a decrease in vimentin positive mesenchymal cells invading three-dimensional collagen gels. However, arsenite does not block TGFβ2 mediated smooth muscle cell differentiation by epicardial cells. Overall these results show that arsenic exposure blocks developmental EMT gene programming in murine coronary progenitor cells by disrupting TGFβ2 signals and Smad activation, and that smooth muscle cell differentiation is refractory to this arsenic toxicity. - Highlights: • Arsenic blocks TGFβ2 induced expression of EMT genes. • Arsenic blocks TGFβ2 triggered Smad2/3 phosphorylation and nuclear translocation. • Arsenic blocks epicardial cell differentiation into cardiac mesenchyme.

  15. Checkpoints Studies Using the Budding Yeast Saccharomyces cerevisiae: Analysis of changes in protein level and subcellular localization during cell cycle progression

    OpenAIRE

    Wu, Xiaorong; Liu, Lili; HUANG, Mingxia

    2011-01-01

    Methods are described here to monitor changes in protein level and subcellular localization during the cell cycle progression in the budding yeast S. cerevisiae. Cell synchronization is achieved by an α-factor mediated block-and-release protocol. Cells are collected at different time points for the first two cell cycles upon release. Cellular DNA contents are analyzed by flow cytometry. Trichloroacetic acid protein precipitates are prepared for monitoring levels of cell cycle regulated protei...

  16. PARP-1 inhibition induces a late increase in the level of reactive oxygen species in cells after ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Cieslar-Pobuda, Artur [Biosystems Group, Institute of Automatic Control, Silesian University of Technology, Akademicka 16, 44-100 Gliwice (Poland); Saenko, Yuriy [Biosystems Group, Institute of Automatic Control, Silesian University of Technology, Akademicka 16, 44-100 Gliwice (Poland); Center of Nanotechnology, Ulyanovsk State University, 432700 Ulyanovsk (Russian Federation); Rzeszowska-Wolny, Joanna, E-mail: Joanna.Rzeszowska@polsl.pl [Biosystems Group, Institute of Automatic Control, Silesian University of Technology, Akademicka 16, 44-100 Gliwice (Poland)

    2012-04-01

    Poly(ADP-ribose) polymerase 1 (PARP1), an enzyme activated by DNA strand breaks, synthesizes polymers of poly(ADP-ribose) (PAR) that modify chromatin and other proteins and play a role in DNA repair. Inhibition of PARP1 activity is considered a potentially important strategy in clinical practice, especially to sensitize tumor cells to chemo- and radiotherapy. Here we examined the influence of inhibition of PARP1 on formation of reactive oxygen species (ROS) and on DNA repair in cells exposed to ionizing radiation (IR). K562 (human myelogenous leukaemia) cells were grown and exposed to 4 or 12 Gy of ionizing radiation in presence or absence of the PARP inhibitor NU1025 (100 {mu}M). Intracellular ROS were assayed using the probe 2,7-dichlorofluorescein with detection by flow cytometry and the rejoining of DNA strand breaks were followed by alkaline single cell gel electrophoresis (comet) assays. In untreated cells a significant increase in PAR formation occurred during the first 5 min after IR, followed by a gradual decrease up to 30 min. Addition of a PARP inhibitor arrested the production of PAR almost completely and decreased the rate of rejoining of DNA strand breaks significantly; however, 3 h after irradiation we observed no difference in the amount of DNA strand breaks between PARP inhibitor-treated and untreated cells. Twelve to 48 h after irradiation, an increase of ROS concentration was observed in irradiated cells and ROS levels in PARP inhibitor-treated cells were significantly higher than in cells without inhibitor. Irradiated cells grown in the presence or absence of PARP inhibitor did not differ in the frequencies of apoptotic and necrotic cells or in the activity of caspases at 24, 48 and 72 h after irradiation. Poly(ADP-ribosylation) and inhibition of PARP1 appeared to modulate DNA strand break rejoining and influence the concentration of ROS in irradiated cells.

  17. PARP-1 inhibition induces a late increase in the level of reactive oxygen species in cells after ionizing radiation

    International Nuclear Information System (INIS)

    Poly(ADP-ribose) polymerase 1 (PARP1), an enzyme activated by DNA strand breaks, synthesizes polymers of poly(ADP-ribose) (PAR) that modify chromatin and other proteins and play a role in DNA repair. Inhibition of PARP1 activity is considered a potentially important strategy in clinical practice, especially to sensitize tumor cells to chemo- and radiotherapy. Here we examined the influence of inhibition of PARP1 on formation of reactive oxygen species (ROS) and on DNA repair in cells exposed to ionizing radiation (IR). K562 (human myelogenous leukaemia) cells were grown and exposed to 4 or 12 Gy of ionizing radiation in presence or absence of the PARP inhibitor NU1025 (100 μM). Intracellular ROS were assayed using the probe 2,7-dichlorofluorescein with detection by flow cytometry and the rejoining of DNA strand breaks were followed by alkaline single cell gel electrophoresis (comet) assays. In untreated cells a significant increase in PAR formation occurred during the first 5 min after IR, followed by a gradual decrease up to 30 min. Addition of a PARP inhibitor arrested the production of PAR almost completely and decreased the rate of rejoining of DNA strand breaks significantly; however, 3 h after irradiation we observed no difference in the amount of DNA strand breaks between PARP inhibitor-treated and untreated cells. Twelve to 48 h after irradiation, an increase of ROS concentration was observed in irradiated cells and ROS levels in PARP inhibitor-treated cells were significantly higher than in cells without inhibitor. Irradiated cells grown in the presence or absence of PARP inhibitor did not differ in the frequencies of apoptotic and necrotic cells or in the activity of caspases at 24, 48 and 72 h after irradiation. Poly(ADP-ribosylation) and inhibition of PARP1 appeared to modulate DNA strand break rejoining and influence the concentration of ROS in irradiated cells.

  18. Chronically Elevated Levels of Short-Chain Fatty Acids Induce T Cell-Mediated Ureteritis and Hydronephrosis.

    Science.gov (United States)

    Park, Jeongho; Goergen, Craig J; HogenEsch, Harm; Kim, Chang H

    2016-03-01

    Short-chain fatty acids (SCFAs) are major products of gut microbial fermentation and profoundly affect host health and disease. SCFAs generate IL-10(+) regulatory T cells, which may promote immune tolerance. However, SCFAs can also induce Th1 and Th17 cells upon immunological challenges and, therefore, also have the potential to induce inflammatory responses. Because of the seemingly paradoxical SCFA activities in regulating T cells, we investigated, in depth, the impact of elevated SCFA levels on T cells and tissue inflammation in mice. Orally administered SCFAs induced effector (Th1 and Th17) and regulatory T cells in ureter and kidney tissues, and they induced T cell-mediated ureteritis, leading to kidney hydronephrosis (hereafter called acetate-induced renal disease, or C2RD). Kidney hydronephrosis in C2RD was caused by ureteral obstruction, which was, in turn, induced by SCFA-induced inflammation in the ureteropelvic junction and proximal ureter. Oral administration of all major SCFAs, such as acetate, propionate, and butyrate, induced the disease. We found that C2RD development is dependent on mammalian target of rapamycin activation, T cell-derived inflammatory cytokines such as IFN-γ and IL-17, and gut microbiota. Young or male animals were more susceptible than old or female animals, respectively. However, SCFA receptor (GPR41 or GPR43) deficiency did not affect C2RD development. Thus, SCFAs, when systemically administered at levels higher than physiological levels, cause dysregulated T cell responses and tissue inflammation in the renal system. The results provide insights into the immunological and pathological effects of chronically elevated SCFAs. PMID:26819206

  19. Alveolar Type II cell transplantation restores pulmonary surfactant protein levels in lung fibrosis

    OpenAIRE

    Guillamat-Prats, Raquel; Gay-Jordi, Gemma; Xaubet, Antoni; Peinado, Victor; Serrano-Mollar, Anna

    2014-01-01

    Background Alveolar Type II cell transplantation has been proposed as a cell therapy for the treatment of idiopathic pulmonary fibrosis. Its long-term benefits include repair of lung fibrosis, but its success partly depends on the restoration of lung homeostasis. Our aim was to evaluate surfactant protein restoration after alveolar Type II cell transplantation in an experimental model of bleomycin-induced lung fibrosis in rats. Methods Lung fibrosis was induced by intratracheal instillation o...

  20. Expression Levels of Histone Deacetylases Determine the Cell Fate of Hematopoietic Progenitors*

    OpenAIRE

    Wada, Taeko; Kikuchi, Jiro; Nishimura, Noriko; Shimizu, Rumi; Kitamura, Toshio; Furukawa, Yusuke

    2009-01-01

    Histone deacetylases (HDACs) are globally implicated in the growth and differentiation of mammalian cells; however, relatively little is known about their specific roles in hematopoiesis. In this study, we investigated the expression of HDACs in human hematopoietic cells and their functions during hematopoiesis. The expression of HDACs was very low in hematopoietic progenitor cells, which was accompanied by histone hyperacetylation. HDACs were detectable in more differentiated progenitors and...

  1. Defining cell-type specificity at the transcriptional level in human disease

    OpenAIRE

    Ju, Wenjun; Greene, Casey S; Eichinger, Felix; Nair, Viji; Hodgin, Jeffrey B.; Bitzer, Markus; Lee, Young-Suk; Zhu, Qian; Kehata, Masami; Li, Min; Jiang, Song; Rastaldi, Maria Pia; Cohen, Clemens D; Troyanskaya, Olga G.; Kretzler, Matthias

    2013-01-01

    Cell-lineage–specific transcripts are essential for differentiated tissue function, implicated in hereditary organ failure, and mediate acquired chronic diseases. However, experimental identification of cell-lineage–specific genes in a genome-scale manner is infeasible for most solid human tissues. We developed the first genome-scale method to identify genes with cell-lineage–specific expression, even in lineages not separable by experimental microdissection. Our machine-learning–based approa...

  2. Amorphous silicon solar cells with graded low-level doped i-layerscharacterised by bifacial measurements

    OpenAIRE

    Fischer, Diego; Wyrsch, Nicolas; Fortmann, C.M.; Shah, Arvind

    2008-01-01

    Bifacial spectral response characterization of solar cells under near operating condition illumination is used in conjuncture with a novel bifacial DICE analysis to establish the collection efficiency as a function of i-layer position in p-i-n amorphous silicon solar cells. A significant portion of solar cell degradation can be explained in terms of electric field distortions which increase recombination losses. Unlike carrier lifetime reductions, the field distortions can be reduced. The num...

  3. Modulation of phosphatidylinositol 4-phosphate levels by CaBP7 controls cytokinesis in mammalian cells

    OpenAIRE

    Rajamanoharan, Dayani; Hannah V McCue; Burgoyne, Robert D.; Haynes, Lee P

    2015-01-01

    Calcium and phosphoinositide signaling regulate cell division in model systems, but their significance in mammalian cells is unclear. Calcium-binding protein-7 (CaBP7) is a phosphatidylinositol 4-kinaseIIIβ (PI4KIIIβ) inhibitor required during cytokinesis in mammalian cells, hinting at a link between these pathways. Here we characterize a novel association of CaBP7 with lysosomes that cluster at the intercellular bridge during cytokinesis in HeLa cells. We show that CaBP7 regulates lysosome c...

  4. Low-level laser therapy on MCF-7 cells: a micro-Fourier transform infrared spectroscopy study

    Science.gov (United States)

    Magrini, Taciana D.; dos Santos, Nathalia Villa; Milazzotto, Marcella Pecora; Cerchiaro, Giselle; da Silva Martinho, Herculano

    2012-10-01

    Low-level laser therapy (LLLT) is an emerging therapeutic approach for several clinical conditions. The clinical effects induced by LLLT presumably scale from photobiostimulation/photobioinhibition at the cellular level to the molecular level. The detailed mechanism underlying this effect remains unknown. This study quantifies some relevant aspects of LLLT related to molecular and cellular variations. Malignant breast cells (MCF-7) were exposed to spatially filtered light from a He-Ne laser (633 nm) with fluences of 5, 28.8, and 1000 mJ/cm2. The cell viability was evaluated by optical microscopy using the Trypan Blue viability test. The micro-Fourier transform infrared technique was employed to obtain the vibrational spectra of each experimental group (control and irradiated) and identify the relevant biochemical alterations that occurred due to the process. It was observed that the red light influenced the RNA, phosphate, and serine/threonine/tyrosine bands. We found that light can influence cell metabolism depending on the laser fluence. For 5 mJ/cm2, MCF-7 cells suffer bioinhibition with decreased metabolic rates. In contrast, for the 1 J/cm2 laser fluence, cells present biostimulation accompanied by a metabolic rate elevation. Surprisingly, at the intermediate fluence, 28.8 mJ/cm2, the metabolic rate is increased despite the absence of proliferative results. The data were interpreted within the retrograde signaling pathway mechanism activated with light irradiation.

  5. Keratin 19-positive cutaneous squamous cell carcinoma with elevated serum cytokeratin 19 fragment 21-1 level: A case report

    Science.gov (United States)

    UEDA, MISATO; YASUDA, MICHINORI; SHIBAOKA, YOSHIE; KUROKAWA, ICHIRO; KAKUNO, AYAKO; TSUBURA, AIRO

    2016-01-01

    cytokeratin 19 fragment 21-1 (CYFRA21-1) is a marker of lung cancer useful for evaluating clinical diagnosis and prognosis. To the best of our knowledge, there have been no reports of cutaneous squamous cell carcinoma (SCC) with high levels of CYFRA21-1 to date. We herein report a case of a 79-year-old man with a large subcutaneous tumor of the left shoulder, which was diagnosed as primary cutaneous poorly differentiated SCC. The tumor nests were composed of poorly differentiated atypical squamous cells exhibiting high-grade malignancy and mitotic figures; multinuclear cells were also identified inside lymph vessels. Keratin 19 (K19) was intensely expressed in tumor cells. A significantly elevated level of CYFRA21-1 (33 ng/ml) was observed preoperatively. After surgery, the level of CYFRA21-1 was significantly decreased (from 33 to 5.0 ng/ml). Our case demonstrated that K19-positive primary cutaneous undifferentiated SCC induced high levels of CYFRA21-1 in the serum. Thus, CYFRA 21-1 may be a marker indicative of poorly differentiated cutaneous SCC exhibiting K19 expression.

  6. Submersible microbial desalination cell for simultaneous ammonia recovery and electricity production from anaerobic reactors containing high levels of ammonia

    DEFF Research Database (Denmark)

    Zhang, Yifeng; Angelidaki, Irini

    2015-01-01

    High ammonia concentration in anaerobic reactors can seriously inhibit the anaerobic digestion process. In this study, a submersible microbial desalination cell (SMDC) was developed as an innovative method to lower the ammonia level in a continuous stirred tank reactor (CSTR) by in situ ammonia...

  7. In vivo MRI of cancer cell fate at the single-cell level in a mouse model of breast cancer metastasis to the brain.

    Science.gov (United States)

    Heyn, Chris; Ronald, John A; Ramadan, Soha S; Snir, Jonatan A; Barry, Andrea M; MacKenzie, Lisa T; Mikulis, David J; Palmieri, Diane; Bronder, Julie L; Steeg, Patricia S; Yoneda, Toshiyuki; MacDonald, Ian C; Chambers, Ann F; Rutt, Brian K; Foster, Paula J

    2006-11-01

    Metastasis (the spread of cancer from a primary tumor to secondary organs) is responsible for most cancer deaths. The ability to follow the fate of a population of tumor cells over time in an experimental animal would provide a powerful new way to monitor the metastatic process. Here we describe a magnetic resonance imaging (MRI) technique that permits the tracking of breast cancer cells in a mouse model of brain metastasis at the single-cell level. Cancer cells that were injected into the left ventricle of the mouse heart and then delivered to the brain were detectable on MR images. This allowed the visualization of the initial delivery and distribution of cells, as well as the growth of tumors from a subset of these cells within the whole intact brain volume. The ability to follow the metastatic process from the single-cell stage through metastatic growth, and to quantify and monitor the presence of solitary undivided cells will facilitate progress in understanding the mechanisms of brain metastasis and tumor dormancy, and the development of therapeutics to treat this disease. PMID:17029229

  8. A computational approach to resolve cell level contributions to early glandular epithelial cancer progression

    Directory of Open Access Journals (Sweden)

    Park Sunwoo

    2009-12-01

    Full Text Available Abstract Background Three-dimensional (3D embedded cell cultures provide an appropriate physiological environment to reconstruct features of early glandular epithelial cancer. Although these are orders of magnitude simpler than tissues, they too are complex systems that have proven challenging to understand. We used agent-based, discrete event simulation modeling methods to build working hypotheses of mechanisms of epithelial 3D culture phenotype and early cancer progression. Starting with an earlier software analogue, we validated an improved in silico epithelial analogue (ISEA for cardinal features of a normally developed MDCK cyst. A set of axiomatic operating principles defined simulated cell actions. We explored selective disruption of individual simulated cell actions. New framework features enabled recording detailed measures of ISEA cell activities and morphology. Results Enabled by a small set of cell operating principles, ISEA cells multiplied and self-organized into cyst-like structures that mimicked those of MDCK cells in a 3D embedded cell culture. Selective disruption of "anoikis" or directional cell division caused the ISEA to develop phenotypic features resembling those of in vitro tumor reconstruction models and cancerous tissues in vivo. Disrupting either process, or both, altered cell activity patterns that resulted in morphologically similar outcomes. Increased disruption led to a prolonged presence of intraluminal cells. Conclusions ISEA mechanisms, behaviors, and morphological properties may have biological counterparts. To the extent that in silico-to-in vitro mappings are valid, the results suggest plausible, additional mechanisms of in vitro cancer reconstruction or reversion, and raise potentially significant implications for early cancer diagnosis based on histology. Further ISEA development and use are expected to provide a viable platform to complement in vitro methods for unraveling the mechanistic basis of

  9. Serum Total Tryptase Level Confirms Itself as a More Reliable Marker of Mast Cells Burden in Mast Cell Leukaemia (Aleukaemic Variant

    Directory of Open Access Journals (Sweden)

    P. Savini

    2015-01-01

    Full Text Available Mast cell leukemia (MCL is a very rare form of systemic mastocytosis (SM with a short median survival of 6 months. We describe a case of a 65-year-old woman with aleukaemic variant of MCL with a very high serum total tryptase level of 2255 μg/L at diagnosis, which occurred following an episode of hypotensive shock. She fulfilled the diagnostic criteria of SM, with a bone marrow smear infiltration of 50–60% of atypical mast cells (MCs. She tested negative for the KIT D816V mutation, without any sign of organ damage (no B- or C-findings and only few mediator-related symptoms. She was treated with antihistamine alone and then with imatinib for the appearance of anemia. She maintained stable tryptase level and a very indolent clinical course for twenty-two months; then, she suddenly progressed to acute MCL with a serum tryptase level up to 12960 μg/L. The patient died due to haemorrhagic diathesis twenty-four months after diagnosis. This clinical case maybe represents an example of the chronic form of mast cell leukemia, described as unpredictable disease, in which the serum total tryptase level has confirmed itself as a reliable marker of mast cells burden regardless of the presence of other signs or symptoms.

  10. Study of serum fucose and serum sialic acid levels in oral squamous cell carcinomia.

    OpenAIRE

    Shashikanth M; Rao B

    1994-01-01

    Serum fucose and sialic acid levels were determined in 50 oral cancer patients and 25 healthy controls. A statistically significant increase was noted in the study group. The increase in serum fucose level correlated well with the clinical staging in the study group whereas sialic acid did not. These values were independent of age, sex and histopathological grading. The result suggest that the serum fucose level is a better biochemical tumor marker than sialic acid level. However its usefulne...

  11. Real-time quantification of protein expression at the single-cell level via dynamic protein synthesis translocation reporters

    Science.gov (United States)

    Aymoz, Delphine; Wosika, Victoria; Durandau, Eric; Pelet, Serge

    2016-01-01

    Protein expression is a dynamic process, which can be rapidly induced by extracellular signals. It is widely appreciated that single cells can display large variations in the level of gene induction. However, the variability in the dynamics of this process in individual cells is difficult to quantify using standard fluorescent protein (FP) expression assays, due to the slow maturation of their fluorophore. Here we have developed expression reporters that accurately measure both the levels and dynamics of protein synthesis in live single cells with a temporal resolution under a minute. Our system relies on the quantification of the translocation of a constitutively expressed FP into the nucleus. As a proof of concept, we used these reporters to measure the transient protein synthesis arising from two promoters responding to the yeast hyper osmolarity glycerol mitogen-activated protein kinase pathway (pSTL1 and pGPD1). They display distinct expression dynamics giving rise to strikingly different instantaneous expression noise. PMID:27098003

  12. Real-time quantification of protein expression at the single-cell level via dynamic protein synthesis translocation reporters.

    Science.gov (United States)

    Aymoz, Delphine; Wosika, Victoria; Durandau, Eric; Pelet, Serge

    2016-01-01

    Protein expression is a dynamic process, which can be rapidly induced by extracellular signals. It is widely appreciated that single cells can display large variations in the level of gene induction. However, the variability in the dynamics of this process in individual cells is difficult to quantify using standard fluorescent protein (FP) expression assays, due to the slow maturation of their fluorophore. Here we have developed expression reporters that accurately measure both the levels and dynamics of protein synthesis in live single cells with a temporal resolution under a minute. Our system relies on the quantification of the translocation of a constitutively expressed FP into the nucleus. As a proof of concept, we used these reporters to measure the transient protein synthesis arising from two promoters responding to the yeast hyper osmolarity glycerol mitogen-activated protein kinase pathway (pSTL1 and pGPD1). They display distinct expression dynamics giving rise to strikingly different instantaneous expression noise. PMID:27098003

  13. Squamocin modulates histone H3 phosphorylation levels and induces G1 phase arrest and apoptosis in cancer cells

    International Nuclear Information System (INIS)

    Histone modifications in tumorigenesis are increasingly recognized as important epigenetic factors leading to cancer. Increased phosphorylation levels of histone H3 as a result of aurora B and pMSK1 overexpression were observed in various tumors. We selected aurora B and MSK1 as representatives for testing various compounds and drugs, and found that squamocin, a bis-tetrahydrofuran annonaceous acetogenin, exerted a potent effect on histone H3 phosphorylation. GBM8401, Huh-7, and SW620 cells were incubated with 15, 30, and 60 μM squamocin for 24 h. The expressions of mRNA and proteins were analyzed by qRT-PCR and Western blotting, respectively. The cell viability was determined by an MTT assay. Cell cycle distribution and apoptotic cells were analyzed by flow cytometry. Our results showed that squamocin inhibited the proliferation of GBM8401, Huh-7, and SW620 cells, arrested the cell cycle at the G1 phase, and activated both intrinsic and extrinsic pathways to apoptosis. In addition, we demonstrated that squamocin had the ability to modulate the phosphorylation levels of H3S10 (H3S10p) and H3S28 (H3S28p) in association with the downregulation of aurora B and pMSK1 expressions. This study is the first to show that squamocin affects epigenetic alterations by modulating histone H3 phosphorylation at S10 and S28, providing a novel view of the antitumor mechanism of squamocin

  14. Biotechnology Apprenticeship for Secondary-Level Students: Teaching Advanced Cell Culture Techniques for Research.

    Science.gov (United States)

    Lewis, Jennifer R.; Kotur, Mark S.; Butt, Omar; Kulcarni, Sumant; Riley, Alyssa A.; Ferrell, Nick; Sullivan, Kathryn D.; Ferrari, Mauro

    2002-01-01

    Discusses small-group apprenticeships (SGAs) as a method for introducing cell culture techniques to high school participants. Teaches cell culture practices and introduces advance imaging techniques to solve various biomedical engineering problems. Clarifies and illuminates the value of small-group laboratory apprenticeships. (Author/KHR)

  15. Keeping the intracellular vitamin C at a physiologically relevant level in endothelial cell culture

    DEFF Research Database (Denmark)

    Frikke-Schmidt, Henriette Rønne; Lykkesfeldt, Jens

    2010-01-01

    It is generally accepted that the addition of vitamin C to cell culture medium improves cell growth. However, once added, the vitamin C concentration declines rapidly. This situation differs from the in vivo environment where the endothelium is constantly supplied with ascorbate from the blood...

  16. Hexaazatrinaphthylene Derivatives: Efficient Electron-Transporting Materials with Tunable Energy Levels for Inverted Perovskite Solar Cells.

    Science.gov (United States)

    Zhao, Dongbing; Zhu, Zonglong; Kuo, Ming-Yu; Chueh, Chu-Chen; Jen, Alex K-Y

    2016-07-25

    Hexaazatrinaphthylene (HATNA) derivatives have been successfully shown to function as efficient electron-transporting materials (ETMs) for perovskite solar cells (PVSCs). The cells demonstrate a superior power conversion efficiency (PCE) of 17.6 % with negligible hysteresis. This study provides one of the first nonfullerene small-molecule-based ETMs for high-performance p-i-n PVSCs. PMID:27273656

  17. Increased levels of (class switched) memory B cells in peripheral blood of current smokers

    NARCIS (Netherlands)

    Brandsma, Corry-Anke; Hylkema, Machteld N.; Geerlings, Marie; van Geffen, Wouter; Postma, Dirkje S.; Timens, Wim; Kerstjens, Huib A. M.

    2009-01-01

    There is increasing evidence that a specific immune response contributes to the pathogenesis of COPD. B-cell follicles are present in lung tissue and increased anti-elastin titers have been found in plasma of COPD patients. Additionally, regulatory T cells (Tregs) have been implicated in its pathoge

  18. High-level expression of human calmodulin in E. coli and its effects on cell proliferation

    Institute of Scientific and Technical Information of China (English)

    Xiao Jun Li; Jian Guo Wu; Jun Ling Si; Da Wen Guo; Jian Ping Xu

    2000-01-01

    Calmodulin (CaM), widely distributed in almost all eukaryotic cells, is a major intracellular calcium receptor responsible for mediating the Ca2 + signal to a multitude of different enzyme systems and is thought to play a vital role in the regulation of cell proliferative cycle[1,2]. Recently, many studies showed that CaM is also present in extracellular fluid such as cell culture media and normal body fluid and has been reported to stimulate proliferation in a range of normal and neoplastic cells, apparently acting as an autocrine growth factor[3-11]. In 1988, Crocker et al reported for the first time that addition of extracellular pure pig brain CaM could promote DNA synthesis and cell [7]proliferation in K562 human leukaemic lymphocytes[7].After that, more and more research was done on extracellular CaM and evidences demonstrated that extracellular CaM could also stimulate cell proliferation in normal human umbilical vein endothelial cells[5], keratinocytes[4], suspension-cultured cells of Angelica Dahurica, etc[6]. CaM is a monomeric protein of 148 amino acids that contains four homologous Ca2 + -binding domains. CaM has been highly conserved throughout the evolution. Only 1 out of 148 amino acids of human CaM is different from that of fish CaM. Complementary DNAs encoding rat, eel, chicken, human, and trypanosome CaM have been cloned.

  19. Transgenic Chinese hamster V79 cell lines which exhibit variable levels of gpt mutagenesis

    International Nuclear Information System (INIS)

    The Escherichia coli gpt gene coding for xanthine-guanine phosphoribosyl transferase has been stably transfected into HPRT- Chinese hamster V79 cells. Several gpt- cell lines have been established, which retain the sequence(s) even after long-term culture without selection for gpt. While spontaneous mutagenesis to gpt- occurs rather frequently for most cell lines, it cannot be correlated with either the number of plasmid integration sites or deletion of the plasmid sequence(s). One transgenic cell line (g12), which continuously maintains a low spontaneous mutation frequency was used in comparative mutagenesis studies with wild-type V79 cells (gpt vs. hprt). Alkylating agents such as N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and β-propiolactone (BPL) are shown to be equally toxic and mutagenic in both g12 and V79 cells. UV and X-rays are also equally toxic to both cell lines. The data presented here suggests that g12 cells may be useful to study mammalian mutagenesis by agents which yield limited response at the hprt locus

  20. Metabolic protein interactions in Bacillus subtilis studied at the single cell level

    NARCIS (Netherlands)

    Detert Oude Weme, Ruud Gerardus Johannes

    2015-01-01

    We have investigated protein-protein interactions in live Bacillus subtilis cells (a bacterium). B. subtilis’ natural habitat is the soil and the roots of plants, but also the human microbiota. B. subtilis is used worldwide as a model organism. Unlike eukaryotic cells, bacteria do not have organelle

  1. Determination of optimum sunlight concentration level in space for 3-5 cascade solar cells

    Science.gov (United States)

    Curtis, H. B.

    1982-01-01

    Current-voltage curves were calculated for each cell in a cascade structure using a solar cell diode equation and superposition. Terms for the light generated current, diffusion current, space charge recombination current and series and shunt resistance are included. Individual current voltage curves are added in series with ohmic resistance losses for the cell interconnects to obtain the cascade cell performance. Temperature was varied with concentration, using several models, and ranged from 55 C at one Sun to between 80 and 200 C at 100 Suns. A variety of series resistance and internal resistances were used. Coefficients of the diffusion and recombination terms are strongly temperature dependent. The study indicates that maximum efficiency (30%) occurs in the 50 to 100X Sun concentration range, provided series resistance is below 0.015 ohm-sq cm and cell temperature is 80 C at 100 Suns.

  2. Inhibition of myeloperoxidase-mediated oxidative damage by nitrite in SH-SY5Y cells: Relevance to neuroprotection in neurodegenerative diseases.

    Science.gov (United States)

    Lu, Naihao; Ding, Yun; Tian, Rong; Peng, Yi-Yuan

    2016-06-01