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21.
Summary As sources of natural products with potential human therapeutic value, marine sponges are important subjects for cell culture
studies. A critical component of any cell culture system is its growth medium. Proceeding from the hypotheses that the thawed,
cryopreserved, primary cells would display detectable differential responses and that those responses could be comparatively
quantified, this study has established that multiwell screening assays are useful tools for improving medium formulations
in cell cultures of the marine sponge, Teichaxinella morchella. Fluorescent probe signals were correlated with known cell densities and viabilities in a 96-well format. Analysis of variance
and post-test methods were applied to judge the significance of signal differences seen in a variety of medium formulations.
Results from a series of experiments suggested that reducing glutamine and selenium concentrations in the standard medium
would result in greater DNA, protein, and esterase activity signals. This was confirmed by the direct comparison of the standard
and improved medium formulations. Significantly higher protein content and esterase activity were associated with the improved
medium. DNA content was also higher, though not significantly. The result is a new medium formulation that may be more able
to support cell growth and division, providing an improved cell culture system for marine sponge cell studies. The assays
can be used in additional studies to further improve the in vitro conditions for marine sponge cell culture. 相似文献
22.
We purified a fraction that showed NAD+-linked methylglyoxal dehydrogenase activity, directly catalyzing methylglyoxal oxidation to pyruvate, which was significantly increased in glutathione-depleted Candida albicans. It also showed NADH-linked methylglyoxal-reducing activity. The fraction was identified as a NAD+-linked alcohol dehydrogenase (ADH1) through mass spectrometric analyses. In ADH1-disruptants of both the wild type and glutathione-depleted cells, the intracellular methylglyoxal concentration increased significantly; defects in growth, differentiation, and virulence were observed; and G2-phase arrest was induced. 相似文献
23.
Hiroshi Shiraishi Hideaki Okamoto Hiromitsu Hara Hiroki Yoshida 《Biochimica et Biophysica Acta (BBA)/General Subjects》2010
Background
Various forms of cell death, such as apoptotic, autophagic and non-lysosomal types, are implicated in normal physiological processes. Apoptotic protease activating factor 1 (Apaf1) is an important component of the intrinsic apoptotic pathway. Deficiency of Apaf1 results in an accumulation of neural progenitor cells (NPCs) in the developing central nervous system and thus, in perinatal lethality. A small percentage of the mutant mice, however, are viable and grow to maturity. The occurrence of such normal mutants implicates alternative cell death pathways during neurogenesis.Methods
NPCs prepared from wild-type or Apaf1-deficient embryos were cultured in growth factor-deprived medium and examined for cell death, caspase activation and morphological alterations. Generation of reactive oxygen species (ROS) and the effects of antioxidants were examined.Results
Wild-type NPCs underwent apoptosis within 24 hours of withdrawal of epidermal growth factor (EGF) or insulin, whereas Apaf1-deficient NPCs underwent cell death but showed no signs of apoptosis. Autophagy was not necessarily accompanied by cell death. Cell death of the Apaf1-deficient NPCs resembled necroptosis—necrosis-like programmed cell death. The necroptosis inhibitor necrostatin-1, however, failed to inhibit the cell death. ROS accumulation was detected in NPCs deprived of growth factors, and an antioxidant partially suppressed the non-apoptotic cell death of Apaf1-deficient NPCs.Conclusions
These data indicate that after withdrawal EGF or insulin withdrawal, the Apaf1-deficient cells underwent non-apoptotic cell death. ROS generation may partially participate in the cell death.General Significance
Non-apoptotic cell death in NPCs may be a compensatory mechanism in the developing CNS of Apaf1-deficient embryos. 相似文献24.
E.S. Abdel-Halim Mostafa M.G. Fouda I. Hamdy F.A. Abdel-Mohdy S.M. El-Sawy 《Carbohydrate polymers》2010,79(1):137
Linear electron beam radiation was used to induce grafting of glycidyl methacrylate/β-cyclodextrin mixture onto cotton fabric. Chlorohexidin diacetate was incorporated to the cavities of cyclodextrin fixed on the cotton fabric to form an inclusion complex having antimicrobial activity. After incorporating chlorohexidin diacetate, the fabric was subjected to several washing cycles to examine the durability of the antimicrobial finishing. Control and grafted cotton fabrics (before and after loading with antimicrobial agent) were characterized for their antimicrobial activity against different kinds of bacteria and fungi.Grafted fabrics loaded with antimicrobial agent were found to show good antimicrobial activity in comparison with control and grafted fabrics which are not loaded with antimicrobial agent. The grafted fabrics loaded with antimicrobial agent were found also to exhibit good antimicrobial activity after five washings and this lasting antimicrobial activity can be attributed to the inclusion complex formed between chlorohexidin diacetate molecules and the cavities of cyclodextrin. 相似文献
25.
Wei Zhang Hiroshi Kudo Shiho Fujisaka Toshiro Sugiyama Naihong Chen Terumi Takahara 《Biochemical and biophysical research communications》2010,391(4):1731-1736
Non-alcoholic steatohepatitis (NASH) develops in a subset of patients with non-alcoholic fatty liver disease (NAFLD), but the exact mechanisms involved in the progression of NAFLD to NASH remain poorly understood. We investigated the role of tumor necrosis factor-α (TNF-α) in the apoptosis of hepatocytes that is related to the severity of NASH. We separated primary hepatocytes from the NAFLD liver caused by a high-fat diet. The production of intracellular reactive oxygen species was increased in steatotic hepatocytes, which were also sensitive to TNF-α. This factor induced significant apoptosis through the signal-regulating kinase 1 (ASK1) and c-Jun N-terminal kinase (JNK) pathway. We describe here a novel culture model of steatotic hepatocytes separated from the NAFLD liver, and demonstrate that TNF-α induces their apoptosis in vitro. 相似文献
26.
A method is described for the isolation of protoplasts from rapidly-growing, friable embryogenic and organogenic cell cultures of corn. A Sepharose 6MB cyanogen-bromide-activated macrobead column coupled with Cellulase RS was used to separate contaminating cells from protoplasts. The column consists of layering 1.5 cm of the coupled-macrobeads into a 2.2-cm diameter column. Contamination of protoplasts by cells possessing partial or complete walls was reduced from 25% to near zero after a single passage through the column. The column was capable of retaining in excess of 30 million cells and recovering 99% cell-free preparations from culture material consisting of less than 1% protoplasts. Coupled-macrobeads were easily recovered, washed free of cells and stored for repeated use. Corn protoplasts appeared undamaged by the column and rapeseed (Brassica napus) protoplasts which were passed through the column have divided and formed colonies in culture. Uncoupled macrobeads were not as efficient as coupled macrobeads in reducing cellular contamination. 相似文献
27.
Devienne KF Cálgaro-Helena AF Dorta DJ Prado IM Raddi MS Vilegas W Uyemura SA Santos AC Curti C 《Phytochemistry》2007,68(7):1075-1080
The isocoumarins (1-50 microM) paepalantine (9,10-dihydroxy-5,7-dimethoxy-1H-naptho(2,3c)pyran-1-one), 8,8'-paepalantine dimer, and vioxanthin isolated from Paepalanthus bromelioides, were assessed for antioxidant activity using isolated rat liver mitochondria and non-mitochondrial systems, and compared with the flavonoid quercetin. The paepalantine and paepalantine dimers, but not vioxanthin, were effective at scavenging both 1,1-diphenyl-2-picrylhydrazyl (DPPH(*)) and superoxide (O(2)(-)) radicals in non-mitochondrial systems, and protected mitochondria from tert-butylhydroperoxide-induced H(2)O(2) accumulation and Fe(2+)-citrate-mediated mitochondrial membrane lipid peroxidation, with almost the same potency as quercetin. These results point towards paepalantine, followed by paepalantine dimer, as being a powerful agent affording protection, apparently via O(2)(-) scavenging, from oxidative stress conditions imposed on mitochondria, the main intracellular source and target of those reactive oxygen species. This strong antioxidant action of paepalantine was reproduced in HepG2 cells exposed to oxidative stress condition induced by H(2)O(2). 相似文献
28.
29.
Does the cellular labile iron pool participate in the oxidation of 2',7'-dichlorodihydrofluorescein?
The fluorogenic probe 2',7'-dichlorodihydrofluorescein diacetate (H2DCF-DA) is widely used for the estimation of oxidative stress in cells. It is known that 2',7'-dichlorodihydrofluorescein (H2DCF), product of intracellular hydrolysis of H2DCF-DA, is oxidized to the fluorescent compound, DCF, mainly by hydrogen peroxide (H2O2) in the presence of catalysts. The present study was aimed at answering the question whether the labile iron pool (LIP) may contribute to the oxidation of H2DCF in cellular systems. The membrane-permeable lipophilic iron chelator salicylaldehyde isonicotinoyl hydrazone (SIH) was found to inhibit oxidation of the probe by H2O2 dependent on ferrous ions but not by peroxidase or superoxide dismutase in defined in vitro systems. When applied to cells, the probe inhibited considerably oxidation of H2DCF in V79 Chinese hamster fibroblasts and two murine lymphoma L5178Y(LY) sublines (LY-R, LY-S) differing in LIP level, the extent of inhibition being greater in the LY-R line of higher LIP level. These results demonstrate that LIP is a significant factor determining the rate of intracellular H2DCF oxidation. 相似文献
30.
Song YH Choi E Park SH Lee SH Cho H Ho WK Ryu SY 《Free radical biology & medicine》2011,51(9):1708-1716
Oxidative stress remodels Ca2+ signaling in cardiomyocytes, which promotes altered heart function in various heart diseases. Ca2+/calmodulin-dependent protein kinase II (CaMKII) was shown to be activated by oxidation, but whether and how CaMKII links oxidative stress to pathophysiological long-term changes in Ca2+ signaling remain unknown. Here, we present evidence demonstrating the role of CaMKII in transient oxidative stress-induced long-term facilitation (LTF) of L-type Ca2+ current (ICa,L) in rat cardiomyocytes. A 5-min exposure of 1 mM H2O2 induced an increase in ICa,L, and this increase was sustained for ~ 1 h. The CaMKII inhibitor KN-93 fully reversed H2O2-induced LTF of ICa,L, indicating that sustained CaMKII activity underlies this oxidative stress-induced memory. Simultaneous inhibition of oxidation and autophosphorylation of CaMKII prevented the maintenance of LTF, suggesting that both mechanisms contribute to sustained CaMKII activity. We further found that sarcoplasmic reticulum Ca2+ release and mitochondrial ROS generation have critical roles in sustaining CaMKII activity via autophosphorylation- and oxidation-dependent mechanisms. Finally, we show that long-term remodeling of the cardiac action potential is induced by H2O2 via CaMKII. In conclusion, CaMKII and mitochondria confer oxidative stress-induced pathological cellular memory that leads to cardiac arrhythmia. 相似文献