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Cell Suspensions (cell + suspension)
Kinds of Cell Suspensions Terms modified by Cell Suspensions Selected AbstractsOptimization of Rosmarinic Acid Production by Lavandula vera MM Plant Cell Suspension in a Laboratory BioreactorBIOTECHNOLOGY PROGRESS, Issue 2 2005Atanas I. Pavlov The all-round effect of dissolved oxygen concentration, agitation speed, and temperature on the rosmarinic acid production by Lavandula veraMM cell suspension was studied in a 3-L laboratory bioreactor by means of the modified Simplex method. Polynomial regression models were elaborated for description of the process of rosmarinic acid production (Y) in the bioreactor as a consequence of the variation of the dissolved oxygen (X1) concentration between 10% and 50%; agitation (X2) between 100 and 400 rpm; and temperature (X3) between 22 and 30 °C. The optimization made it possible to establish the optimal conditions for the biosynthesis of rosmarinic acid by L. veraMM: dissolved oxygen (X1*), 50% of air saturation; agitation (X2*), 400 rpm; and temperature (X3*), 29.9 °C, where maximal yield (Ymax) of 3489.4 mg/L of rosmarinic acid was achieved (2 times higher compared with the shake-flasks cultivation). [source] Endogenous Fluorescence Spectroscopy of Cell Suspensions for Chemopreventive Drug Monitoring,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 1 2005Nathaniel D. Kirkpatrick ABSTRACT Cancer chemopreventive agents such as N -4-(hydroxyphenyl)-retinamide (4HPR) are thought to prevent cancers by suppressing growth or inducing apoptosis in precancerous cells. Mechanisms by which these drugs affect cells are often not known, and the means to monitor their effects is not available. In this study endogenous fluorescence spectroscopy was used to measure metabolic changes in response to treatment with 4HPR in ovarian and bladder cancer cell lines. Fluorescence signals consistent with nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD) and tryptophan were measured to monitor cellular activity through redox status and protein content. Cells were treated with varying concentrations of 4HPR and measured in a stable environment with a sensitive fluorescence spectrometer. Results suggest that redox signal of all cells changed in a similar dose-dependant manner but started at different baseline levels. Redox signal changes depended primarily on changes consistent with NADH fluorescence, whereas the FAD fluorescence remained relatively constant. Similarly, tryptophan fluorescence decreased with increased drug treatment, suggesting a decrease in protein production. Given that each cell line has been shown to have a different apoptotic response to 4HPR, fluorescence redox values along with changes in tryptophan fluorescence may be a response as well as an endpoint marker for chemopreventive drugs. [source] A Study on Hydrodynamics and Heat Transfer in a Bubble Column Reactor with Yeast and Bacterial Cell SuspensionsTHE CANADIAN JOURNAL OF CHEMICAL ENGINEERING, Issue 4 2005Nigar Kantarci Abstract Hydrodynamics and heat transfer experiments were carried out in a slurry bubble column with air-water-yeast cells and air-water-bacteria cells systems to investigate gas hold-up, bubble characteristics and heat transfer coefficients with cell concentrations of 0.1% w/w and 0.4% w/w and superficial gas velocity up to 0.20 m/s. The gas hold-ups and heat transfer coefficients were found to increase with increasing gas velocity and cell concentration. The heat transfer coefficients were higher at the centre of the column as compared to the near wall region. The development of empirical correlations to predict the heat transfer coefficient in two- and three-phase systems was carried out with ±15% confidence interval at most. On a réalisé des expériences d'hydrodynamique et de transfert de chaleur dans une colonne triphasique gaz-liquide-solide avec des systèmes de cellules air-eau-levure et de cellules air-eau-bactéries afin d'étudier la rétention de gaz, les caractéristiques des bulles et les coefficients de transfert de chaleur avec des concentrations de cellules de 0,1 % en poids et 0,4 % en poids et des vitesses de gaz superficielles jusqu'à 0,20 m/s. On a trouvé que les rétentions de gaz et les coefficients de transfert de chaleur augmentaient avec la vitesse de gaz et la concentration en cellules. Les coefficients de transfert de chaleur sont plus grands au centre de la colonne que dans la région proche de la paroi. Des corrélations empiriques pour prédire le coefficient de transfert de chaleur dans des systèmes bi et triphasiques ont été établies avec un écart de confiance inférieur ou égal à ± 15%. [source] Overexpression of CD7 in classical Hodgkin lymphoma-infiltrating T lymphocytes,CYTOMETRY, Issue 3 2009Adam C. Seegmiller Abstract Background: Diagnosis of Hodgkin lymphoma (HL) is sometimes complicated by the scarcity of neoplastic cells in a reactive inflammatory background. Immunophenotyping by flow cytometry (FC) has not played a significant role in HL diagnosis because of its consistent failure to identify these neoplastic cells. However, HL-infiltrating T cells have been shown to play a role in HL pathogenesis. This study characterizes the FC immunophenotype of these T lymphocytes to determine whether they can be used to assist in the diagnosis of HL. Methods: Cell suspensions from 76 lymph nodes involved by HL and 156 lymph nodes with reactive lymphadenopathy (LAD) were analyzed by flow cytometry to assess the expression of T-cell antigens. Results: The CD4:CD8 ratio and CD7 expression in both CD4(+) and CD8(+) T cells are increased in HL compared with reactive lymph nodes and there are significant differences between these features in different subtypes of HL. However, only the expression of CD7 in CD4(+) T cells distinguishes between HL and reactive LAD. This is especially true for classical HL in younger patients. Using a CD7 mean fluorescence intensity (MFI) cutoff value generated by this data, 37/47 FNA specimens were correctly diagnosed. Conclusions: There are significant differences in the immunophenotypes of HL-infiltrating T cells. Of these, the CD7 expression in CD4(+) T cells discriminates between HL and reactive LAD, suggesting that this could be a useful and practical adjunctive tool in the diagnosis of HL. It may also further our understanding of the pathophysiology of this disease. © 2008 Clinical Cytometry Society [source] Irradiated cultured apoptotic peripheral blood mononuclear cells regenerate infarcted myocardiumEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 6 2009H. J. Ankersmit Abstract Background, Acute myocardial infarction (AMI) is followed by post AMI cardiac remodelling, often leading to congestive heart failure. Homing of c-kit+ endothelial progenitor cells (EPC) has been thought to be the optimal source for regenerating infarcted myocardium. Methods, Immune function of viable peripheral blood mononuclear cells (PBMC) was evaluated after co-culture with irradiated apoptotic PBMC (IA-PBMC) in vitro. Viable PBMC, IA-PBMC and culture supernatants (SN) thereof were obtained after 24 h. Reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay were utilized to quantify interleukin-8 (IL-8), vascular endothelial growth factor, matrix metalloproteinase-9 (MMP9) in PBMC, SN and SN exposed fibroblasts. Cell suspensions of viable- and IA-PBMC were infused in an experimental rat AMI model. Immunohistological analysis was performed to detect inflammatory and pro-angiogenic cells within 72 h post-infarction. Functional data and determination of infarction size were quantified by echocardiography and Elastica van Gieson staining. Results, The IA-PBMC attenuated immune reactivity and resulted in secretion of pro-angiogenic IL-8 and MMP9 in vitro. Fibroblasts exposed to viable and IA-PBMC derived SN caused RNA increment of IL-8 and MMP9. AMI rats that were infused with IA-PBMC cell suspension evidenced enhanced homing of endothelial progenitor cells within 72 h as compared to control (medium alone, viable-PBMC). Echocardiography showed a significant reduction in infarction size and improvement in post AMI remodelling as evidenced by an attenuated loss of ejection fraction. Conclusion, These data indicate that infusion of IA-PBMC cell suspension in experimental AMI circumvented inflammation, caused preferential homing of regenerative EPC and replaced infarcted myocardium. [source] The Effect of Helicobacter pylori Infection on Levels of DNA Damage in Gastric Epithelial CellsHELICOBACTER, Issue 5 2002S. M. Everett Abstract Background.Helicobacter pylori infection leads to an increased risk of developing gastric cancer. The mechanism through which this occurs is not known. We aimed to determine the effect of H. pylori and gastritis on levels of DNA damage in gastric epithelial cells. Methods. Epithelial cells were isolated from antral biopsies from 111 patients. DNA damage was determined using single cell gel electrophoresis and the proportion of cells with damage calculated before and 6 weeks after eradication of H. pylori. Cell suspensions generated by sequential digestions of the same biopsies were assayed to determine the effect of cell position within the gastric pit on DNA damage. Results. DNA damage was significantly higher in normal gastric mucosa than in H. pylori gastritis [median (interquartile range) 65% (58.5,75.8), n = 18 and 21% (11.9,29.8), n = 65, respectively, p < .001]. Intermediate levels were found in reactive gastritis [55.5% (41.3,71.7), n = 13] and H. pylori negative chronic gastritis [50.5% (36.3,60.0), n = 15]. DNA damage rose 6 weeks after successful eradication of H. pylori[to 39.5% (26.3,51.0), p = .007] but was still lower than in normal mucosa. Chronic inflammation was the most important histological factor that determined DNA damage. DNA damage fell with increasing digestion times (r = ,.92 and ,.88 for normal mucosa and H. pylori gastritis, respectively). Conclusions. Lower levels of DNA damage in cells isolated from H. pylori infected gastric biopsies may be a reflection of increased cell turnover in H. pylori gastritis. The investigation of mature gastric epithelial cells for DNA damage is unlikely to elucidate the mechanisms underlying gastric carcinogenesis. [source] In vitro Selection for Fusarium Wilt Resistance in GladiolusJOURNAL OF INTEGRATIVE PLANT BIOLOGY, Issue 5 2008Idrees Ahmad Nasir Abstract Cormels pieces of four Fusarium susceptible Gladiolus cultivars (Friendship, Peter Pears, Victor Borge and Novalux) formed friable calli when cultured in vitro on Murashige and Skoog basal medium containing various concentrations of auxin and cytokinin. The friable calli established cell suspensions. Plantlet regeneration was obtained from the control callus, control cell suspension derived callus and in vitro selected Fusarium oxysporum Schlecht. resistant cell-lines of Friendship. The in vitro cormlets showed 85,95% germination after breaking dormancy of 8 weeks at 4 °C. Cell suspensions of all four Gladiolus cultivars were found to be highly sensitive to fusaric acid. Gradual increase in fusaric acid concentrations to the cell-suspension cultures decreased cell growth considerably. One albino plant was found from the second generation of the in vitro selected cell line of Friendship. The albino plant was found to be highly susceptible to F. oxysporum. The cormlets of all in vitro selected cell lines of Friendship were inoculated with a conidial suspension of the F. oxysporum before planting and were also sprayed with the same spore suspension for further characterization when the height of plants was about 6 cm. The four selected cell lines showed the same response whether or not they were inoculated with conidia of the F. oxysporum. Plantlets of all of the selected cell lines exhibited significant growth as compared with the control after application of conidia of the F. oxysporum. [source] Morphology of Cultured Human Epidermal Melanocytes Observed by Atomic Force MicroscopyPIGMENT CELL & MELANOMA RESEARCH, Issue 1 2004Ru-zhi Zhang The objective of this study was to image the surface structure of cultured human epidermal melanocytes using atomic force microscopy (AFM). Epidermis obtained from human foreskins was treated with 0.5% dispase. Cell suspensions of the epidermis were prepared and seeded in six-well plates, in which sheets of mica had been placed. Samples for AFM were fixed on mica and scanning AFM images were captured by contacting and tapping modes operated under normal atmospheric pressure and temperature. Human epidermal melanocytes exhibited rounded, oval, triangular or quadrangular perikarya from which eight to 10 thick dendrites arose. These dendrites first bifurcated near the soma and then divided profusely into daughter branches, which spread out in all directions. We observed string-like long thin projections, growth cones and shorter thicker projections, which arose from the dendritic shafts, in which groups of melanosomes were arrayed. In addition to such structures, the most striking feature was the presence of filopodia arising from the melanocyte dendrite tips and the melanocyte cell body, many of which contained melanosomes. The termini of dendrites formed unbranched terminal protrusions (approximately 1500,2000 nm wide) consisting of two to three melanosomes wrapped in an arc, with their filopodia extending outwards. The tips of these structures also appeared to be squeezed and finally pinched off by the melanocyte to form a pouch filled with numerous melanosomes. We conclude that secondary and tertiary branches and subordinate branches might take part in transferring melanosomes into keratinocytes in addition to the transfer through the tips of the dendritic shafts. The melanin granules were expelled by exocytosis. [source] Irradiated cultured apoptotic peripheral blood mononuclear cells regenerate infarcted myocardiumEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 6 2009H. J. Ankersmit Abstract Background, Acute myocardial infarction (AMI) is followed by post AMI cardiac remodelling, often leading to congestive heart failure. Homing of c-kit+ endothelial progenitor cells (EPC) has been thought to be the optimal source for regenerating infarcted myocardium. Methods, Immune function of viable peripheral blood mononuclear cells (PBMC) was evaluated after co-culture with irradiated apoptotic PBMC (IA-PBMC) in vitro. Viable PBMC, IA-PBMC and culture supernatants (SN) thereof were obtained after 24 h. Reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay were utilized to quantify interleukin-8 (IL-8), vascular endothelial growth factor, matrix metalloproteinase-9 (MMP9) in PBMC, SN and SN exposed fibroblasts. Cell suspensions of viable- and IA-PBMC were infused in an experimental rat AMI model. Immunohistological analysis was performed to detect inflammatory and pro-angiogenic cells within 72 h post-infarction. Functional data and determination of infarction size were quantified by echocardiography and Elastica van Gieson staining. Results, The IA-PBMC attenuated immune reactivity and resulted in secretion of pro-angiogenic IL-8 and MMP9 in vitro. Fibroblasts exposed to viable and IA-PBMC derived SN caused RNA increment of IL-8 and MMP9. AMI rats that were infused with IA-PBMC cell suspension evidenced enhanced homing of endothelial progenitor cells within 72 h as compared to control (medium alone, viable-PBMC). Echocardiography showed a significant reduction in infarction size and improvement in post AMI remodelling as evidenced by an attenuated loss of ejection fraction. Conclusion, These data indicate that infusion of IA-PBMC cell suspension in experimental AMI circumvented inflammation, caused preferential homing of regenerative EPC and replaced infarcted myocardium. [source] CD5+ B cells with the features of subepithelial B cells found in human tonsilsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 8 2007Mariella Dono Abstract This study describes a CD5+ B cell that differs from the majority of the CD5+ B cells from human tonsils. This cell, isolated from in vivo activated B cells, expressed activation markers and featured a CD23,, IgMhigh, IgDlow surface phenotype, responded to T cell-independent type-2 antigens in vitro, and was detected in the subepithelial (SE) areas, the tonsil equivalent of the splenic marginal zone (MZ). Most of the cells utilized unmutated Ig VH genes, although cells with mutated genes also were found, a finding confirmed by single-cell studies. Mutated sequences were more frequent in suspensions enriched for CD27+ cells. Repeated VDJ gene sequences were observed in different molecular clones from the same cell suspension, suggesting in situ expansion. These CD5+ B cells seem to share features with previously characterized tonsil CD5, SE B cells and differ from the majority of tonsil CD5+ B cells, which have the surface phenotype of follicular mantle B cells, lack activation markers, do not respond to T cell-independent antigens, and utilize unmutated VH genes. These data are discussed considering the present views on the origin of B cell subset populations and the relationships between MZ and B1 cells. [source] CD4 is expressed by epidermal Langerhans' cells predominantly as covalent dimersEXPERIMENTAL DERMATOLOGY, Issue 5 2003G. W. Lynch Abstract:, Langerhans' cells (LC) of skin are CD4 expressing, dendritic, antigen-presenting cells, that are essential for activation of primary immune responses and are productively infected by HIV. We have shown previously that lymphocytes and monocytes express CD4 both as monomers and covalently linked homodimers. In those cells the 55-kDa monomer structure predominates. LC in un-fractionated human epidermal cell (EC) suspension also expresses both forms of CD4, but in EC the dimer form is predominant. Because isolation of LC into single cell suspension by trypsin, as is routinely used for LC isolation, degrades CD4, a systematic study for an alternate procedure for LC isolation was performed. Thus it was found that collagenase blend F treatment can efficiently release LC into suspension, under conditions of only minimal degradation of control soluble recombinant CD4 or CEM-T4 or THP-1 cell CD4, or importantly of LC surface CD4. SDS,PAGE immunoblotting of purified LC extracted from EC by collagenase confirmed CD4 structure as predominantly 110-kDa dimers, with only minimal 55-kDa monomers. The suitability of LC prepared thus for functional studies was demonstrated with binding of functional ligand HIV gp120. It remains to be determined, however, why tissue embedded LC express mainly CD4 dimers, but single-celled blood lymphocytes and monocytes mainly monomers. [source] Cell adhesion regulates platelet-derived growth factor,induced MAP kinase and PI-3 kinase activation in stellate cellsHEPATOLOGY, Issue 3 2002Vinicio Carloni The biologic effects of growth factors are dependent on cell adhesion, and a cross talk occurs between growth factors and adhesion complexes. The aim of the present study was to evaluate the influence of cell adhesion on the major intracellular signaling pathways elicited by platelet-derived growth factor (PDGF) in hepatic stellate cells (HSC). PDGF signaling was investigated in an experimental condition characterized by lack of cell adhesion for different intervals of time. Basal and PDGF-induced focal adhesion kinase (FAK) tyrosine phosphorylation was maintained in a condition of cell suspension for 2, 4, and 6 hours, whereas it was completely lost after 12 and 24 hours. We examined MAP kinase activity at 2 and 24 hours, corresponding to the higher and lower levels of FAK phosphorylation. In these experiments, MAP kinase activity correlated with FAK phosphorylation. Stimulation with PDGF was able to cause Ras-GTP loading only in adherent cells. The ability of PDGF to induce phosphatidylinositol 3-kinase (PI 3-K) activity was abrogated in cells maintained in suspension. The Ser473 phosphorylation of Akt was only marginally affected by the lack of cell adhesion. We then evaluated the association of FAK with c-Src. This association was found to be cell adhesion dependent, and it did not appear to be dependent from phosphorylated FAK. These changes in PDGF-induced intracellular signaling were associated with a remarkable reduction of PDGF-proliferative potential in nonadherent cells, although no marked differences in the apoptotic rate were observed. In conclusion, these results suggest that cell adhesion differentially regulates major signaling pathways activated by PDGF in HSC. [source] Close relation of arterial ICC-like cells to the contractile phenotype of vascular smooth muscle cellJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 4 2007Vladimír Pucovský Abstract This work aimed to establish the lineage of cells similar to the interstitial cells of Cajal (ICC), the arterial ICC-like (AIL) cells, which have recently been described in resistance arteries, and to study their location in the artery wall. Segments of guinea-pig mesenteric arteries and single AIL cells freshly isolated from them were used. Confocal imaging of immunostained cells or segments and electron microscopy of artery segments were used to test for the presence and cellular localization of selected markers, and to localize AIL cells in intact artery segments. AIL cells were negative for PGP9.5, a neural marker, and for von Willebrand factor (vWF), an endothelial cell marker. They were positive for smooth muscle ,-actin and smooth muscle myosin heavy chain (SM-MHC), but expressed only a small amount of smoothelin, a marker of contractile smooth muscle cells (SMC), and of myosin light chain kinase (MLCK), a critical enzyme in the regulation of smooth muscle contraction. Cell isolation in the presence of latrunculin B, an actin polymerization inhibitor, did not cause the disappearance of AIL cells from cell suspension. The fluorescence of basal lamina protein collagen IV was comparable between the AIL cells and the vascular SMCs and the fluorescence of laminin was higher in AIL cells compared to vascular SMCs. Moreover, cells with thin processes were found in the tunica media of small resistance arteries using transmis-sion electron microscopy. The results suggest that AIL cells are immature or phenotypically modulated vascular SMCs constitutively present in resistance arteries. [source] Inactivation of Shigella boydii 18 IDPH and Listeria monocytogenes Scott A with Power Ultrasound at Different Acoustic Energy Densities and TemperaturesJOURNAL OF FOOD SCIENCE, Issue 4 2007Edgar Ugarte-Romero ABSTRACT:, The effect of acoustic energy density (AED) on inactivation of Shigella boydii 18 IDPH and Listeria monocytogenes Scott A in a cell suspension was studied at sublethal temperatures and at AEDs of 0.49, 0.85, and 1.43 W/mL. The effect of temperature on ultrasonic inactivation of L. monocytogenes Scott A at 35, 50, and 65 °C was examined at an AED of 1.43 W/mL. Increasing AED increased the rate of inactivation for both S. boydii and L. monocytogenes. The destruction of S. boydii and L. monocytogenes followed 1st order kinetics in a 20-min treatment, except for S. boydii inactivation at 1.43 W/mL where a tailing effect was observed after 15 min. At sublethal temperatures, the D-values of S. boydii were 8.8, 4.3, and 2.5 min for AEDs of 0.49, 0.85, and 1.43 W/mL, whereas those for L. monocytogenes at the 3 AED levels were 31.5, 13.5, and 7.3 min, respectively. Ultrasonic treatment of L. monocytogenes at 35 and 50 °C enhanced inactivation. However, at 65 °C, application of ultrasound did not result in additional inactivation compared to thermal treatment alone at the same temperature. With the experimental conditions and the ultrasound system used in this study, an upper temperature limit for thermosonication was evident above which no added killing due to ultrasound was observed. [source] In vitro Selection for Fusarium Wilt Resistance in GladiolusJOURNAL OF INTEGRATIVE PLANT BIOLOGY, Issue 5 2008Idrees Ahmad Nasir Abstract Cormels pieces of four Fusarium susceptible Gladiolus cultivars (Friendship, Peter Pears, Victor Borge and Novalux) formed friable calli when cultured in vitro on Murashige and Skoog basal medium containing various concentrations of auxin and cytokinin. The friable calli established cell suspensions. Plantlet regeneration was obtained from the control callus, control cell suspension derived callus and in vitro selected Fusarium oxysporum Schlecht. resistant cell-lines of Friendship. The in vitro cormlets showed 85,95% germination after breaking dormancy of 8 weeks at 4 °C. Cell suspensions of all four Gladiolus cultivars were found to be highly sensitive to fusaric acid. Gradual increase in fusaric acid concentrations to the cell-suspension cultures decreased cell growth considerably. One albino plant was found from the second generation of the in vitro selected cell line of Friendship. The albino plant was found to be highly susceptible to F. oxysporum. The cormlets of all in vitro selected cell lines of Friendship were inoculated with a conidial suspension of the F. oxysporum before planting and were also sprayed with the same spore suspension for further characterization when the height of plants was about 6 cm. The four selected cell lines showed the same response whether or not they were inoculated with conidia of the F. oxysporum. Plantlets of all of the selected cell lines exhibited significant growth as compared with the control after application of conidia of the F. oxysporum. [source] Optimal conditions for in vivo induction of dopaminergic neurons from embryonic stem cells through stromal cell-derived inducing activityJOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2002Asuka Morizane Abstract A method of inducing dopamine (DA) neurons from mouse embryonic stem (ES) cells by stromal cell-derived inducing activity (SDIA) was previously reported. When transplanted, SDIA-induced DA neurons integrate into the mouse striatum and remain positive for tyrosine hydroxylase (TH) expression. In the present study, to optimize the transplantation efficiency, we treated mouse ES cells with SDIA for various numbers of days (8,14 days). SDIA-treated ES cell colonies were isolated by papain treatment and then grafted into the 6-hydroxydopamine (6-OHDA)-lesioned mouse striatum. The ratio of the number of surviving TH-positive cells to the total number of grafted cells was highest when ES cells were treated with SDIA for 12 days before transplantation. This ratio revealed that grafting cell colonies was more efficient for obtaining TH-positive cells in vivo than grafting cell suspensions. When we grafted a cell suspension of 2 × 105, 2 × 104, or 2 × 103 cells into the 6-OHDA-lesioned mouse striatum, we observed only a few surviving TH-positive cells. In conclusion, inducing DA neurons from mouse ES cells by SDIA for 12 days and grafting cell colonies into mouse striatum was the most effective method for the survival of TH-positive neurons in vivo. © 2002 Wiley-Liss, Inc. [source] HEMOLYTIC ACTIVITY OF HETEROCAPSA CIRCULARISQUAMA (DINOPHYCEAE) AND ITS POSSIBLE INVOLVEMENT IN SHELLFISH TOXICITYJOURNAL OF PHYCOLOGY, Issue 4 2001Tatsuya Oda Heterocapsa circularisquama Horiguchi is lethal to shellfish, particularly bivalves such as pearl oysters (Pinctada fucata Gould). No detrimental effects of this flagellate on fish have been observed thus far. In this study, we found that H. circularisquama causes mammalian erythrocytes to lyse. Among the erythrocytes tested, rabbit erythrocytes showed the highest susceptibility, whereas erythrocytes from cattle, sheep, and human were relatively insensitive. Heterocapsa triquetra Stein, which is morphologically similar to H. circularisquama but not toxic to bivalves, showed no hemolytic activity toward rabbit erythrocytes. Culture supernatant or ultrasonic-ruptured cells of H. circularisquama showed only weak hemolytic activity. Hemolytic activity was found in the ethanol extract of H. circularisquama cells, suggesting that the hemolytic agents may be more stable in ethanol than in aqueous solution. Both an intact flagellate cell suspension and the ethanol extract caused morphological changes and eventual collapse of unfertilized eggs of Pacific oyster. Furthermore, the ethanol extract was lethal to the microzooplankton rotifer Brachionus plicatilis Müller, which is highly sensitive to H. circularisquama. Our results suggest that a hemolytic toxin produced by H. circularisquama may be one of the causative agents responsible for the shellfish toxicity. [source] Melatonin modulates the action of near infrared radiation on cell adhesionJOURNAL OF PINEAL RESEARCH, Issue 3 2003Tiina I. Karu Abstract: The adhesion of human cervical cancer (HeLa) cells to a glass matrix is evaluated following their irradiation in a suspension with a pulsed near-infrared (IR) light-emitting diode (wavelength 820 nm, pulse repetition frequency 10 Hz, irradiation dose 16,120 J/m2) when melatonin (4 × 10,11 to 4 × 10,5 m) is added to cell suspension immediately before or after the irradiation. Also, the dependence of visible-to-near-IR radiation (600,840 nm, 52 J/m2) on cell adhesion (action spectrum) is recorded in absence and presence of melatonin (4 × 10,6 m). It is found that melatonin in pharmacological concentrations (but not in physiological range) inhibited cell adherence. Irradiation of cells before or after melatonin treatment normalizes cell adhesion to control level. Melatonin in pharmacological concentrations eliminates stimulation of cell attachment induced by irradiation. Pre-treatment (but not post-treatment) with melatonin in the physiological concentration eliminates cell adhesion stimulation induced by irradiation. Melatonin modifies the light action spectrum significantly in near IR region (760,840 nm only). Thus, the peak at 820,830 nm characteristic for the light action spectrum is fully reduced. [source] Inhibitory effect of propolis extract on the growth of Listeria monocytogenes and the mutagenicity of 4-nitroquinoline- N -oxideJOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 6 2006Hsin-Yi Yang Abstract Propolis originates from a resinous substance collected by honeybees from the buds and leaves of trees and plants, which is then mixed with pollen as well as enzymes secreted by the bees. In the present study, the susceptibility of Listeria monocytogenes to the ethanol extract of propolis (EEP) as influenced by EEP concentration, incubation temperature, pH, and cell age was investigated. In addition, the antimutagenic action of EEP against 4-nitroquinoline- N -oxide (4-NQO) was also examined. Results revealed that EEP at a dosage of 7.5 µg mL,1 or higher exerted a bactericidal effect on L. monocytogenes. L. monocytogenes was most susceptible to EEP at 37 °C followed by 25 and 4 °C. At acid pH values, cells of the test organism were more sensitive to EEP than at neutral pH, while most resistant at alkaline pH values. Cell age was also found to affect the susceptibility of L. monocytogenes to EEP. Cells in the mid-exponential phase showed the highest susceptibility, followed by cells in the late-exponential phase and stationary phase. EEP caused cell leakage of the test organism. A marked increase in the absorbance at 260 nm, UV-absorbing material in the supernatant of cell suspension, and irregularly shaped materials around the cell surface were noted after cells of L. monocytogenes were exposed to EEP. Furthermore, EEP at a dosage of 7.5,60.0 µg per plate was found to suppress 4-NQO-induced mutation by 17.6,88.8%. Copyright © 2006 Society of Chemical Industry [source] PAR and UV Effects on Vertical Migration and Photosynthesis in Euglena gracilis,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2007Peter Richter Recently it was shown that the unicellular flagellate Euglena gracilis changes the sign of gravitaxis from negative to positive upon excessive radiation. This sign change persists in a cell culture for hours even if subsequently transferred to dim light. To test the ecological relevance of this behavior, a vertical column experiment was performed (max. depth 65 cm) to test distribution, photosynthetic efficiency and motility in different horizons of the column (surface, 20, 40 and 65 cm). One column was covered with a UV cut-off filter, which transmits photosynthetically active radiation (PAR) only, the other with a filter which transmits PAR and UV. The columns were irradiated with a solar simulator (PAR 162 W m,2, UV-A 32.6 W m,2, UV-B 1.9 W m,2). The experiment was conducted for 10 days, normally with a light/dim light cycle of 12 h:12 h, but in some cases the light regime was changed (dim light instead of full radiation). Under irradiation the largest fraction of cells was found at the bottom of the column. The cell density decreased toward the surface. Photosynthetic efficiency, determined with a pulse amplitude modulated fluorometer, was negligible at the surface and increased toward the bottom. While the cell suspension showed a positive gravitaxis at the bottom, the cells in the 40 cm horizon were bimodally oriented (about the same percentage of cells swimming upward and downward, respectively). At 20 cm and at the surface the cells showed negative gravitaxis. Positive gravitaxis was more pronounced in the UV + PAR samples. At the surface and in the 20 and 40 cm horizons photosynthetic efficiency was better in the PAR-only samples than in the PAR + UV samples. At the bottom photosynthetic efficiency was similar in both light treatments. The data suggest that high light reverses gravitaxis of the cells, so that they move downward in the water column. At the bottom the light intensity is lower (attenuation of the water column and self shading of the cells) and the cells recover. After recovery the cells swim upward again until the negative gravitaxis is reversed again. [source] Ultraviolet B radiation suppresses Langerhans cell migration in the dermis by down-regulation of ,4 integrinPHOTODERMATOLOGY, PHOTOIMMUNOLOGY & PHOTOMEDICINE, Issue 3 2006Motoko Hamakawa Background/Purpose: Ultraviolet B (UVB) radiation affects the migration and function of epidermal Langerhans cells (LC) and causes immunosuppression of contact hypersensitivity. It is known that LC leaves the epidermis after exposure to UVB. To know the behavior of LC in the dermis after UVB radiation, we studied the effect of UVB radiation on the expression of integrin families on freshly isolated or cultured murine LC. We also examined whether UVB radiation affects the migration of LC to secondary lymphoid tissue chemokine (SLC/6Ckine). Methods: Integrin expressions of murine LC cultured in epidermal cell suspension were analyzed using flowcytometry. We used murine LC sorted flowcytometrically for binding assay to extracellular matrix and for migration assay to chemokine. Skin explant assay and immnohistochemical staining for ,cords formation' were performed as previously described. Results: Twenty and 40 mJ/cm2 of UVB radiation down-regulated the expression of ,4 integrin on 24 h-cultured LC, but not that of ,6, ,1, or ,4 integrin. The number of cultured LC adhered to fibronectin, a ligand for ,4 integrin, was decreased after UVB irradiation, while that to laminin, a ligand for ,6 integrin, was not influenced. UVB radiation reduced the number of migrating LC to SLC. Furthermore, skin sheet explant experiments showed that UVB radiation inhibited the ,cords' formation in dermal vessels of the 48 h-cultured skin. Conclusions: These data suggest that UVB radiation may suppress the migration of LC from the dermis to lymphatic vessels. UVB radiation may downregulate the adherence of LC to dermal fibronectin and migration to SLC, and consequently suppress the migration of LC from the UVB-irradiated dermis to lymphatics. [source] Auxin-induced, SCFTIR1 -mediated poly-ubiquitination marks AUX/IAA proteins for degradationTHE PLANT JOURNAL, Issue 1 2009Felipe Dos Santos Maraschin Summary The plant hormone auxin (indole-3-acetic acid or IAA) regulates plant development by inducing rapid cellular responses and changes in gene expression. Auxin promotes the degradation of Aux/IAA transcriptional repressors, thereby allowing auxin response factors (ARFs) to activate the transcription of auxin-responsive genes. Auxin enhances the binding of Aux/IAA proteins to the receptor TIR1, which is an F-box protein that is part of the E3 ubiquitin ligase complex SCFTIR1. Binding of Aux/IAA proteins leads to degradation via the 26S proteasome, but evidence for SCFTIR1 -mediated poly-ubiquitination of Aux/IAA proteins is lacking. Here we used an Arabidopsis cell suspension-based protoplast system to find evidence for SCFTIR1 -mediated ubiquitination of the Aux/IAA proteins SHY2/IAA3 and BDL/IAA12. Each of these proteins showed a distinct abundance and repressor activity when expressed in this cell system. Moreover, the amount of endogenous TIR1 protein appeared to be rate-limiting for a proper auxin response measured by the co-transfected DR5::GUS reporter construct. Co-transfection with 35S::TIR1 led to auxin-dependent degradation, and excess of 35S::TIR1 even led to degradation of Aux/IAAs in the absence of auxin treatment. Expression of the mutant tir1-1 protein or the related F-box protein COI1, which is involved in jasmonate signaling, had no effect on Aux/IAA degradation. Our results show that SHY2/IAA3 and BDL/IAA12 are poly-ubiquitinated and degraded in response to increased auxin or TIR1 levels. In conclusion, our data provide experimental support for the model that SCFTIR1 -dependent poly-ubiquitination of Aux/IAA proteins marks these proteins for degradation by the 26S proteasome, leading to activation of auxin-responsive gene expression. [source] Identification of a novel family of 70 kDa microtubule-associated proteins in Arabidopsis cellsTHE PLANT JOURNAL, Issue 4 2005Andrey V. Korolev Summary Most plant microtubule-associated proteins (MAPs) have homologues across the phylogenetic spectrum. To find potential plant-specific MAPs that will have evaded bioinformatic searches we devised a low stringency method for isolating proteins from an Arabidopsis cell suspension on endogenous taxol-microtubules. By tryptic peptide mass fingerprinting we identified 55 proteins that were enriched on taxol-microtubules. Amongst a range of known MAPs, such as kinesins, MAP65 isoforms and MOR1, we detected ,unknown' 70 kDa proteins that belong to a family of five closely related Arabidopsis proteins having no known homologues amongst non-plant organisms. To verify that AtMAP70-1 associates with microtubules in vivo, it was expressed as a GFP fusion. This confirmed that the protein decorates all four microtubule arrays in both transiently infected Arabidopsis and stably transformed tobacco BY-2 suspension cells. Microtubule-directed drugs perturbed the localization of AtMAP70-1 but cytochalasin D did not. AtMAP70-1 contains four predicted coiled-coil domains and truncation studies identified a central domain that targets the fusion protein to microtubules in vivo. This study therefore introduces a novel family of plant-specific proteins that interact with microtubules. [source] SAR and efficiency evaluation of a 900 MHz waveguide chamber for cell exposureBIOELECTROMAGNETICS, Issue 6 2008Giuseppe De Prisco Abstract In this work we present the results of numerical and experimental dosimetry carried out for an in vitro exposure device to irradiate sample groups at 900 MHz. The cells are kept in 8 and 15 ml cell cultures, contained, respectively in T25 and T75 rectangular flasks. The dosimetric assessment of the distribution of the specific absorption rate (SAR) is performed for both the bottom of the flask and the whole volume of the sample to provide results for experiments on either the cell layer or the cell suspension. The irradiating chamber is a rectangular waveguide (WG). Different configurations are considered to assess the optimum orientation and positioning of the cell cultures inside the WG. The system performance is optimal when the electric field is parallel to the sample and the WG is terminated by a matched load. In this condition two 15 or four 8 ml cells cultures can be exposed. The efficiency (ratio between the power absorbed by the sample and the incident power) and the non-uniformity degree (ratio between the standard deviation of SAR values and the average SAR over the sample) are calculated and successfully verified through measurements of the scattering parameters and local temperature increases. In the chosen exposure configuration, the efficiency is 0.40 and the non-uniformity degree is 39% for the 15 ml samples. For the 8 ml samples, the efficiency is 0.19 and a low non-uniformity degree (15%) is found. Bioelectromagnetics 29:429,438, 2008. © 2008 Wiley-Liss, Inc. [source] Development and characterization of a tissue engineered pancreatic substitute based on recombinant intestinal endocrine L-cellsBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2009Heather Bara Abstract A tissue engineered pancreatic substitute (TEPS) consisting of insulin-producing cells appropriately designed and encapsulated to support cellular function and prevent interaction with the host may provide physiological blood glucose regulation for the treatment of insulin dependent diabetes (IDD). The performance of agarose-based constructs which contained either a single cell suspension of GLUTag-INS cells, a suspension of pre-aggregated GLUTag-INS spheroids, or GLUTag-INS cells on small intestinal submucosa (SIS), was evaluated in vitro for total cell number, weekly glucose consumption and insulin secretion rates (GCR and ISR), and induced insulin secretion function. The three types of TEPS studied displayed similar number of cells, GCR, and ISR throughout 4 weeks of culture. However, the TEPS, which incorporated SIS as a substrate for the GLUTag-INS cells, was the only type of TEPS tested which was able to retain the induced insulin secretion function of non-encapsulated GLUTag-INS cells. Though improvements in the expression level of GLUTag-INS cells and/or the number of viable cells contained within the TEPS are needed for successful treatment of a murine model of IDD, this study has revealed a potential method for promoting proper cellular function of recombinant L-cells upon incorporation into an implantable three-dimensional TEPS. Biotechnol. Bioeng. 2009;103: 828,834. © 2009 Wiley Periodicals, Inc. [source] A high-rate perfusion bioreactor for plant cellsBIOTECHNOLOGY & BIOENGINEERING, Issue 6 2006C. De Dobbeleer Abstract A perfusion bioreactor allowing continuous extraction of secondary metabolites was designed and challenged for Eschscholtzia californica plant cell suspensions. Four sedimentation columns mounted inside a 2.5-L bioreactor separated single cells and cell aggregates from the culture medium. Cells were elicited with chitin at day 4 and the liquid medium free of cells and debris was then continuously pumped to the extraction columns containing fluidized XAD-7 resins, and then recirculated back to the cell suspension. A medium upward velocity corresponding to cell sedimentation velocity maintained a stable cell/medium separation front in the columns for sedimented cell volume (SCV) of 90% (70% packed cell volume, PCV). Two perfusion bioreactor cultures of 10 and 14 days were performed. A maximum dilution rate of 20.4/day was reached from day 4 to day 6, and was then reduced to 5/day at day 9 for 55% SCV. Control cultures were performed without and with free extraction resins into the cell suspension. Perfusion cultures showed similar specific growth rates of 0.24,±,0.04/day before and after elicitation. However, production level in the perfusion cultures was similar to that from the culture without resins with a maximum of 2.06 µmole/gDW total alkaloids, with 1.54 µmole/gDW in the resins. Cultures with free resins resulted in 30.94 µmole/gDW with 28.4,±,8.8 µmole/gDW in the resins. Difference in the cells nutritional state from elicitation was identified as a major cause in the production reduction. However, pathway to chelilutine was favored in the continuous extraction culture. © 2006 Wiley Periodicals, Inc. [source] Differentiation and lineage selection of mouse embryonic stem cells in a stirred bench scale bioreactor with automated process controlBIOTECHNOLOGY & BIOENGINEERING, Issue 7 2005Magnus Schroeder Abstract It is well established that embryonic stem (ES) cells can differentiate into functional cardiomyocytes in vitro. ES-derived cardiomyocytes could be used for pharmaceutical and therapeutic applications, provided that they can be generated in sufficient quantity and with sufficient purity. To enable large-scale culture of ES-derived cells, we have developed a robust and scalable bioprocess that allows direct embryoid body (EB) formation in a fully controlled, stirred 2 L bioreactor following inoculation with a single cell suspension of mouse ES cells. Utilizing a pitched-blade-turbine, parameters for optimal cell expansion as well as efficient ES cell differentiation were established. Optimization of stirring conditions resulted in the generation of high-density suspension cultures containing 12.5,×,106 cells/mL after 9 days of differentiation. Approximately 30%,40% of the EBs formed in this process vigorously contracted, indicating robust cardiomyogenic induction. An ES cell clone carrying a recombinant DNA molecule comprised of the cardiomyocyte-restricted alpha myosin heavy chain (,MHC) promoter and a neomycin resistance gene was used to establish the utility of this bioprocess to efficiently generate ES-derived cardiomyocytes. The genetically engineered ES cells were cultured directly in the stirred bioreactor for 9 days, followed by antibiotic treatment for another 9 days. The protocol resulted in the generation of essentially pure cardiomyocyte cultures, with a total yield of 1.28,×,109 cells in a single 2 L bioreactor run. This study thus provides an important step towards the large-scale generation of ES-derived cells for therapeutic and industrial applications. © 2005 Wiley Periodicals, Inc. [source] Fractionation of cell mixtures using acoustic and laminar flow fieldsBIOTECHNOLOGY & BIOENGINEERING, Issue 2 2005Manoj Kumar Abstract A fractionation method applicable to different populations of cells in a suspension is reported. The separation was accomplished by subjecting the suspension to a resonant ultrasonic field and a laminar flow field propagating in orthogonal directions within a thin, rectangular chamber. Steady, laminar flow transports the cell suspension along the chamber, while the ultrasonic field causes the suspended cells to migrate to the mid-plane of the chamber at rates related to their size and physical properties. A thin flow splitter positioned near the outlet divides the effluent cell suspension into two product streams, thereby allowing cells that respond faster to the acoustic field to be separated from those cells that respond more slowly. Modeling of the trajectories of individual cells through the chamber shows that by altering the strength of the flow relative to that of the acoustic field, the desired fractionation can be controlled. Proof-of-concept experiments were performed using hybridoma cells and Lactobacillus rhamnosus cells. The two populations of cells could be effectively separated using this technique, resulting in hybridoma/Lactobacillus ratios in the left and right product streams, normalized to the feed ratio, of 6.9 ± 1.8 and 0.39 ± 0.01 (vol/vol), respectively. The acoustic method is fast, efficient, and could be operated continuously with a high degree of selectivity and yield and with low power consumption. © 2004 Wiley Periodicals, Inc. [source] Alkaloid production in Vernonia cinerea: Callus, cell suspension and root culturesBIOTECHNOLOGY JOURNAL, Issue 8 2007Priti Maheshwari Abstract Fast-growing callus, cell suspension and root cultures of Vernonia cinerea, a medicinal plant, were analyzed for the presence of alkaloids. Callus and root cultures were established from young leaf explants in Murashige and Skoog (MS) basal media supplemented with combinations of auxins and cytokinins, whereas cell suspension cultures were established from callus cultures. Maximum biomass of callus, cell suspension and root cultures were obtained in the medium supplemented with 1 mg/L ,-naphthaleneacetic acid (NAA) and 5 mg/L benzylaminopurine (BA), 1.0 mg/L NAA and 0.1 mg/L BA and 1.5 mg/L NAA, respectively. The 5-week-old callus cultures resulted in maximum biomass and alkaloid contents (750 ,g/g). Cell suspension growth and alkaloid contents were maximal in 20-day-old cultures and alkaloid contents were 1.15 mg/g. A 0.2-g sample of root tissue regenerated in semi-solid medium upon transfer to liquid MS medium containing 1.5 mg/L NAA regenerated a maximum increase in biomass of 6.3-fold over a period of 5 weeks. The highest root growth and alkaloid contents of 2 mg/g dry weight were obtained in 5-week-old cultures. Maximum alkaloid contents were obtained in root cultures in vitro compared to all others including the alkaloid content of in vivo obtained with aerial parts and roots (800 ,g/g and 1.2 mg/g dry weight, respectively) of V. cinerea. [source] Optimization of Rosmarinic Acid Production by Lavandula vera MM Plant Cell Suspension in a Laboratory BioreactorBIOTECHNOLOGY PROGRESS, Issue 2 2005Atanas I. Pavlov The all-round effect of dissolved oxygen concentration, agitation speed, and temperature on the rosmarinic acid production by Lavandula veraMM cell suspension was studied in a 3-L laboratory bioreactor by means of the modified Simplex method. Polynomial regression models were elaborated for description of the process of rosmarinic acid production (Y) in the bioreactor as a consequence of the variation of the dissolved oxygen (X1) concentration between 10% and 50%; agitation (X2) between 100 and 400 rpm; and temperature (X3) between 22 and 30 °C. The optimization made it possible to establish the optimal conditions for the biosynthesis of rosmarinic acid by L. veraMM: dissolved oxygen (X1*), 50% of air saturation; agitation (X2*), 400 rpm; and temperature (X3*), 29.9 °C, where maximal yield (Ymax) of 3489.4 mg/L of rosmarinic acid was achieved (2 times higher compared with the shake-flasks cultivation). [source] | |||||||||||||||||||||||||||||||||||||