Sample records for l929 cell lines
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Toxicity evaluation of ZnO nanostructures on L929 fibroblast cell line using MTS assay
NASA Astrophysics Data System (ADS)
Bakhori, Siti Khadijah Mohd; Mahmud, Shahrom; Ann, Ling Chuo; Mohamed, Azman Seeni; Saifuddin, Siti Nazmin; Masudi, Sam'an Malik; Mohamad, Dasmawati
2015-04-01
ZnO has wide applications in medical and dentistry apart from being used as optoelectronic devices such as solar cells, photodetectors, sensors and light emitting diodes (LEDs). Therefore, the toxicity evaluation is important to know the toxicity level on normal cell line. The toxicity of two grades ZnO nanostructures, ZnO-4 and ZnO-8 have been carried out using cytotoxicity test of MTS assay on L929 rat fibroblast cell line. Prior to that, ZnO-4 and ZnO-8 were characterized for its morphology, structure and optical properties using FESEM, X-ray diffraction, and Photoluminescence respectively. The two groups revealed difference in morphology and exhibit slightly shifted of near band edge emission of Photoluminescence other than having a similar calculated crystallite size of nanostructures. The viability of cells after 72h were obtained and the statistical significance value was calculated using SPSS v20. The p value is more than 0.05 between untreated and treated cell with ZnO. This insignificant value of p>0.05 can be summarized as a non-toxic level of ZnO-4 and ZnO-8 on the L929 cell line.
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Toxicity evaluation of ZnO nanostructures on L929 fibroblast cell line using MTS assay
DOE Office of Scientific and Technical Information (OSTI.GOV)
Bakhori, Siti Khadijah Mohd; Mahmud, Shahrom; Ann, Ling Chuo
2015-04-24
ZnO has wide applications in medical and dentistry apart from being used as optoelectronic devices such as solar cells, photodetectors, sensors and light emitting diodes (LEDs). Therefore, the toxicity evaluation is important to know the toxicity level on normal cell line. The toxicity of two grades ZnO nanostructures, ZnO-4 and ZnO-8 have been carried out using cytotoxicity test of MTS assay on L929 rat fibroblast cell line. Prior to that, ZnO-4 and ZnO-8 were characterized for its morphology, structure and optical properties using FESEM, X-ray diffraction, and Photoluminescence respectively. The two groups revealed difference in morphology and exhibit slightly shiftedmore » of near band edge emission of Photoluminescence other than having a similar calculated crystallite size of nanostructures. The viability of cells after 72h were obtained and the statistical significance value was calculated using SPSS v20. The p value is more than 0.05 between untreated and treated cell with ZnO. This insignificant value of p>0.05 can be summarized as a non-toxic level of ZnO-4 and ZnO-8 on the L929 cell line.« less
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Chen, Fengying; Wu, Tianfu; Cheng, Xiangrong
2014-03-01
To date, there have been very little data on the cytotoxic responses of different cell lines to denture adhesives. To determine the cytotoxicity of three denture adhesives on primary human oral keratinocytes (HOKs), fibroblasts (HOFs) and permanent mouse fibroblasts cell lines (L929). Three commercial denture adhesives (two creams and one powder) were prepared for indirect contact using the agar diffusion test, as well as extracts in MTT assay. The results of the MTT assay were statistically analysed by one-way anova and Tukey's test (p < 0.05). All of the tested denture adhesives showed mild to moderate cytotoxicity to primary HOKs (p < 0.001), whereas none of three was toxic to L929 cells (p > 0.05) in both assays. For primary HOFs cultures, slight cytotoxicity was observed for one of the products from the agar diffusion test and undiluted eluates of all tested adhesives with MTT assay (p < 0.01). Denture adhesives are toxic to the primary HOKs and HOFs cultures, whereas non-toxic to L929 cells. The results suggest that primary human oral mucosal cells may provide more valuable information in toxicity screening of denture adhesives. © 2012 John Wiley & Sons A/S and The Gerodontology Association. Published by John Wiley & Sons Ltd.
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Type I collagen gel protects murine fibrosarcoma L929 cells from TNFα-induced cell death
DOE Office of Scientific and Technical Information (OSTI.GOV)
Wang, Hong-Ju; He, Wen-Qi; Chen, Ling
Murine fibrosarcoma L929 cells have been used to test efficacy of proinflammatory cytokine TNFα. In the present study, we reported on protective effect of type I collagen gel used as L929 cell culture. L929 cell grew and proliferated well on collagen gel. However, the L929 cells exhibited cobblestone-like morphology which was much different from the spread fusiform shape when cultured on conventional cell dishes as well as the cells tended to aggregate. On conventional cell culture dishes, the cells treated with TNFα became round in shape and eventually died in a necroptotic manner. The cells cultured on collagen gel, however,more » were completely unaffected. TNFα treatment was reported to induce autophagy in L929 cells on the plastic dish, and therefore we investigated the effect of collagen gel on induction of autophagy. The results indicated that autophagy induced by TNFα treatment was much reduced when the cells were cultured on collagen gel. In conclusion, type I collagen gel protected L929 cell from TNFα-induced cell death. - Highlights: • Collagen gel culture changed the morphology of L929 cells. • L929 cell cultured on collagen gel were resistant to TNFα-induced cell death. • Collagen gel culture inhibited TNFα-induced autophagy in L929 cells.« less
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Hadjzadeh, Mosa-Al-Reza; Ghanbari, Habib; Keshavarzi, Zakieh; Tavakol-Afshari, Jalil
2014-01-01
Background Although the incidence of gastric cancer is declining during the last half century, this cancer still is the second morbid cancer in the world after lung cancer. The incidence of gastric cancer is 26 per 100,000 in Iran. This study evaluated the effect of Alpinia galangal on AGS cells (human gastric adenocarcinoma epithelial cell line) and L929 cells (as a standard cell line originated from mouse fibroblast cells). Methods After culturing the cells in Roswell Park Memorial Institute (RPMI) medium, the cells were incubated with different doses of Alpinia galangal (0 (control), 125, 250, 500, 750 and 1000 µg/ml) in 24, 48 and 72 hour periods and then, cells viability were assessed using MTT based cell proliferation assay. Results After 24 hours, the percentage of living AGS cells compared to the control group showed no significant decrease at the concentrations of 125 and 250µg/ml. But in the rest concentrations were significant (p<0.05). Only, the percentage of surviving L929 cells at concentration of 125µg/ml of the extract was not significant, but these percentages in the other concentrations were significant. After 48 and 72h incubation, in the last three extract concentrations, the percentage of living AGS and L929 cells significantly decreased compared to control cells (p<0.05). Conclusion We have demonstrated, using cell culture model, anti-proliferative effect of aqueous extract of Alpinia galangal on human gastric tumor (AGS) and L929 cell lines. This effect was prominent in high concentrations. PMID:25250165
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Alabi, Ola D.; Gunnink, Stephen M.; Kuiper, Benjamin D.; Kerk, Samuel A.; Braun, Emily; Louters, Larry L.
2016-01-01
Aims Osthole, a coumarin derivative, has been used in Chinese medicine and studies have suggested a potential use in treatment of diabetes and cancers. Therefore, we investigated the effects of osthole and other coumarins on GLUT1 activity in two cell lines that exclusively express GLUT1. Main Methods We measured the magnitude and time frame of the effects of osthole and related coumarins on glucose uptake in two cells lines; L929 fibroblast cells which have low GLUT1 expression levels and low basal glucose uptake and HCLE cells which have high GLUT1 concentrations and high basal uptake. We also explored the effects of these coumarins in combination with other GLUT1 activators. Key findings Osthole activates glucose uptake in L929 cells with a modest maximum 1.7-fold activation achieved by 50 µM with both activation and recovery occurring within minutes. However, osthole blocks full acute activation of glucose uptake by other, more robust activators. This behavior mimics the effects of other thiol reactive compounds and suggests that osthole is interacting with cysteine residues, possibly within GLUT1 itself. Coumarin, 7-hydroxycoumarin, and 7-methoxycoumarin, do not affect glucose uptake, which is consistent with the notion that the isoprenoid structure in osthole may be important to gain membrane access to GLUT1. In contrast to its effects in L929 cells, osthole inhibits basal glucose uptake in the more active HCLE cells. Significance The differential effects of osthole in L929 and HCLE cells indicated that regulation of GLUT1 varies, likely depending on its membrane concentration. PMID:24657891
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Alabi, Ola D; Gunnink, Stephen M; Kuiper, Benjamin D; Kerk, Samuel A; Braun, Emily; Louters, Larry L
2014-05-02
Osthole, a coumarin derivative, has been used in Chinese medicine and studies have suggested a potential use in treatment of diabetes and cancers. Therefore, we investigated the effects of osthole and other coumarins on GLUT1 activity in two cell lines that exclusively express GLUT1. We measured the magnitude and time frame of the effects of osthole and related coumarins on glucose uptake in two cells lines; L929 fibroblast cells which have low GLUT1 expression levels and low basal glucose uptake and HCLE cells which have high GLUT1 concentrations and high basal uptake. We also explored the effects of these coumarins in combination with other GLUT1 activators. Osthole activates glucose uptake in L929 cells with a modest maximum 1.7-fold activation achieved by 50 μM with both activation and recovery occurring within minutes. However, osthole blocks full acute activation of glucose uptake by other, more robust activators. This behavior mimics the effects of other thiol reactive compounds and suggests that osthole is interacting with cysteine residues, possibly within GLUT1 itself. Coumarin, 7-hydroxycoumarin, and 7-methoxycoumarin, do not affect glucose uptake, which is consistent with the notion that the isoprenoid structure in osthole may be important to gain membrane access to GLUT1. In contrast to its effects in L929 cells, osthole inhibits basal glucose uptake in the more active HCLE cells. The differential effects of osthole in L929 and HCLE cells indicated that regulation of GLUT1 varies, likely depending on its membrane concentration. Copyright © 2014 Elsevier Inc. All rights reserved.
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hua Yu, Jing; yu Liu, Chun; bin Zheng, Gui; Zhang, Li Ying; hui Yan, Ming; yan Zhang, Wen; ying Meng, Xian; fang Yu, Xiao
2013-01-01
Objective: PAB induced various cancer cell apoptosis, cell cycle arrest and senescence. But in cell line murine fibrosarcoma L929, PAB did not induce apoptosis, but autophagy, therefore it was thought by us as a good model to research the relationship of cell cycle arrest, autophagy and senescence bypass apoptosis. Methods: Inhibitory ratio was assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis. Phase contrast microscopy visualized cell morphology. Hoechst 33258 staining for nuclear change, propidium iodode (PI) staining for cell cycle, monodansylcadaverine (MDC) staining for autophagy, and rodanmine 123 staining for mitochondrial membrane potential (MMP) were measured by fluorescence microscopy or flowcytometry. Apoptosis was determined by DNA ladder test. Protein kinase C (PKC) activity was detected by PKC assay kit. SA-β-galactosidase assay was used to detect senescence. Protein expression was examined by western blot. Results: PAB inhibited L929 cell growth in time-and dose-dependent manner. At 12 h, 80 μmol/L PAB induced obvious mitotic arrest; at 24 h, PAB began to induce autophagy; at 36 h, cell-treated with PAB slip into G1 cell cycle; and 3 d PAB induced senescence. In time sequence PAB induced firstly cell cycle arrest, then autophagy, then slippage into G1 phase, lastly senescence. Senescent cells had high level of autophagy, inhibiting autophagy led to apoptosis, and no senescence. PAB activated PKC activity to induce cell cycle arrest, autophagy and senescence, inhibiting PKC activity suppressed cell cycle arrest, autophagy and senescence. Conclusion: PAB induced cell cycle arrest, autophagy and senescence in murine fibrosarcoma L929 cell through PKC. PMID:23630435
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Wang, Hong-Ju; Li, Meng-Qi; Liu, Wei; Yao, Guo-Dong; Xia, Ming-Yu; Hayashi, Toshihiko; Fujisaki, Hitomi; Hattori, Shunji; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi
2016-07-01
Gelatin has been considered to exist as intermediate substance of collagen catabolism in tissue remodeling or under inflammatory conditions. We have initiated the study on possible biological functions of gelatin that can exist temporally and locally under the conditions of remodeling and inflammation Materials and methods: To this purpose, we investigated cell proliferation and survival on gelatin-coated dishes and the response to tumor necrosis factor α (TNFα)-induced cytotoxicity in L929 cells. Autophagy level, ATP level, and ROS generation are examined. L929 cells detached from the gelatin-coated dishes and formed multicellular aggregates. TNFα-induced cytotoxicity in L929 cells was inhibited by gelatin-coating culture. The cells on gelatin-coated dishes showed reduced cellular ATP levels and increased adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) phosphorylation, leading to increased ROS generation and autophagy. This study showed that gelatin-coated culture protected L929 cells from TNFα-induced cytotoxicity and suggested for a possible pathophysiological function of gelatin in regulating cellular functions.
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Collagen gel protects L929 cells from TNFα-induced death by activating NF-κB.
Wang, Hong-Ju; Li, Meng-Qi; Liu, Wei-Wei; Hayashi, Toshihiko; Fujisaki, Hitomi; Hattori, Shunji; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi
2017-09-01
Type I collagen is one of the most abundant components of extracellular matrix. We previously illustrated that murine fibrosarcoma L929 cells grew well on type I collagen gel and escaped from TNFα-induced cell death. In this study, we investigated the mechanism underlying the protective effect of collagen gel. We used western blot, confocal microscopy, MTT assay and flow cytometry by introducing fluorescence staining to determine the expression levels of nuclear factor kappa B (NF-κB), inhibitory ratio and autophagy. L929 cells on collagen gel showed higher expression of NF-κB in the nucleus. Inhibition of NF-κB with pyrrolidine dithiocarbamate hydrochloride (PDTC) or knockdown by NF-κB-siRNA canceled the protective effect of collagen gel on L929 cells from TNFα-induced death, suggesting for the role of NF-κB in the protection from cell death. We found a new aspect of the effect of PDTC on L929 cells cultured on collagen gel. PDTC alone without TNFα induced apoptosis in the L929 cells cultured on collagen gel but not the cells on plastic dish. The apoptosis induction of the L929 cells cultured on collagen gel with PDTC was repressed by inhibiting autophagy with chloroquine, an autophagy inhibitor, suggesting that autophagy contributes to the death induced by the treatment with PDTC. Possible underlying mechanism of this finding is discussed. NF-κB played an important role in protecting the L929 cells cultured on collagen gel from TNFα-induced death.
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Electrochemical properties of 316L stainless steel with culturing L929 fibroblasts
Hiromoto, Sachiko; Hanawa, Takao
2005-01-01
Potentiodynamic polarization and impedance tests were carried out on 316L stainless steel with culturing murine fibroblast L929 cells to elucidate the corrosion behaviour of 316L steel with L929 cells and to understand the electrochemical interface between 316L steel and cells, respectively. Potential step test was carried out on 316L steel with type I collagen coating and culturing L929 cells to compare the effects of collagen and L929 cells. The open-circuit potential of 316L steel slightly shifted in a negative manner and passive current density increased with cells, indicating a decrease in the protective ability of passive oxide film. The pitting potential decreased with cells, indicating a decrease in the pitting corrosion resistance. In addition, a decrease in diffusivity at the interface was indicated from the decrease in the cathodic current density and the increase in the diffusion resistance parameter in the impedance test. The anodic peak current in the potential step test decreased with cells and collagen. Consequently, the corrosion resistance of 316L steel decreases with L929 cells. In addition, collagen coating would provide an environment for anodic reaction similar to that with culturing cells. PMID:16849246
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Fomenko, A. N., E-mail: alserova@ispms.tsc.ru; Korovin, M. S., E-mail: msk@ispms.tsc.ru
The paper presents the toxicity evaluation of nanostructures on the basis of alumina of different shape (nanofibers, nanoplates, nanosheets, nanosheet agglomerates) and with similar physical and chemical properties (particle size, specific surface area, phase composition, and zeta potential). The nanostructures were examined by transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD), low-temperature nitrogen adsorption, and dynamic light scattering (DLS). The cytotoxicity of nanostructures was estimated using L929 fibroblast cells and Neuro-2a tumor cells. It has been found that the L929 cells are less subject to the influence of alumina nanoparticles than the Neuro-2a tumor cells. Probably, themore » differences in the proliferation activity of normal and tumor cells in contact with the synthesized nanostructures are due to a change in the pH of the cell microenvironment.« less
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NASA Astrophysics Data System (ADS)
Fomenko, A. N.; Korovin, M. S.
2016-08-01
The paper presents the toxicity evaluation of nanostructures on the basis of alumina of different shape (nanofibers, nanoplates, nanosheets, nanosheet agglomerates) and with similar physical and chemical properties (particle size, specific surface area, phase composition, and zeta potential). The nanostructures were examined by transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD), low-temperature nitrogen adsorption, and dynamic light scattering (DLS). The cytotoxicity of nanostructures was estimated using L929 fibroblast cells and Neuro-2a tumor cells. It has been found that the L929 cells are less subject to the influence of alumina nanoparticles than the Neuro-2a tumor cells. Probably, the differences in the proliferation activity of normal and tumor cells in contact with the synthesized nanostructures are due to a change in the pH of the cell microenvironment.
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Martyniszyn, Lech; Szulc, Lidia; Boratyńska, Anna; Niemiałtowski, Marek G
2011-12-01
Several reports have brought to light new and interesting findings on the involvement of autophagy and apoptosis in pathogenesis of viral and bacterial diseases, as well as presentation of foreign antigens. Our model studies focused on the involvement of apoptosis during replication of highly virulent Moscow strain of ectromelia virus (ECTV-MOS). Here, we show evidence that autophagy is induced during mousepox replication in a cell line. Fluorescence microscopy revealed increase of LC3 (microtubule-associated protein 1 light chain 3) aggregation in infected as opposed to non-infected control L929 cells. Furthermore, Western blot analysis showed that replication of ECTV-MOS in L929 cells led to the increase in LC3-II (marker of autophagic activity) expression. Beclin 1 strongly colocalized with extranuclear viral replication centers in infected cells, whereas expression of Bcl-2 decreased in those centers as shown by fluorescence microscopy. Loss of Beclin 1-Bcl-2 interaction may lead to autophagy in virus-infected L929 cells. To assess if Beclin 1 has a role in regulation of apoptosis during ECTV-MOS infection, we used small interfering RNA directed against beclin 1 following infection. Early and late apoptotic cells were analyzed by flow cytometry after AnnexinV and propidium iodide staining. Silencing of beclin 1 resulted in decreased percentage of early and late apoptotic cells in the late stage of ECTV-MOS infection in L929 cells. We conclude that Beclin 1 plays an important role in regulation of both, autophagy and apoptosis, during ECTV-MOS replication in L929 permissive cells.
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Toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells.
Li, Juntao; Chen, Sifan; Li, Wenxue; Yang, Guangyu; Zhu, Wei
2015-04-01
To evaluate the toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells. We followed national standards to prepare the extracts from disposable chopsticks, toothpicks, and paper cups used for the cell culture media, and the morphology of L-929 cells was observed with an optical microscope. The loss rate for adherent cells was evaluated with the trypan blue exclusion method, and cell proliferation was determined using the WST-1 assay. Compared with the control group, the cells cultured in media containing the extracts showed signs of apoptosis and necrosis after culturing for 4 or 7 days, and the loss rate for adherent cells was significantly increased (P < 0.05). An obvious decrease in cell viability was also observed (P < 0.05). The extracts from disposable chopsticks, toothpicks, and paper cups can affect the growth and proliferation of L-929 cells and are potentially toxic to humans.
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[Observation of the L929 cell membrane after infrasound exposure with atomic force microscope].
Wang, Bing-shui; Chen, Jing-zao; Liu, Bin; Li, Ling; Yi, Nan; Liu, Jing; Zhang, Sa
2005-12-01
To observe the changes of L929 cell membrane with atomic force microscope (AFM) after infrasound exposure and to explore the mechanisms of effect of infrasound on cell membrane. After primary culture, the L929 cells were exposed to infrasound with intensity output of 130 dB and frequency of 16 Hz 2 hours each day for 3 days. The subsequent changes in the membrane of the control cells and the cells exposed to the infrasound were determined by nano-scale scanning with AFM. After infrasound exposure, the normal prominence of the membrane became short and the dent became shallow in the 7.5 microm x 7.5 microm and 4.0 microm x 4.0 microm photographs. The prominence appeared as cobblestones. The surface of the membrane became smooth. The membrane structure of the L929 cells can be changed by infrasound exposure with intensity of 130 dB and frequency of 16 Hz. The change might be one of the characteristics of effect of infrasound on cell membrane.
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Cell death induced by hydroxyapatite on L929 fibroblast cells.
Inayat-Hussain, S H; Rajab, N F; Roslie, H; Hussin, A A; Ali, A M; Annuar, B O
2004-05-01
Biomaterials intended for end-use application as bone-graft substitutes have to undergo safety evaluation. In this study, we investigated the in vitro cytotoxic effects especially to determine the mode of death of two hydroxyapatite compounds (HA2, HA3) which were synthesized locally. The methods used for cytotoxicity was the standard MTT assay whereas AO/PI staining was performed to determine the mode of cell death in HA treated L929 fibroblasts. Our results demonstrated that both HA2 and HA3 were not significantly cytotoxic as more than 75% cells after 72 hours treatment were viable. Furthermore, we found that the major mode of cell death in HA treated cells was apoptosis. In conclusion, our results demonstrated that these hydroxyapatite compounds are not cytotoxic where the mode of death was primarily via apoptosis.
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Alkaline pH activates the transport activity of GLUT1 in L929 fibroblast cells.
Gunnink, Stephen M; Kerk, Samuel A; Kuiper, Benjamin D; Alabi, Ola D; Kuipers, David P; Praamsma, Riemer C; Wrobel, Kathryn E; Louters, Larry L
2014-04-01
The widely expressed mammalian glucose transporter, GLUT1, can be acutely activated in L929 fibroblast cells by a variety of conditions, including glucose deprivation, or treatment with various respiration inhibitors. Known thiol reactive compounds including phenylarsine oxide and nitroxyl are the fastest acting stimulators of glucose uptake, implicating cysteine biochemistry as critical to the acute activation of GLUT1. In this study, we report that in L929 cells glucose uptake increases 6-fold as the pH of the uptake solution is increased from 6 to 9 with the half-maximal activation at pH 7.5; consistent with the pKa of cysteine residues. This pH effect is essentially blocked by the pretreatment of the cells with either iodoacetamide or cinnamaldehyde, compounds that form covalent adducts with reduced cysteine residues. In addition, the activation by alkaline pH is not additive at pH 8 with known thiol reactive activators such as phenylarsine oxide or hydroxylamine. Kinetic analysis in L929 cells at pH 7 and 8 indicate that alkaline conditions both increases the Vmax and decreases the Km of transport. This is consistent with the observation that pH activation is additive to methylene blue, which activates uptake by increasing the Vmax, as well as to berberine, which activates uptake by decreasing the Km. This suggests that cysteine biochemistry is utilized in both methylene blue and berberine activation of glucose uptake. In contrast a pH increase from 7 to 8 in HCLE cells does not further activate glucose uptake. HCLE cells have a 25-fold higher basal glucose uptake rate than L929 cells and the lack of a pH effect suggests that the cysteine biochemistry has already occurred in HCLE cells. The data are consistent with pH having a complex mechanism of action, but one likely mediated by cysteine biochemistry. Copyright © 2013 Elsevier Masson SAS. All rights reserved.
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Wu, Jin-Nan; Huang, Jian; Yang, Jia; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi
2008-09-01
Oridonin, a diterpenoid isolated from Rabdosia rubescences, has been reported to have antitumor effects. In this study, the growth-inhibitory activity of oridonin for L929 cells was exerted in a time-and dose-dependent manner. After treatment with oridonin for 24 h, L929 cells underwent both apoptosis and necrosis as measured by an lactate dehydrogenase (LDH) activity-based assay. A rapid generation of reactive oxygen species (ROS) was triggered by oridonin, and subsequently up-regulation of phospho-p53 (ser 15) expression and an increased expression ratio of Bax/Bcl-2 was observed. Furthermore, there was a significant fall in mitochondrial membrane potential (MMP) and increase in caspase-3 activity after exposure to oridonin for 24 h. Surprisingly, the pan-caspase inhibitor z-VAD-fmk and caspase3 inhibitor z-DEVD-fmk rendered L929 cells more sensitive to oridonin, rather than preventing oridonin-induced cell death. Oridonin and z-VAD-fmk co-treatment not only resulted in an even higher ROS production, but also made a more significant reduction in the MMP. Pretreatment of ROS scavenger N-acetylcysteine (NAC) led to a complete inhibition of oridonin-induced cell death, intracellular ROS generation, and MMP collapse. NAC treatment also reversed the potentiation of cell death by the pan-caspase inhibitor z-VAD-fmk. Taken together, these observations showed that oridonin-induced cell death in L929 cells involved intracellular ROS generation, activation of phospho-p53 (ser 15), and up-regulation of the Bax/Bcl-2 ratio; and the augmented cell death by z-VAD-fmk was dependent on an increased ROS production.
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Konjhodzic-Prcic, Alma; Jakupovic, Selma; Hasic-Brankovic, Lajla; Vukovic, Amra
2015-01-01
Introduction: The purpose of the current study was to estimate the biocompatibility of endodontic sealers with different bases on L929 mouse fibroblasts permanent cell line using Multiscan EX Spectrophotometer. Materials and Methods: Endodontics sealers used in this study were GuttaFlow (Roeko) silicone based sealer, AH plus (De Tray-DENTSPLY) epoxy resin based, Apexit (Vivadent) calcium hydroxide based and Endorez (Ultradent) methacrylate based sealer. Sealer were tested trough time, freshly mixed 24 h, 48h and 7 days after setting. Biocompatibility was determinate on permanent cell lines L929 mouse fibroblasts trough cytotoxicity using MTT assay. Level of absorption was measured with multi scan EX spectrophotometer on length 420-600 nm. Results: Sealer based on calcium hydroxide Apexit Plus, GuttaFlow silicone based sealer and AH plus epoxy resin based sealer, have shown a low cytotoxicity through the all periods of time on culture of L292 mouse fibroblasts. Methacrylate based sealer, Endorez showed moderate cytotoxicity when freshly mixed and after 7 days. After 24 hours the visibility of the cells was 74,0% and after 48 hours 65,1%. which is slightly cytotoxic. Conclusions: According to results of this study there is a statistically significant difference among the groups p<0,05 for all the tested sealers. Apexit Plus, GuttaFlow and AH plus can be considered as biocompatibile. EndoREZ sealer which is based on methacrylate, after 7 days shows 50,1% of visible live cells which is considered as moderate cytotoxicity. PMID:26236077
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Konjhodzic-Prcic, Alma; Jakupovic, Selma; Hasic-Brankovic, Lajla; Vukovic, Amra
2015-06-01
The purpose of the current study was to estimate the biocompatibility of endodontic sealers with different bases on L929 mouse fibroblasts permanent cell line using Multiscan EX Spectrophotometer. Endodontics sealers used in this study were GuttaFlow (Roeko) silicone based sealer, AH plus (De Tray-DENTSPLY) epoxy resin based, Apexit (Vivadent) calcium hydroxide based and Endorez (Ultradent) methacrylate based sealer. Sealer were tested trough time, freshly mixed 24 h, 48h and 7 days after setting. Biocompatibility was determinate on permanent cell lines L929 mouse fibroblasts trough cytotoxicity using MTT assay. Level of absorption was measured with multi scan EX spectrophotometer on length 420-600 nm. Sealer based on calcium hydroxide Apexit Plus, GuttaFlow silicone based sealer and AH plus epoxy resin based sealer, have shown a low cytotoxicity through the all periods of time on culture of L292 mouse fibroblasts. Methacrylate based sealer, Endorez showed moderate cytotoxicity when freshly mixed and after 7 days. After 24 hours the visibility of the cells was 74,0% and after 48 hours 65,1%. which is slightly cytotoxic. According to results of this study there is a statistically significant difference among the groups p<0,05 for all the tested sealers. Apexit Plus, GuttaFlow and AH plus can be considered as biocompatibile. EndoREZ sealer which is based on methacrylate, after 7 days shows 50,1% of visible live cells which is considered as moderate cytotoxicity.
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Shimizu, Masaya; Muramatsu, Yuki; Tada, Eiko; Kurosawa, Takeshi; Yamaura, Erika; Nakamura, Hiroyuki; Fujino, Hiromichi; Houjyo, Yuuya; Miyasaka, Yuri; Koide, Yuuki; Nishida, Atsushi; Murayama, Toshihiko
2009-03-01
Sphingolipid metabolites including ceramide, sphingosine, and their phosphorylated products [sphingosine-1-phosphate (S1P) and ceramide-1-phosphate] regulate cell functions including arachidonic acid (AA) metabolism and cell death. The development of analogs of S1P may be useful for regulating these mediator-induced cellular responses. We synthesized new analogs of S1P and examined their effects on the release of AA and cell death in L929 mouse fibrosarcoma cells. Among the analogs tested, several compounds including DMB-mC11S [dimethyl (2S,3R)-2-tert-butoxycarbonylamino-3-hydroxy-3-(3'-undecyl)phenylpropyl phosphate] and DMB-mC9S [dimethyl (2S,3R)-2-tert-butoxycarbonylamino-3-hydroxy-3-(3'-nonyl)phenylpropyl phosphate] released AA within 1 h and caused cell death 6 h after treatment. The release of AA was observed in C12 cells [a L929 variant lacking a type alpha cytosolic phospholipase A(2) (cPLA(2)alpha)] and L929-cPLAalpha-siRNA cells (L929 cells treated with small interference RNA for cPLA(2)alpha). Treatment with pharmacological inhibitors of secretory and Ca(2+)-independent PLA(2)s decreased the DMB-mC11S-induced release of AA. The effect of the S1P analogs tested on the release of AA was comparable to that on cell death in L929 cells, and a high correlation coefficient was observed. Two analogs lacking a butoxycarbonyl moiety [DMAc-mC11S (dimethyl (2S,3R)-2-acetamino-3-hydroxy-3-(3'-undecyl)phenylpropyl phosphate] and DMAm-mC11S [dimethyl (2S,3R)-2-amino-3-hydroxy-3-(3'-undecyl)phenylpropyl phosphate)] had inhibitory effects on the release of AA and cell toxicity induced by DMB-mC11S. Synthetic phosphorylated lipid analogs may be useful for studying PLA(2) activity and its toxicity in cells. [Supplementary Fig. 1: available only at http://dx.doi.org/10.1254/jphs.08284FP].
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L929 cell cytotoxicity associated with experimental and commercial dental flosses
NASA Astrophysics Data System (ADS)
Tua-ngam, P.; Supanitayanon, L.; Dechkunakorn, S.; Anuwongnukroh, N.; Srikhirin, T.; Roongrujimek, P.
2017-11-01
This aim of the study was to investigate the cytotoxicity of two commercial and two experimental dental flosses. Two commercial, Oral B® Essential Floss (nylon-waxed) and Thai Silk Floss (silk-waxed), and two experimental, Floss X (nylon-waxed) and Floss Xu (nylon-unwaxed) dental flosses were used. The cytotoxic assay was performed by using cell cultures (L929) which were subjected to cell viability test with methyl-tetrazolium. Each floss specimen (0.4 g) was placed in 1 ml of Minimum Essential Medium at 37°C with 5% CO2 at 100% humidity in an incubator for 24 hours. After incubation, the cell mitochondrial activity was evaluated for detecting viable cells using optical density as per the guidelines of ISO 10993-5:2009(E). Cytotoxic effects were evaluated by measuring percentage of cell viability at 3 points of time- 5 mins, 30 mins, and 1 hr. The results showed that two commercial dental flosses and Floss X had cell viability about 90% at the three time points; however, the experimental Floss Xu presented 80% cell viability at 5 min and <70% cell viability at 30 min and 1 hr. The results concluded that the commercial dental flosses and the experimental dental floss with wax tested in this study were acceptable for clinical use.
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Fan, Weiwei; Lin, Chun Shi; Potluri, Prasanth; Procaccio, Vincent; Wallace, Douglas C.
2012-01-01
The role of mitochondrial DNA (mtDNA) mutations and mtDNA recombination in cancer cell proliferation and developmental biology remains controversial. While analyzing the mtDNAs of several mouse L cell lines, we discovered that every cell line harbored multiple mtDNA mutants. These included four missense mutations, two frameshift mutations, and one tRNA homopolymer expansion. The LA9 cell lines lacked wild-type mtDNAs but harbored a heteroplasmic mixture of mtDNAs, each with a different combination of these variants. We isolated each of the mtDNAs in a separate cybrid cell line. This permitted determination of the linkage phase of each mtDNA and its physiological characteristics. All of the polypeptide mutations inhibited their oxidative phosphorylation (OXPHOS) complexes. However, they also increased mitochondrial reactive oxygen species (ROS) production, and the level of ROS production was proportional to the cellular proliferation rate. By comparing the mtDNA haplotypes of the different cell lines, we were able to reconstruct the mtDNA mutational history of the L–L929 cell line. This revealed that every heteroplasmic L-cell line harbored a mtDNA that had been generated by intracellular mtDNA homologous recombination. Therefore, deleterious mtDNA mutations that increase ROS production can provide a proliferative advantage to cancer or stem cells, and optimal combinations of mutant loci can be generated through recombination. PMID:22345519
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Mier-Giraldo, Helen; Díaz-Barrera, Luis Eduardo; Delgado-Murcia, Lucy Gabriela; Valero-Valdivieso, Manuel Fernando; Cáez-Ramírez, Gabriela
2017-10-01
It was purposed to evaluate the biological potential of ethanol and isopropanol crude extracts of ripe Physalis peruviana fruits. Cytotoxic and immunomodulatory effects of the expression of interleukin-6, interleukin-8, and monocyte chemoattractant protein-1 (MCP-1) were evaluated on human cervical cancer (HeLa) and murine fibroblast (L929) cells. The composition was evaluated by high-performance liquid chromatography diode-array detection and high-performance liquid chromatography ultraviolet/visible detection. The presence of ursolic acid and rosmarinic acid was found in both solvents. However, gallic acid, quercetin, and epicatechin were higher in isopropanol extracts ( P < .05). The results indicated a relationship among the total polyphenol content, antioxidant activity, and cytotoxic activity that was dependent on the solvent used. Isopropanol extracts presented a half-maximal inhibition concentration value (IC 50 ) of 60.48 ± 3.8 μg/mL for HeLa cells and 66.62 ± 2.67 μg/mL for L929 fibroblasts. The extracts reduced the release of interleukin-6, interleukin-8, and MCP-1 in a dose-dependent manner. Extracts showed anticancer and immunomodulatory potential for new complementary pharmaceutical products development.
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Mier-Giraldo, Helen; Díaz-Barrera, Luis Eduardo; Delgado-Murcia, Lucy Gabriela; Valero-Valdivieso, Manuel Fernando; Cáez-Ramírez, Gabriela
2017-01-01
It was purposed to evaluate the biological potential of ethanol and isopropanol crude extracts of ripe Physalis peruviana fruits. Cytotoxic and immunomodulatory effects of the expression of interleukin-6, interleukin-8, and monocyte chemoattractant protein-1 (MCP-1) were evaluated on human cervical cancer (HeLa) and murine fibroblast (L929) cells. The composition was evaluated by high-performance liquid chromatography diode-array detection and high-performance liquid chromatography ultraviolet/visible detection. The presence of ursolic acid and rosmarinic acid was found in both solvents. However, gallic acid, quercetin, and epicatechin were higher in isopropanol extracts (P < .05). The results indicated a relationship among the total polyphenol content, antioxidant activity, and cytotoxic activity that was dependent on the solvent used. Isopropanol extracts presented a half-maximal inhibition concentration value (IC50) of 60.48 ± 3.8 μg/mL for HeLa cells and 66.62 ± 2.67 μg/mL for L929 fibroblasts. The extracts reduced the release of interleukin-6, interleukin-8, and MCP-1 in a dose-dependent manner. Extracts showed anticancer and immunomodulatory potential for new complementary pharmaceutical products development. PMID:28719984
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Faria, Gisele; Cardoso, Cristina R.B.; Department of Biological Sciences, Federal University of Triangulo Mineiro, Uberaba, Minas Gerais
Chlorhexidine (CHX), widely used as antiseptic and therapeutic agent in medicine and dentistry, has a toxic effect both in vivo and in vitro. The intrinsic mechanism underlying CHX-induced cytotoxicity in eukaryotic cells is, however, still unknown. A recent study from our laboratory has suggested that CHX may induce death in cultured L929 fibroblasts via endoplasmic reticulum (ER) stress. This hypothesis was further tested by means of light and electron microscopy, quantification of apoptosis and necrosis by flow cytometry, fluorescence visualization of the cytoskeleton and endoplasmic reticulum, and evaluation of the expression of 78-kDa glucose-regulated protein 78 (Grp78), a marker ofmore » activation of the unfolded protein response (UPR) in cultured L929 fibroblasts. Our finding showing increased Grp 78 expression in CHX-treated cells and the results of flow cytometry, cytoskeleton and endoplasmic reticulum fluorescence visualization, and scanning and transmission electron microscopy allowed us to suggest that CHX elicits accumulation of proteins in the endoplasmic reticulum, which causes ER overload, resulting in ER stress and cell death either by necrosis or apoptosis. It must be pointed out, however, that this does not necessarily mean that ER stress is the only way that CHX kills L929 fibroblasts, but rather that ER stress is an important target or indicator of cell death induced by this drug.« less
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Cao, Ting-Ting; Zhang, Yu-Qing
2015-09-01
Cell cultures often require the addition of animal serum and other supplements. In this study, silk sericin, a bioactive protein, recovered from the waste of silk floss production was hydrolysed into three pepsin-degraded sericin peptides with different ranges of molecular mass. Normal animal cells, tumour cells and hybridoma cells were cultured systematically in FBS culture media containing sericin as a supplement or serum substitute. The culture test and microscopic observation of L929 cells showed that the smaller molecular weight of the degraded sericin is most suitable for cell culture. The cell culture results showed that with the degradation of sericin, for normal mouse fibroblast L929 cells, addition of 0.75 % sericin into FBS culture medium yields cell viability that is superior to FBS culture medium alone. When all serum was replaced by sericin, cell viability in the sericin medium could reach about one half of that in FBS medium. When in a medium containing a mixture of FBS: sericin (6:4, v/v), the cell culture effect is about 80 %. For the cultures of four tumour and one hybridoma cells, regardless of the molecular weight range, these degraded sericin peptides could substitute all serum in FBS media. The cell viability and proliferation of these tumour and hybridoma cells are equivalent or superior to that in FBS medium. In other words, cell viability and proliferation of these tumour and hybridoma cells in sericin media are more preferable to serum media. The mechanism of the sericin protein to promote cell growth and proliferation will be further investigated later.
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Changes in the mitochondrial network during ectromelia virus infection of permissive L929 cells.
Gregorczyk, Karolina P; Szulc-Dąbrowska, Lidia; Wyżewski, Zbigniew; Struzik, Justyna; Niemiałtowski, Marek
2014-01-01
Mitochondria are extremely important organelles in the life of a cell. Recent studies indicate that mitochondria also play a fundamental role in the cellular innate immune mechanisms against viral infections. Moreover, mitochondria are able to alter their shape continuously through fusion and fission. These tightly regulated processes are activated or inhibited under physiological or pathological (e.g. viral infection) conditions to help restore homeostasis. However, many types of viruses, such as orthopoxviruses, have developed various strategies to evade the mitochondrial-mediated antiviral innate immune responses. Moreover, orthopoxviruses exploit the mitochondria for their survival. Such viral activity has been reported during vaccinia virus (VACV) infection. Our study shows that the Moscow strain of ectromelia virus (ECTV-MOS), an orthopoxvirus, alters the mitochondrial network in permissive L929 cells. Upon infection, the branching structure of the mitochondrial network collapses and becomes disorganized followed by destruction of mitochondrial tubules during the late stage of infection. Small, discrete mitochondria co-localize with progeny virions, close to the cell membrane. Furthermore, clustering of mitochondria is observed around viral factories, particularly between the nucleus and viroplasm. Our findings suggest that ECTV-MOS modulates mitochondrial cellular distribution during later stages of the replication cycle, probably enabling viral replication and/or assembly as well as transport of progeny virions inside the cell. However, this requires further investigation.
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Mosafer, Jafar; Teymouri, Manouchehr; Abnous, Khalil; Tafaghodi, Mohsen; Ramezani, Mohammad
2017-03-01
Magnetic nanoparticulate systems based on polymeric materials such as poly (lactic-co-glycolic acid) (PLGA 1 ) are being studied for their potential applications in targeted therapy and imaging of malignant tumors. In the current study, superparamagnetic iron oxide nanocrystals (SPIONs 2 ) and doxorubicin (Dox 3 ) were entrapped in the PLGA-based nanoparticles via a modified multiple emulsion solvent evaporation method. Furthermore, SPIO/Dox-NPs 4 were conjugated to anti-nucleolin AS1411 aptamer (Apt 5 ) and their targeting ability was investigated in high nucleolin-expressing C6 glioma cells compared to low nucleolin-expressing L929 cells. The NPs exhibited a narrow size distribution with mean diameter of ~170nm and an appropriate SPION content (~18% of total polymer weight) with a sufficient saturation magnetization value of 5.9emu/g which is suitable for imaging objectives. They manifested an increased Dox release at pH5.5 compared to pH7.4, with initial burst release (within 24h) followed by sustained release of Dox for 36days. The Apt conjugation to NPs enhanced cellular uptake of Dox in C6 glioma cells compared to L929 cells. Similarly, the Apt-NPs increased the cytotoxicity effect of Dox compared with NPs and Dox solution (f-Dox) alone. In conclusion, the Apt-NPs were found to be a promising delivery system for therapeutic and diagnostic purposes. Copyright © 2016 Elsevier B.V. All rights reserved.
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Rajab, N F; Yaakob, T A; Ong, B Y; Hamid, M; Ali, A M; Annuar, B O; Inayat-Hussain, S H
2004-05-01
Hydroxyapatite is the main component of the bone which is a potential biomaterial substance that can be applied in orthopaedics. In this study, the biocompatibility of this biomaterial was assessed using an in vitro technique. The cytotoxicity and genotoxicity effect of HA2 and HA3 against L929 fibroblast cell was evaluated using the MTT Assay and Alkaline Comet Assay respectively. Both HA2 and HA3 compound showed low cytotoxicity effect as determined using MTT Assay. Cells viability following 72 hours incubation at maximum concentration of both HA2 and HA3 (200 mg/ml) were 75.3 +/- 8.8% and 86.7 +/- 13.1% respectively. However, the cytotoxicity effect of ZnSO4.7H2O as a positive control showed an IC50 values of 46 mg/ml (160 microM). On the other hand, both HA2 and HA3 compound showed a slight genotoxicity effect as determined using the Alkaline Comet Assay following incubation at the concentration 200 mg/ml for 72 hours. This assay has been widely used in genetic toxicology to detect DNA strand breaks and alkali-labile site. The percentage of the cells with DNA damage for both substance was 27.7 +/- 1.3% and 15.6 +/- 1.0% for HA2 and HA3 respectively. Incubation of the cells for 24 hours with 38 microg/ml (IC25) of positive control showed an increase in percentage of cells with DNA damage (67.5 +/- 0.7%). In conclusion, our study indicated that both hydroxyapatite compounds showed a good biocompatibility in fibroblast cells.
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Hashimoto, Masanori; Yamaguchi, Satoshi; Sasaki, Jun-Ichi; Kawai, Koji; Kawakami, Hayato; Iwasaki, Yasuhiko; Imazato, Satoshi
2016-02-01
This study evaluated the inhibition of matrix metalloproteases (MMPs) and cellular responses elicited by gold (Au) and platinum (Pt) nanoparticles (NPs). The interaction of MMP-1 and NPs was evaluated using an MMP assay kit. The cultured L929 cells were exposed to various concentrations of NPs. The cellular responses to NPs were examined using a cytotoxicity assay (that evaluated cell viability and lactic dehydrogenase production), real-time polymerase chain reaction (RT-qPCR), and transmission electron microscopy. Both types of NPs, when used at concentrations above 10 μg ml(-1), inhibited MMP-1 activity. No cytotoxic effects were found when the cells were exposed to AuNPs. In contrast, PtNPs, at both 100 and 400 μg ml(-1), induced cytotoxicity. No inflammatory responses (production of interleukin-6 and tumor necrosis factor-alpha) to NPs were identified by RT-qPCR. The negative surface charge of NPs (COOH(-)) binds to the Zn(2+) of the MMP active center by chelation, leading to MMP inhibition. Gold nanoparticles are plausible candidates for MMP inhibitors in resin-bonding materials because they effectively inhibit MMP-1 activity without cytotoxic or inflammatory effects. © 2015 Eur J Oral Sci.
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Catalán, Elena; Charni, Seyma; Jaime, Paula; Aguiló, Juan Ignacio; Enríquez, José Antonio; Naval, Javier; Pardo, Julián; Villalba, Martín; Anel, Alberto
2015-01-01
Tumor cells have a tendency to use glucose fermentation to obtain energy instead of mitochondrial oxidative phosphorylation (OXPHOS). We demonstrated that this phenotype correlated with loss of ERK5 expression and with reduced MHC class I expression. Consequently, tumor cells could evade cytotoxic T lymphocyte (CTL)-mediated immune surveillance, but also increase their sensitivity to natural killer (NK) cells. These outcomes were evaluated using two cellular models: leukemic EL4 cells and L929 transformed fibroblasts and their derived ρ° cell lines, which lack mitochondrial DNA. We have also used a L929 cell sub-line that spontaneously lost matrix attachment (L929dt), reminiscent of metastasis generation, that also downregulated MHC-I and ERK5 expression. MHC-I expression is lower in ρ° cells than in the parental cell lines, but they were equally sensitive to CTL. On the contrary, ρ° cells were more sensitive to activated NK cells than parental cells. On the other hand, L929dt cells were resistant to CTL and NK cells, showed reduced viability when forced to perform OXPHOS, and surviving cells increased MHC-I expression and became sensitive to CTL. The present results suggest that when the reduction in MHC-I levels in tumor cells due to glycolytic metabolism is partial, the increase in sensitivity to NK cells seems to predominate. However, when tumor cells completely lose MHC-I expression, the combination of treatments that increase OXPHOS with CTL-mediated immunotherapy could be a promising therapeutic approach. PMID:25949869
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Catalán, Elena; Charni, Seyma; Jaime, Paula; Aguiló, Juan Ignacio; Enríquez, José Antonio; Naval, Javier; Pardo, Julián; Villalba, Martín; Anel, Alberto
2015-01-01
Tumor cells have a tendency to use glucose fermentation to obtain energy instead of mitochondrial oxidative phosphorylation (OXPHOS). We demonstrated that this phenotype correlated with loss of ERK5 expression and with reduced MHC class I expression. Consequently, tumor cells could evade cytotoxic T lymphocyte (CTL)-mediated immune surveillance, but also increase their sensitivity to natural killer (NK) cells. These outcomes were evaluated using two cellular models: leukemic EL4 cells and L929 transformed fibroblasts and their derived ρ° cell lines, which lack mitochondrial DNA. We have also used a L929 cell sub-line that spontaneously lost matrix attachment (L929dt), reminiscent of metastasis generation, that also downregulated MHC-I and ERK5 expression. MHC-I expression is lower in ρ° cells than in the parental cell lines, but they were equally sensitive to CTL. On the contrary, ρ° cells were more sensitive to activated NK cells than parental cells. On the other hand, L929dt cells were resistant to CTL and NK cells, showed reduced viability when forced to perform OXPHOS, and surviving cells increased MHC-I expression and became sensitive to CTL. The present results suggest that when the reduction in MHC-I levels in tumor cells due to glycolytic metabolism is partial, the increase in sensitivity to NK cells seems to predominate. However, when tumor cells completely lose MHC-I expression, the combination of treatments that increase OXPHOS with CTL-mediated immunotherapy could be a promising therapeutic approach.
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Tokunaga, Etsuko; Akiyama, Hidehiko; Soloshonok, Vadim A.; Inoue, Yuki; Hara, Hideaki
2017-01-01
Over the last few years, thalidomide has become one of the most important anti-tumour drugs for the treatment of relapsed-refractory multiple myeloma. However, besides its undesirable teratogenic side effect, its configurational instability critically limits any further therapeutic improvements of this drug. In 1999, we developed fluoro-thalidomide which is a bioisostere of thalidomide, but, in sharp contrast to the latter, it is configurationally stable and readily available in both enantiomeric forms. The biological activity of fluoro-thalidomide however, still remains virtually unstudied, with the exception that fluoro-thalidomide is not teratogenic. Herein, we report the first biological evaluation of fluoro-thalidomide in racemic and in both (R)- and (S)-enantiomerically pure forms against (in vitro) H929 cells of multiple myeloma (MM) using an annexin V assay. We demonstrate that all fluoro-thalidomides inhibited the growth of H929 MM cells without any in-vivo activation. Furthermore, we report that the enantiomeric forms of fluoro-thalidomide display different anti-tumour activities, with the (S)-enantiomer being noticeably more potent. The angiogenesis of fluoro-thalidomides is also investigated and compared to thalidomide. The data obtained in this study paves the way towards novel pharmaceutical research on fluoro-thalidomides. PMID:28763493
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Jaber, Basem M; Petroianu, Georg A; Rizvi, Syed A; Borai, Anwar; Saleh, Nada A; Hala, Sharif M; Saleh, Ayman M
2018-03-01
This study was performed to evaluate the protective efficacy of metoclopramide (MCP) against the organophosphates paraoxon (POX)- and malathion (MLT)-induced apoptosis in the murine L929 skin fibroblasts. L929 cells were exposed to either POX (10 nm) or 1.0 μm MLT in the absence and presence of increased concentrations of MCP. The protective effect of MCP on these organophosphate-stimulated apoptotic events was evaluated by flow cytometry analysis after staining with annexin-V/propidium iodide, processing and activation of the executioner caspase-3, cleavage of the poly-ADP ribose polymerase, fragmentation of the nucleosomal DNA and disruption of the mitochondrial membrane potential (Δψ). Our results showed that increased doses of MCP alone (≥10 μm) did not induce apoptosis or activation of caspase-3. Pretreatment of the cells with MCP attenuated all the apoptotic events triggered by the organophosphate compounds in a dose-dependent manner reaching ~70-80% protection when they were preincubated at 1 and 5 μm of the drug before the addition of POX and MLT, respectively. Interestingly, MCP did not offer a significant protective effect against the cytotoxicity of tumor necrosis factor-α, cisplatinum, etoposide or paclitaxel, which stimulate apoptosis by various mechanisms, suggesting that the anti-apoptotic effect of the drug is specific to organophosphates. The strong and specific anti-apoptotic activity of subclinical doses of MCP against the cytotoxicity of organophosphate compounds suggests its potential clinical application in treating their poisoning. Copyright © 2017 John Wiley & Sons, Ltd.
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Zhang, Mengjiao; Chen, Xianchun; Pu, Ximing; Liao, Xiaoming; Huang, Zhongbing; Yin, Guangfu
2014-04-01
The effects in vitro of a novel multiphase glass-ceramic (with nominal composition of 43.19% CaO, 7.68% MgO, and 49.13% SiO2 in weight percent) on cell adhesion, proliferation, differentiation and ultrastructure of human osteosarcoma cell line MG63, mouse fibroblasts L929, and human lung adenocarcinoma epithelial cell line A549 were investigated in this research. Scanning electron microscopy (SEM) micrographs revealed that the surface morphology of this glass-ceramic was beneficial to cell adhesion. The glass-ceramic extracts at certain concentrations could stimulate the proliferation and differentiation of MG63 and L929 cells, whereas inhibit A549 proliferation, which might be resulted from the released Si ions. In addition, when cultured with 0.1mg/mL glass-ceramic powder suspension, the cell ultrastructure of MG63 showed abundant organelles and L929 displayed the phenomena of cellular stress response. While more interestingly, A549 exhibited chromatin condensation, mitochondria swell and RER expansion, which was presumed to be early signs of apoptosis. These results suggest that this novel CaO-MgO-SiO2-based multiphase glass-ceramic has potential for bone regeneration and tissue engineering applications. Copyright © 2013 Elsevier B.V. All rights reserved.
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de S Wisintainer, G G N; Scola, G; Moura, S; Lemos, T L G; Pessoa, C; de Moraes, M O; Souza, L G S; Roesch-Ely, M; Henriques, J A P
2015-12-21
Biflorin is an o-naphthoquinone isolated from the roots of the plant Capraria biflora L. (Scrophulariaceae). In this study, the cytotoxic effects of biflorin were verified, and late apoptosis was detected in various cancer cell lines by in situ analysis. The cytotoxicity was further evaluated exclusively for 48 h of treatment in different tumor and non-tumor cell lines (Hep-2, HeLa, HT-29, A-375, and A-549, and HEK-293, respectively). The results indicated that biflorin induced selective cytotoxicity in tumor cells. HeLa cells were more susceptible to biflorin, followed by HT-29, A-549, A-375, and Hep-2 at all concentrations (range 5-50 μg/mL), and the highest half-maximal inhibitory concentration IC50 (56.01 ± 1.17 μg/mL) was observed in HEK-293 cells. Late apoptotic/necrotic events, observed by in situ immunostaining with Annexin V, varied with each cell line; an increase in late apoptotic events was observed corresponding to the increase in biflorin dosage. Hep-2 cells showed a greater percentage of late apoptotic events among the tumor cell lines when treated with higher concentrations of biflorin (69.63 ± 2.28%). The non-tumor HEK-293 line showed greater resistance to late apoptotic events, as well as a lower level of cytotoxicity (77.69 ± 6.68%) than the tested tumor lines. The data presented indicate that biflorin showed an important, possibly selective, cytotoxicity against tumor cell lines, thereby revealing a promising novel substance with potential anticancer activity for tumor therapy.
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Waiwijit, Uraiwan; Maturos, Thitima; Pakapongpan, Saithip; Phokharatkul, Ditsayut; Wisitsoraat, Anurat; Tuantranont, Adisorn
2016-08-01
Recently, three-dimensional graphene interconnected network has attracted great interest as a scaffold structure for tissue engineering due to its high biocompatibility, high electrical conductivity, high specific surface area and high porosity. However, free-standing three-dimensional graphene exhibits poor flexibility and stability due to ease of disintegration during processing. In this work, three-dimensional graphene is composited with polydimethylsiloxane to improve the structural flexibility and stability by a new simple two-step process comprising dip coating of polydimethylsiloxane on chemical vapor deposited graphene/Ni foam and wet etching of nickel foam. Structural characterizations confirmed an interconnected three-dimensional multi-layer graphene structure with thin polydimethylsiloxane scaffold. The composite was employed as a substrate for culture of L929 fibroblast cells and its cytocompatibility was evaluated by cell viability (Alamar blue assay), reactive oxygen species production and vinculin immunofluorescence imaging. The result revealed that cell viability on three-dimensional graphene/polydimethylsiloxane composite increased with increasing culture time and was slightly different from a polystyrene substrate (control). Moreover, cells cultured on three-dimensional graphene/polydimethylsiloxane composite generated less ROS than the control at culture times of 3-6 h. The results of immunofluorescence staining demonstrated that fibroblast cells expressed adhesion protein (vinculin) and adhered well on three-dimensional graphene/polydimethylsiloxane surface. Good cell adhesion could be attributed to suitable surface properties of three-dimensional graphene/polydimethylsiloxane with moderate contact angle and small negative zeta potential in culture solution. The results of electrochemical study by cyclic voltammetry showed that an oxidation current signal with no apparent peak was induced by fibroblast cells and the oxidation current at an
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Hasegawa, Kiyoshi; Suzuki, Machiko; Ishikawa, Kunimi; Yasue, Akira; Kato, Rina; Nakamura, Azumi; Kuroki, Jun; Udagawa, Yasuhiro
2003-03-01
A new cell line of human uterine endometrial undifferentiated carcinoma, designated as TMG-L, was established from the metastatic lymph node of 56-year-old patient TMG-L cells have been cultured with Ham's F-12 medium supplemented with 10% FCS and grew as a loosely adherent monolayer with polygonal or spindle-shaped cells exhibiting poor cell-cell contact and piled up against each other, showing a tendency to grow as floating cells. The doubling time of this cell line was about 48 hours, and chromosomal analysis revealed aneuploidy at passage 25. The cells formed tumors in SCID mouse, the histology of which was similar to that of undifferentiated carcinoma component of primary tumor. TMG-L cells showed the loss of expression and membranous localization of either E-cadherin or alpha-catenin, implied corresponding loss of their adhesive function. And this dysfunction implicated the biological aggressive behavior of uterine endometrial undifferentiated carcinoma. This cell line appears to provide a useful system for studying uterine undifferentiated carcinoma in vivo and in vitro.
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Rumex L. species induce apoptosis in 1301, EOL-1 and H-9 cell lines.
Wegiera, Magdalena; Smolarz, Helena D; Bogucka-Kocka, Anna
2012-01-01
The Rumex L. (dock) species for many centuries have been used in medical treatment, through their adstringent, spasmolitic or cholagogic action. In the present study, the in vitro screening of cytotoxic activities of ethanol extract from roots, leaves and fruits of six Rumex species: R. acetosa L., R. acetosella L., R. confertus Willd., R. crispus L., R. hydrolapathum Huds. and R. obtusifolius L. were performed. We found remarkable cytotoxic activities on leukemic 1301 and EOL-1 cell lines and T cell line at concentration dependent manners. Cytotoxic activity was determined in two ways: trypan blue test and annexin-V FITC and propidium iodide assay. Received IC50 values of investigated extracts on 1301, EOL-1 and H-9 cell lines ranged from 0.22, 0.17 and 0.04 to 2.56, 1.91 and 1.83 mg/mL, respectively. Analysis of morphological changes demonstrated that the extract exerted cell-death via apoptosis. The overall activities of Rumex species support the traditional use of the extract from the fruits of R. confertus, R. crispus, R. hydrolapathum and R. obtusifolius in the treatment of cancer.
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[Establishment of L-periaxin gene knock-out RSC96 cell line].
Liang, Min; Peng, Tingting; Shi, Yawei
2016-12-25
Periaxin, a protein of noncompact myelin, is specifically expressed in the peripheral nervous system (PNS). There are two protein isoform L-periaxin and S-Periaxin by alternative splicing of periaxin gene, playing an important role in the initiation of myelin formation. So far, 18 different mutation sites in L-periaxin gene have been found to induce the peripheral demyelinating neurological charcot-marie-tooth diseases subtype 4F (CMT4F). The technique of activation of transcription activator-like effector nucleases (TALENS) was used to knock out the L-periaxin gene in RSC 96 cell line of Rattus. According to the design principle, the knock-out site of L-periaxin was assured to NLS domain of L-periaxin, which is target sequence of left and right arms of TALEN. The knock-out vectors of TALEN-L and TALEN-R were established and transfected into RSC96 cell. After puromycin screening, L-periaxin was knocked out successfully in RSC96 cell, which is confirmed by DNA sequence. The mutation efficiency is 21.6%. S-periaxin, not L-periaxin can be detected by Western blotting in L-periaxin gene knock-out RSC96 cell. The cell growth rate was decreased and the number of cells in G1 increased and decreased in S phase in L-periaxin gene knock-out RSC96 cell by flow cytometry and MTT assay.
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Influence of PD-L1 cross-linking on cell death in PD-L1-expressing cell lines and bovine lymphocytes
Ikebuchi, Ryoyo; Konnai, Satoru; Okagawa, Tomohiro; Yokoyama, Kazumasa; Nakajima, Chie; Suzuki, Yasuhiko; Murata, Shiro; Ohashi, Kazuhiko
2014-01-01
Programmed death-ligand 1 (PD-L1) blockade is accepted as a novel strategy for the reactivation of exhausted T cells that express programmed death-1 (PD-1). However, the mechanism of PD-L1-mediated inhibitory signalling after PD-L1 cross-linking by anti-PD-L1 monoclonal antibody (mAb) or PD-1–immunogloblin fusion protein (PD-1-Ig) is still unknown, although it may induce cell death of PD-L1+ cells required for regular immune reactions. In this study, PD-1-Ig or anti-PD-L1 mAb treatment was tested in cell lines that expressed PD-L1 and bovine lymphocytes to investigate whether the treatment induces immune reactivation or PD-L1-mediated cell death. PD-L1 cross-linking by PD-1-Ig or anti-PD-L1 mAb primarily increased the number of dead cells in PD-L1high cells, but not in PD-L1low cells; these cells were prepared from Cos-7 cells in which bovine PD-L1 expression was induced by transfection. The PD-L1-mediated cell death also occurred in Cos-7 and HeLa cells transfected with vectors only encoding the extracellular region of PD-L1. In bovine lymphocytes, the anti-PD-L1 mAb treatment up-regulated interferon-γ (IFN-γ) production, whereas PD-1-Ig treatment decreased this cytokine production and cell proliferation. The IFN-γ production in B-cell-depleted peripheral blood mononuclear cells was not reduced by PD-1-Ig treatment and the percentages of dead cells in PD-L1+ B cells were increased by PD-1-Ig treatment, indicating that PD-1-Ig-induced immunosuppression in bovine lymphocytes could be caused by PD-L1-mediated B-cell death. This study provides novel information for the understanding of signalling through PD-L1. PMID:24405267
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Chen, Lixiang; Wang, Cong; Li, Shun; Yu, Xin; Liu, Xue; Ren, Rongrong; Liu, Wenwen; Zhou, Xiaojing; Zhang, Xiaonan; Zhou, Xiaohui
2016-04-28
Chlamydiae, obligate intracellular bacteria, are associated with a variety of human diseases. The chlamydial life cycle undergoes a biphasic development: replicative reticulate bodies (RBs) phase and infectious elementary bodies (EBs) phase. At the end of the chlamydial intracellular life cycle, EBs have to be released to the surrounded cells. Therefore, the interactions between Chlamydiae and cell death pathways could greatly influence the outcomes of Chlamydia infection. However, the underlying molecular mechanisms remain elusive. Here, we investigated host cell death after Chlamydia infection in vitro, in L929 cells, and showed that Chlamydia infection induces cell necrosis, as detected by the propidium iodide (PI)-Annexin V double-staining flow-cytometric assay and Lactate dehydrogenase (LDH) release assay. The production of reactive oxygen species (ROS), an important factor in induction of necrosis, was increased after Chlamydia infection, and inhibition of ROS with specific pharmacological inhibitors, diphenylene iodonium (DPI) or butylated hydroxyanisole (BHA), led to significant suppression of necrosis. Interestingly, live-cell imaging revealed that Chlamydia infection induced lysosome membrane permeabilization (LMP). When an inhibitor upstream of LMP, CA-074-Me, was added to cells, the production of ROS was reduced with concomitant inhibition of necrosis. Taken together, our results indicate that Chlamydia infection elicits the production of ROS, which is dependent on LMP at least partially, followed by induction of host-cell necrosis. To our best knowledge, this is the first live-cell-imaging observation of LMP post Chlamydia infection and report on the link of LMP to ROS to necrosis during Chlamydia infection.
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[Effect of DOT1L gene silence on proliferation of acute monocytic leukemia cell line THP-1].
Zhang, Yu-Juan; Li, Hua-Wen; Chang, Guo-Qiang; Zhang, Hong-Ju; Wang, Jian; Lin, Ya-Ni; Zhou, Jia-Xi; Li, Qing-Hua; Pang, Tian-Xiang
2013-08-01
This study was aimed to investigate the influence of short hairpin RNA (shRNA) on proliferation of human leukemia cell line THP-1. The shRNA targeting the site 732-752 of DOT1L mRNA was designed and chemically synthesized, then a single-vector lentiviral, tet-inducible shRNA-DOT1L system (Plko-Tet-On) was generated. Thereafter, the THP-1 cells with lentivirus were infected to create stable cell line with regulatable shRNA expression. The expression of DOT1L in the THP-1 cell line was assayed by RT-PCR. Effect of shRNA-DOT1L on the proliferation of THP-1 cells was detected with MTT method,and the change of colony forming potential of THP-1 cells was analyzed by colony forming unit test. Cell cycle distribution was tested by flow cytometry. The results indicated that the expression of DOT1L was statistically lower than that in the control groups. The proliferation and colony forming capacity of THP-1 cells were significantly inhibited. The percentage of cells at G0/G1 phase increased in THP-1/shRNA cells treated with Dox while the percentage of cells at S phase significantly decreased as compared with that in the control group. It is concluded that the shRNA targeting DOT1L can effectively inhibit the proliferation of acute monocytic leukemia cell line THP-1.
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Bello, Felio J; Mejía, Astrid J; Corena, María del Pilar; Ayala, Martha; Sarmiento, Ladys; Zuñiga, Claudio; Palau, María T
2005-10-01
The present work describes the in vitro infection of a cell line Lulo, derived from Lutzomyia longipalpis embryonic tissue, by Leishmania chagasi promastigotes. This infection process is compared with a parallel one developed using the J774 cell line. The L. chagasi MH/CO/84/CI-044B strain was used for experimental infection in two cell lines. The cells were seeded on glass coverslips in 24-well plates to reach a final number of 2 x 10(5) cells/well. Parasites were added to the adhered Lulo and J774 cells in a 10:1 ratio and were incubated at 28 and 37 masculineC respectively. After 2, 4, 6, 8, and 10 days post-infection, the cells were extensively washed with PBS, fixed with methanol, and stained with Giemsa. The number of internalized parasites was determined by counting at least 400 cultured cells on each coverslip. The results showed continuous interaction between L. chagasi promastigotes with the cell lines. Some ultrastructural characteristics of the amastigote forms were observed using transmission electron microscopy. The highest percentage of infection in Lulo cells was registered on day 6 post-infection (29.6%) and on day 4 in the J774 cells (51%). This work shows similarities and differences in the L. chagasi experimental infection process in the two cell lines. However, Lulo cells emerge as a new model to study the life-cycle of this parasite.
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Wong, Yu Hua; Tan, Wai Yan; Tan, Chin Ping; Long, Kamariah; Nyam, Kar Lin
2014-05-01
To examine the cytotoxic properties of both the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cervical cancer, human breast cancer, human colon cancer and human lung cancer cell lines. The in vitro cytotoxic activity of the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cancer cell lines was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and sulforhodamine B assays. Cell morphological changes were observed by using an inverted light microscope. The kenaf seed extract (KSE) exhibited a lower IC50 than kenaf seed oil (KSO) in all of the cancer cell lines. Morphological alterations in the cell lines after KSE and KSO treatment were observed. KSE and KSO possessed effective cytotoxic activities against all the cell lines been selected. KSE and KSO could be potential sources of natural anti-cancer agents. Further investigations on using kenaf seeds for anti-proliferative properties are warranted.
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Wong, Yu Hua; Tan, Wai Yan; Tan, Chin Ping; Long, Kamariah; Nyam, Kar Lin
2014-01-01
Objective To examine the cytotoxic properties of both the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cervical cancer, human breast cancer, human colon cancer and human lung cancer cell lines. Methods The in vitro cytotoxic activity of the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cancer cell lines was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and sulforhodamine B assays. Cell morphological changes were observed by using an inverted light microscope. Results The kenaf seed extract (KSE) exhibited a lower IC50 than kenaf seed oil (KSO) in all of the cancer cell lines. Morphological alterations in the cell lines after KSE and KSO treatment were observed. KSE and KSO possessed effective cytotoxic activities against all the cell lines been selected. Conclusions KSE and KSO could be potential sources of natural anti-cancer agents. Further investigations on using kenaf seeds for anti-proliferative properties are warranted. PMID:25183141
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Spontaneous generation of germline characteristics in mouse fibrosarcoma cells
NASA Astrophysics Data System (ADS)
Ma, Zhan; Hu, Yao; Jiang, Guoying; Hou, Jun; Liu, Ruilai; Lu, Yuan; Liu, Chunfang
2012-10-01
Germline/embryonic-specific genes have been found to be activated in somatic tumors. In this study, we further showed that cells functioning as germline could be present in mouse fibrosarcoma cells (L929 cell line). Early germline-like cells spontaneously appeared in L929 cells and further differentiated into oocyte-like cells. These germline-like cells can, in turn, develop into blastocyst-like structures in vitro and cause teratocarcinomas in vivo, which is consistent with natural germ cells in function. Generation of germline-like cells from somatic tumors might provide a novel way to understand why somatic cancer cells have strong features of embryonic/germline development. It is thought that the germline traits of tumors are associated with the central characteristics of malignancy, such as immortalization, invasion, migration and immune evasion. Therefore, germline-like cells in tumors might provide potential targets to tumor biology, diagnosis and therapy.
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Ye, Yuan-Chao; Wang, Hong-Ju; Yu, Lu; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi
2012-12-01
Tumor necrosis factor alpha (TNFα) induces necroptosis and autophagy; however, the detailed molecular mechanism is not fully understood. In this study, we found that TNFα administration caused mitochondrial dysfunction and reactive oxygen species (ROS) production, which led to necroptosis and autophagy in murine fibrosarcoma L929 cells. Notably, the RIP1 (serine-threonine kinase receptor-interacting protein 1, a main adaptor protein of necroptosis) specific inhibitor necrostatin-1 (Nec-1) recovered mitochondrial dysfunction and ROS production due to TNFα administration. Moreover, pan-caspase inhibitor z-VAD-fmk (zVAD) increased RIP1 expression and exacerbated TNFα-induced mitochondrial dysfunction and ROS production, indicating that RIP1 led to mitochondrial dysfunction and ROS production. In addition, cytochrome c release from mitochondria was accompanied with TNFα administration, and Nec-1 blocked the release of cytochrome c upon TNFα administration, while zVAD enhanced the release. These further suggested that RIP1 induced mitochondrial dysfunction accompanied with cytochrome c release. Furthermore, autophagy inhibitor 3-methyladenine (3MA) did not affect RIP1 expression as well as mitochondrial dysfunction and ROS production. Together with our previous publication that autophagy was a downstream consequence of necroptosis, we concluded that TNFα induced mitochondrial dysfunction accompanied with ROS production and cytochrome c release via RIP1, leading to necroptosis and resulting autophagic cell death. Copyright © 2012 Elsevier B.V. All rights reserved.
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Salazar-Aguilar, Sandra; Ruiz-Posadas, Lucero Del Mar; Cadena-Iñiguez, Jorge; Soto-Hernández, Marcos; Santiago-Osorio, Edelmiro; Aguiñiga-Sánchez, Itzen; Rivera-Martínez, Ana Rocío; Aguirre-Medina, Juan Francisco
2017-07-25
The Sechium edule Perla Negra cultivar is a recently-obtained biological material whose progenitors are S. edule var. nigrum minor and S. edule var. amarus silvestrys, the latter of which has been reported to have antiproliferative activity against the HeLa P-388 and L-929 cancer cell lines. The present study aimed to determine if the methanolic extract of the fruit of the Perla Negra cultivar had the same biological activity. The methanolic extract was phytochemically characterized by thin layer chromatography (TLC) and column chromatography (CC), identifying the terpenes and flavonoids. The compounds identified via high performance liquid chromatography (HPLC) were Cucurbitacins B, D, E, and I for the terpene fractions, and Rutin, Phlorizidin, Myricetin, Quercetin, Naringenin, Phloretin, Apigenin, and Galangin for the flavonoid fractions). Biological activity was evaluated with different concentrations of the methanolic extract in the HeLa cell line and normal lymphocytes. The methanolic extract inhibited the proliferation of HeLa cells (IC 50 1.85 µg·mL -1 ), but the lymphocytes were affected by the extract (IC 50 30.04 µg·mL -1 ). Some fractions, and the pool of all of them, showed inhibition higher than 80% at a concentration of 2.11 µg·mL -1 . Therefore, the biological effect shown by the methanolic extract of the Perla Negra has some specificity in inhibiting tumor cells and not normal cells; an unusual feature among molecules investigated as potential biomedical agents.
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L-N-Acetylcysteine protects against radiation-induced apoptosis in a cochlear cell line.
Low, Wong-Kein; Sun, Li; Tan, Michelle G K; Chua, Alvin W C; Wang, De-Yun
2008-04-01
L-N-Acetylcysteine (L-NAC) significantly reduced reactive oxygen species (ROS) generation and cochlear cell apoptosis after irradiation. The safe and effective use of L-NAC in reducing radiation-induced sensorineural hearing loss (SNHL) should be verified by further in vivo studies. Radiation-induced SNHL is a common complication after radiotherapy of head and neck tumours. There is growing evidence to suggest that ROS play an important role in apoptotic cochlear cell death from ototoxicity, resulting in SNHL. The aim of this study was to evaluate the effectiveness of L-NAC, an antioxidant, on radiation-induced apoptosis in cochlear cells. The OC-k3 cochlear cell line was studied after 0 and 20 Gy of gamma-irradiation. Cell viability assay was performed using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide. Flow cytometry and TUNEL assay were done with and without the addition of 10 mmol/L of L-NAC. Intracellular generation of ROS was detected by 2',7'-dichlorofluorescein diacetate, with comparisons made using fluorescence intensity. L-NAC increased the viability of cells after irradiation. Generation of ROS was demonstrated at 1 h post-irradiation and was significantly reduced by L-NAC (p<0.0001). Flow cytometry and TUNEL assay showed cell apoptosis at 72 h post-irradiation, which was diminished by the addition of L-NAC.
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Infection of nonphagocytic host cells by legionella.
Day, Shandra R; Sifri, Costi D; Hoffman, Paul S
2013-01-01
Legionella pneumophila is an intracellular pathogen of free-living protozoa that can also infect alveolar macrophages, L929 fibroblast cells, and HeLa cells. Infection of nonphagocytic cells by L. pneumophila can be used to study invasion mechanisms, compare infectivity of different strains and identify factors important for virulence. Virulent strains of L. pneumophila exposed to monolayers of L929 cells are able to invade and form virus-like plaques, which can be enumerated as a measure of infectivity. Invasiveness of HeLa cells can also be used to evaluate relative infectivity and to study mechanisms of invasion and to track the development of cyst-like forms. The detailed methods of both the L929 plaque assay and HeLa cell invasion assay are described.
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Evaluation of anticancer potential of Bacopa monnieri L. against MCF-7 and MDA-MB 231 cell line.
Mallick, Md Nasar; Akhtar, Md Salman; Najm, Mohd Zeeshan; Tamboli, E T; Ahmad, Sayeed; Husain, Syed Akhtar
2015-01-01
The ethanolic extract of Bacopa monnieri contains bacoside A and B, brahmin, cucurbitacins, and betulinic acid. Currently, cucurbitacins have also been reported for their strong anti-tumorigenic and anti-proliferative activity by inducing cell cycle arrest at the G2/M phase and formation of multiplied cells. The present study was carried out to evaluate the in vitro cytotoxic activity of ethanolic extract of dichloromethane (DCM) fraction of B. monnieri on two different cell lines. The ethanolic extract of B. monnieri was prepared using soxhlet extraction method and different fractions (hexane, DCM, methanol, acetone, and water) of ethanolic extracts were prepared. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay of ethanolic extract and of all fractions was carried out on MCF-7 and MDA-MB 231 cell lines. The presence of cucurbitacins and betulinic acid in these fractions was confirmed by high-performance thin layer chromatography. The IC50 values of ethanolic extract of B. monnieri in MCF-7 and MDA-MB 231 cell lines were 72.0 μg/mL and 75.0 μg/mL, respectively. The DCM fraction of B. monnieri showed maximum cytotoxic activity among all fraction upto 72 h and was found to be 57.0 μg/mL and 42.0 μg/mL, respectively. The results showed good cytotoxic activity in DCM fraction in both the cell lines may be due to the presence of cucurbitacins and betulinic acid in DCM fraction.
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Evaluation of anticancer potential of Bacopa monnieri L. against MCF-7 and MDA-MB 231 cell line
Mallick, Md. Nasar; Akhtar, Md. Salman; Najm, Mohd. Zeeshan; Tamboli, E. T.; Ahmad, Sayeed; Husain, Syed Akhtar
2015-01-01
Background: The ethanolic extract of Bacopa monnieri contains bacoside A and B, brahmin, cucurbitacins, and betulinic acid. Currently, cucurbitacins have also been reported for their strong anti-tumorigenic and anti-proliferative activity by inducing cell cycle arrest at the G2/M phase and formation of multiplied cells. The present study was carried out to evaluate the in vitro cytotoxic activity of ethanolic extract of dichloromethane (DCM) fraction of B. monnieri on two different cell lines. Materials and Methods: The ethanolic extract of B. monnieri was prepared using soxhlet extraction method and different fractions (hexane, DCM, methanol, acetone, and water) of ethanolic extracts were prepared. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay of ethanolic extract and of all fractions was carried out on MCF-7 and MDA-MB 231 cell lines. The presence of cucurbitacins and betulinic acid in these fractions was confirmed by high-performance thin layer chromatography. Results: The IC50 values of ethanolic extract of B. monnieri in MCF-7 and MDA-MB 231 cell lines were 72.0 μg/mL and 75.0 μg/mL, respectively. The DCM fraction of B. monnieri showed maximum cytotoxic activity among all fraction upto 72 h and was found to be 57.0 μg/mL and 42.0 μg/mL, respectively. Conclusion: The results showed good cytotoxic activity in DCM fraction in both the cell lines may be due to the presence of cucurbitacins and betulinic acid in DCM fraction. PMID:26681894
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Kobayashi, Masaki; Fujita, Itaru; Itagaki, Shirou; Hirano, Takeshi; Iseki, Ken
2005-07-01
Monocarboxylate transporter (MCT), which cotransport L-lactic acid and protons across cell membranes, are important for regulation of muscle pH. However, it has not been demonstrated in detail whether MCT isoform contribute to the transport of L-lactic acid in skeletal muscle. The aim of this study was to characterize L-lactic acid transport using an human rhabdomyosarcoma (RD) cell line as a model of human skeletal muscle. mRNAs of MCT 1, 2 and 4 were found to be expressed in RD cells. The [14C] L-lactic acid uptake was concentration-dependent with a Km of 1.19 mM. This Km value was comparable to its Km values for MCT1 or MCT2. MCT1 mRNA was found to be present markedly greater than that MCT2. Therefore, MCT1 most probably acts on L-lactic acid uptake at RD cells. [14C] L-Lactic acid efflux in RD cells was inhibited by alpha-cyano-4-hydroxycinnamate (CHC) but not by butyric acid, a substrate of MCT1. Accordingly, MCT2 or MCT4 is responsible for L-lactic acid efflux by RD cells. MCT4 mRNA was found to be present significantly greater than that MCT2. We conclude that MCT1 is responsible for L-lactic acid uptake and L-lactic acid efflux is mediated by MCT4 in RD cells.
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Selection. 929.23 Section 929.23 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Administrative Body § 929.23 Selection...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Cranberries. 929.5 Section 929.5 Agriculture... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.5 Cranberries...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Cranberries. 929.5 Section 929.5 Agriculture... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.5 Cranberries...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Cranberries. 929.5 Section 929.5 Agriculture... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.5 Cranberries...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Cranberries. 929.5 Section 929.5 Agriculture... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.5 Cranberries...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Cranberries. 929.5 Section 929.5 Agriculture... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.5 Cranberries...
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Code of Federal Regulations, 2010 CFR
2010-10-01
... 42 Public Health 5 2010-10-01 2010-10-01 false Chemistry. 493.929 Section 493.929 Public Health... Proficiency Testing Programs by Specialty and Subspecialty § 493.929 Chemistry. The subspecialties under the specialty of chemistry for which a proficiency testing program may offer proficiency testing are routine...
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Code of Federal Regulations, 2012 CFR
2012-10-01
... 42 Public Health 5 2012-10-01 2012-10-01 false Chemistry. 493.929 Section 493.929 Public Health... Proficiency Testing Programs by Specialty and Subspecialty § 493.929 Chemistry. The subspecialties under the specialty of chemistry for which a proficiency testing program may offer proficiency testing are routine...
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Code of Federal Regulations, 2014 CFR
2014-10-01
... 42 Public Health 5 2014-10-01 2014-10-01 false Chemistry. 493.929 Section 493.929 Public Health... Proficiency Testing Programs by Specialty and Subspecialty § 493.929 Chemistry. The subspecialties under the specialty of chemistry for which a proficiency testing program may offer proficiency testing are routine...
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Code of Federal Regulations, 2013 CFR
2013-10-01
... 42 Public Health 5 2013-10-01 2013-10-01 false Chemistry. 493.929 Section 493.929 Public Health... Proficiency Testing Programs by Specialty and Subspecialty § 493.929 Chemistry. The subspecialties under the specialty of chemistry for which a proficiency testing program may offer proficiency testing are routine...
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Code of Federal Regulations, 2011 CFR
2011-10-01
... 42 Public Health 5 2011-10-01 2011-10-01 false Chemistry. 493.929 Section 493.929 Public Health... Proficiency Testing Programs by Specialty and Subspecialty § 493.929 Chemistry. The subspecialties under the specialty of chemistry for which a proficiency testing program may offer proficiency testing are routine...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Records. 929.63 Section 929.63 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.63 Records. Each handler shall maintain such records of all cranberries acquired, withheld from handling, handled, and...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Records. 929.63 Section 929.63 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.63 Records. Each handler shall maintain such records of all cranberries acquired, withheld from handling, handled, and...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Records. 929.63 Section 929.63 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.63 Records. Each handler shall maintain such records of all cranberries acquired, withheld from handling, handled, and...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Secretary. 929.1 Section 929.1 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.1 Secretary. Secretary means the Secretary of Agriculture of the United States, or any officer or employee of the United States...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Secretary. 929.1 Section 929.1 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.1 Secretary. Secretary means the Secretary of Agriculture of the United States, or any officer or employee of the United States...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Secretary. 929.1 Section 929.1 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.1 Secretary. Secretary means the Secretary of Agriculture of the United States, or any officer or employee of the United States...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Secretary. 929.1 Section 929.1 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.1 Secretary. Secretary means the Secretary of Agriculture of the United States, or any officer or employee of the United States...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Secretary. 929.1 Section 929.1 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.1 Secretary. Secretary means the Secretary of Agriculture of the United States, or any officer or employee of the United States...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Reports. 929.62 Section 929.62 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.62 Reports. (a) Grower report. Each grower shall file a report with the committee by January 15 of each crop year, or...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Reports. 929.62 Section 929.62 Agriculture Regulations... ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.62 Reports. (a) Grower report. Each grower shall file a report with the committee by January 15 of each crop year, or...
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Tsukamoto, Akira; Higashiyama, Satoru; Yoshida, Kenji; Watanabe, Yoshiaki; Furukawa, Katsuko S; Ushida, Takashi
2011-12-01
An increase in cytoplasmic calcium (Ca(2+) increase) is a second messenger that is often observed under ultrasound irradiation. We hypothesize that cavitation is a physical mechanism that underlies the increase in Ca(2+) in these experiments. To control the presence of cavitation, the wave type was controlled in a sonication chamber. One wave type largely contained a traveling wave (wave type A) while the other wave type largely contained a standing wave (wave type B). Fast Fourier transform (FFT) analysis of a sound field produced by the wave types ascertained that stable cavitation was present only under wave type A ultrasound irradiation. Under the two controlled wave types, the increase in Ca(2+) in L929 fibroblasts was observed with fluorescence imaging. Under wave type A ultrasound irradiation, an increase in Ca(2+) was observed; however, no increase in Ca(2+) was observed under wave type B ultrasound irradiation. We conclude that stable cavitation is involved in the increase of Ca(2+) in cells subjected to pulsed ultrasound. Copyright © 2011 Elsevier B.V. All rights reserved.
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Oktyabri, Dulamsuren; Ishimura, Akihiko; Tange, Shoichiro; Terashima, Minoru; Suzuki, Takeshi
2016-04-01
DOT1L is a histone H3 lysine 79 (H3K79) methyltransferase mainly implicated in leukemia. Here we analyzed the function of DOT1L in breast cancer cells. The expression of DOT1L was up-regulated in malignant breast cancer tissues. Over-expression of DOT1L significantly increased the sphere formation and the cell migration activities of MCF7 breast cancer cell line. In contrast, knockdown of DOT1L reduced the cell migration activity of MDA-MB-231 breast cancer cell line. BCAT1, which encodes a branched-chain amino acid transaminase, was identified as one of the target genes controlled by DOT1L through the regulation of H3K79 methylation. Mechanistic investigation revealed that BCAT1 might be an important regulator responsible for DOT1L-mediated sphere formation and cell migration in breast cancer cells. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.
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Li, Qian; Peng, Jie; Liu, Ting; Zhang, Guiying
2017-09-01
Fas, which is an apoptotic-related protein, has an important role in cell apoptosis. Fas ligand (FasL) binds to Fas and activates apoptosis signal transduction. We previously demonstrated that the efficiency of celecoxib inhibited the proliferation and apoptosis of HT-29 colon cancer cell line. The BGC823 cell line was used as an experimental model to evaluate the potential role of celecoxib on gastric cancer cell apoptosis. Inhibitory effects of celecoxib on cell viability were determined by MTT assay. Cell apoptosis was evaluated by flow cytometric analysis and laser confocal microscopy. The results of the present study demonstrated that celecoxib inhibited the viability of BGC823 cells in a concentration- and time-dependent manner. Furthermore, the effect of BGC823 cells apoptosis was increased in a concentration-dependent manner. Western blotting was used to determine the protein expression levels of Fas, FasL, and B-cell lymphoma-2 (Bcl-2). During the celecoxib-induced apoptosis of BGC823 cells, celecoxib upregulated Fas expression and downregulated FasL and Bcl-2 expression in a concentration-dependent manner. These results suggest that celecoxib inhibited the growth and induced apoptosis of BGC823 gastric cancer cells by regulating the protein expression of Fas, FasL and Bcl-2.
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7 CFR 929.15 - Annual allotment.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Annual allotment. 929.15 Section 929.15 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... by multiplying such grower's sales history by the allotment percentage established pursuant to § 929...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Fiscal period. 929.6 Section 929.6 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.6 Fiscal period...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Fiscal period. 929.6 Section 929.6 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.6 Fiscal period...
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7 CFR 929.46 - Marketing policy.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Marketing policy. 929.46 Section 929.46 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.46 Marketing policy...
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7 CFR 929.46 - Marketing policy.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Marketing policy. 929.46 Section 929.46 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.46 Marketing policy...
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7 CFR 929.46 - Marketing policy.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Marketing policy. 929.46 Section 929.46 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.46 Marketing policy...
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7 CFR 929.46 - Marketing policy.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Marketing policy. 929.46 Section 929.46 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.46 Marketing policy...
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7 CFR 929.46 - Marketing policy.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Marketing policy. 929.46 Section 929.46 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.46 Marketing policy...
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7 CFR 929.59 - Excess cranberries.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Excess cranberries. 929.59 Section 929.59 Agriculture... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.59 Excess cranberries...
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7 CFR 929.59 - Excess cranberries.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Excess cranberries. 929.59 Section 929.59 Agriculture... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.59 Excess cranberries...
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7 CFR 929.59 - Excess cranberries.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Excess cranberries. 929.59 Section 929.59 Agriculture... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.59 Excess cranberries...
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7 CFR 929.59 - Excess cranberries.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Excess cranberries. 929.59 Section 929.59 Agriculture... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.59 Excess cranberries...
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Kulesh, D A; Anderson, L H; Wilson, B; Otis, E J; Elgin, D M; Barker, M J; Mehm, W J; Kearney, G P
1994-08-01
Myoblast cell cultures have been widely employed in conventional (1g) studies of biological processes because characteristics of intact muscle can be readily observed in these cultured cells. We decided to investigate the effects of spaceflight on muscle by utilizing a well characterized myoblast cell line (L8 rat myoblasts) as cultured in the recently designed Space Tissue Loss Flight Module "A" (STL-A). The STL-A is a "state of the art," compact, fully contained, automated cell culture apparatus which replaces a single mid-deck locker on the Space Shuttle. The L8 cells were successfully flown in the STL-A on the Space Shuttle STS-45 mission. Upon return to earth, reculturing of these spaceflown L8 cells (L8SF) resulted in their unexpected failure to fuse and differentiate into myotubes. This inability of the L8SF cells to fuse was found to be a permanent phenotypic alteration. Scanning electron microscopic examination of L8SF cells growing at 1g on fibronectin-coated polypropylene fibers exhibited a strikingly different morphology as compared to control cells. In addition to their failure to fuse into myotubes, L8SF cells also piled up on top of each other. When assayed in fusion-promoting soft agar, L8SF cells gave rise to substantially more and larger colonies than did either preflight (L8AT) or ground control (L8GC) cells. All data to this point indicate that flying L8 rat myoblasts on the Space Shuttle for a duration of 7-10 d at subconfluent densities results in several permanent phenotypic alterations in these cells.
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Sales history. 929.13 Section 929.13 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.13 Sales history. Sales history means the number of barrels of cranberries established for a grower by the committee...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Sales history. 929.13 Section 929.13 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.13 Sales history. Sales history means the number of barrels of cranberries established for a grower by the committee...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Sales history. 929.13 Section 929.13 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.13 Sales history. Sales history means the number of barrels of cranberries established for a grower by the committee...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Sales history. 929.13 Section 929.13 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.13 Sales history. Sales history means the number of barrels of cranberries established for a grower by the committee...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Sales history. 929.13 Section 929.13 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.13 Sales history. Sales history means the number of barrels of cranberries established for a grower by the committee...
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Dynamized Preparations in Cell Culture
Sunila, Ellanzhiyil Surendran; Preethi, Korengath Chandran; Kuttan, Girija
2009-01-01
Although reports on the efficacy of homeopathic medicines in animal models are limited, there are even fewer reports on the in vitro action of these dynamized preparations. We have evaluated the cytotoxic activity of 30C and 200C potencies of ten dynamized medicines against Dalton's Lymphoma Ascites, Ehrlich's Ascites Carcinoma, lung fibroblast (L929) and Chinese Hamster Ovary (CHO) cell lines and compared activity with their mother tinctures during short-term and long-term cell culture. The effect of dynamized medicines to induce apoptosis was also evaluated and we studied how dynamized medicines affected genes expressed during apoptosis. Mother tinctures as well as some dynamized medicines showed significant cytotoxicity to cells during short and long-term incubation. Potentiated alcohol control did not produce any cytotoxicity at concentrations studied. The dynamized medicines were found to inhibit CHO cell colony formation and thymidine uptake in L929 cells and those of Thuja, Hydrastis and Carcinosinum were found to induce apoptosis in DLA cells. Moreover, dynamized Carcinosinum was found to induce the expression of p53 while dynamized Thuja produced characteristic laddering pattern in agarose gel electrophoresis of DNA. These results indicate that dynamized medicines possess cytotoxic as well as apoptosis-inducing properties. PMID:18955237
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Doi, Toshifumi; Ishikawa, Takeshi; Okayama, Tetsuya; Oka, Kaname; Mizushima, Katsura; Yasuda, Tomoyo; Sakamoto, Naoyuki; Katada, Kazuhiro; Kamada, Kazuhiro; Uchiyama, Kazuhiko; Handa, Osamu; Takagi, Tomohisa; Naito, Yuji; Itoh, Yoshito
2017-03-01
Although improvements in the chemotherapy modalities for pancreatic cancer have been realized, pancreatic cancer remains one of the most lethal malignancies. New-generation cancer immunotherapy methods, such as blocking of the PD-1/PD-L1 pathway, are consistently being investigated to improve the survival of pancreatic cancer patients. In the present study, we evaluated the influence of anticancer agents 5-fluorouracil, gemcitabine and paclitaxel on PD-L1 expression in human pancreatic cancer cell lines MIA PaCa-2 and AsPC-1 and in murine pancreatic cancer cell line Pan02. Additionally, we analyzed the molecular mechanisms that facilitated the regulation of PD-L1 expression in these cell lines. We observed that when AsPC-1, MIA PaCa-2 and Pan02 cells were stimulated by 5-fluorouracil, gemcitabine or paclitaxel, PD-L1 surface protein expression was enhanced. Similarly, the mRNA level of PD-L1 was upregulated in the AsPC-1 and Pan02 cells when stimulated by each of the three anticancer agents. The phosphorylation of STAT1 and an increase in total STAT1 were also observed in the AsPC-1 cells when stimulated by each anticancer agent. In response to JAK2 inhibitor treatment, PD-L1 upregulation induced by the anticancer agents was reduced in a dose-dependent manner. These results suggest that i) the JAK2/STAT1 pathway is involved in the anticancer agent-mediated PD-L1 transcription; and ii) the anticancer agents altered the tumor immune response which may induce tumor immune escape. These findings can have an influence on the design of treatments that combine chemotherapy and immunotherapy.
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Vargas-Ramírez, Alba L; Medina-Enríquez, Miriam M; Cordero-Rodríguez, Neira I; Ruiz-Cuello, Tatiana; Aguilar-Faisal, Leopoldo; Trujillo-Ferrara, José G; Alcántara-Farfán, Verónica; Rodríguez-Páez, Lorena
2016-07-01
N-ω-chloroacetyl-L-ornithine (NCAO) is an ornithine decarboxylase (ODC) inhibitor that is known to exert cytotoxic and antiproliferative effects on three neoplastic human cancer cell lines (HeLa, MCF-7, and HepG2). Here, we show that NCAO has antiproliferative activity in 13 cancer cell lines, of diverse tissue origin from human and mice, and in a mouse cancer model in vivo. All cell lines were sensitive to NCAO after 72 h of treatment (the EC50 ranged from 1 to 50.6 µmol/l). The Ca Ski cell line was the most sensitive (EC50=1.18±0.07 µmol/l) and MDA-MB-231 was the least sensitive (EC50=50.6±0.3 µmol/l). This ODC inhibitor showed selectivity for cancer cells, exerting almost no cytotoxic effect on the normal Vero cell line (EC50>1000 µmol/l). NCAO induced apoptosis and inhibited tumor cell migration in vitro. Furthermore, in vivo, this compound (at 50 and 100 mg/kg, daily intraperitoneal injection for 7 days) exerted potent antitumor activity against both solid and ascitic tumors in a mouse model using the myeloma (Ag8) cell line. At these same two doses, the toxicological evaluation showed that NCAO has no obvious systemic toxicity. The current results suggest that the antitumor activity is exerted by apoptosis related not only to a local but also a systemic cytotoxic effect exerted by NCAO on tumor cells. The applications for NCAO as an antitumor agent may be extensive; however, further studies are needed to ascertain the antitumor activity on other types of tumor in vivo and to determine the precise molecular mechanism of its activity.
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Sales history. 929.48 Section 929.48 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.48 Sales history. (a) A sales history for each grower shall be computed by the committee in the following manner: (1) For...
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7 CFR 929.236 - Assessment rate.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Assessment rate. 929.236 Section 929.236 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Assessment Rate § 929.236 Assessment rate. On and after September 1, 2006, an assessment rate of $.28 per barrel is established for cranberries. [72 FR 14654, Mar. 29...
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7 CFR 929.236 - Assessment rate.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Assessment rate. 929.236 Section 929.236 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Assessment Rate § 929.236 Assessment rate. On and after September 1, 2006, an assessment rate of $.28 per barrel is established for cranberries. [72 FR 14654, Mar. 29...
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7 CFR 929.236 - Assessment rate.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Assessment rate. 929.236 Section 929.236 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Assessment Rate § 929.236 Assessment rate. On and after September 1, 2006, an assessment rate of $.28 per barrel is established for cranberries. [72 FR 14654, Mar. 29...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Sales history. 929.48 Section 929.48 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.48 Sales history. (a) A sales history for each grower shall be computed by the committee in the following manner: (1) For...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Sales history. 929.48 Section 929.48 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.48 Sales history. (a) A sales history for each grower shall be computed by the committee in the following manner: (1) For...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Sales history. 929.48 Section 929.48 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.48 Sales history. (a) A sales history for each grower shall be computed by the committee in the following manner: (1) For...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Sales history. 929.48 Section 929.48 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.48 Sales history. (a) A sales history for each grower shall be computed by the committee in the following manner: (1) For...
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7 CFR 929.236 - Assessment rate.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Assessment rate. 929.236 Section 929.236 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Assessment Rate § 929.236 Assessment rate. On and after September 1, 2006, an assessment rate of $.28 per barrel is established for cranberries. [72 FR 14654, Mar. 29...
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7 CFR 929.236 - Assessment rate.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Assessment rate. 929.236 Section 929.236 Agriculture... LONG ISLAND IN THE STATE OF NEW YORK Assessment Rate § 929.236 Assessment rate. On and after September 1, 2006, an assessment rate of $.28 per barrel is established for cranberries. [72 FR 14654, Mar. 29...
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Code of Federal Regulations, 2010 CFR
2010-04-01
... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Sonoma Valley. 9.29 Section 9.29 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS AMERICAN VITICULTURAL AREAS Approved American Viticultural Areas § 9.29 Sonoma Valley. (a) Name. The name of the viticultural...
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Code of Federal Regulations, 2012 CFR
2012-04-01
... 27 Alcohol, Tobacco Products and Firearms 1 2012-04-01 2012-04-01 false Sonoma Valley. 9.29 Section 9.29 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS AMERICAN VITICULTURAL AREAS Approved American Viticultural Areas § 9.29 Sonoma Valley. (a) Name. The name of the viticultural...
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Code of Federal Regulations, 2014 CFR
2014-04-01
... 27 Alcohol, Tobacco Products and Firearms 1 2014-04-01 2014-04-01 false Sonoma Valley. 9.29 Section 9.29 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY ALCOHOL AMERICAN VITICULTURAL AREAS Approved American Viticultural Areas § 9.29 Sonoma Valley. (a) Name. The name of the viticultural...
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Code of Federal Regulations, 2013 CFR
2013-04-01
... 27 Alcohol, Tobacco Products and Firearms 1 2013-04-01 2013-04-01 false Sonoma Valley. 9.29 Section 9.29 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY ALCOHOL AMERICAN VITICULTURAL AREAS Approved American Viticultural Areas § 9.29 Sonoma Valley. (a) Name. The name of the viticultural...
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Code of Federal Regulations, 2011 CFR
2011-04-01
... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Sonoma Valley. 9.29 Section 9.29 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS AMERICAN VITICULTURAL AREAS Approved American Viticultural Areas § 9.29 Sonoma Valley. (a) Name. The name of the viticultural...
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Code of Federal Regulations, 2012 CFR
2012-10-01
... 47 Telecommunication 5 2012-10-01 2012-10-01 false Nameplate. 80.929 Section 80.929 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) SAFETY AND SPECIAL RADIO SERVICES STATIONS IN THE... manufacturer and the type or model number. ...
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de Coupade, Catherine; Solito, Egle; Levine, Jon D
2003-09-01
(1) L-selectin, constitutively expressed by leukocytes, is involved in the initial binding of leukocytes to activated endothelium. Anti-inflammatory drugs like glucocorticoids can induce shedding of L-selectin, but the mechanism is still unknown. Annexin 1, a protein whose synthesis and externalization/secretion are induced during the inflammatory response, has been proposed as a mediator of the anti-inflammatory actions of glucocorticoids. (2) The monocytic cell line U-937 strongly expresses Annexin 1 after 24 h of phorbol 12-myristate 13-acetate (PMA, 1 nm) treatment and externalizes/releases the protein after additional 16 h of dexamethasone (1 microm) treatment. (3) This study investigated the possible regulation of cell surface L-selectin shedding by endogenous Annexin 1, and its role in glucocorticoid-induced L-selectin shedding in the U-937 cell line. (4) PMA- and dexamethasone treatment-induced L-selectin shedding was potentially mediated by Annexin 1, since neutralizing antibodies against Annexin 1 reduced dexamethasone- and Annexin 1-induced shedding. (5) Immunoprecipitation and binding assays provided support for the suggestion that this effect could be mediated by an interaction between externalized Annexin 1 and L-selectin. Such interaction involved the N-terminal domain of Annexin 1 and was calcium-dependent. Confocal microscopy studies demonstrated increased colocalization of Annexin 1 and L-selectin on the cell surface. (6) Overall, our study provides new insights into the potential role of endogenous ANXA1 as a mediator of dexamethasone-induced L-selectin shedding, which may contribute to the anti-inflammatory activity of glucocorticoids.
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Deleterious CHEK2 1100delC and L303X mutants identified among 38 human breast cancer cell lines.
Wasielewski, Marijke; Hanifi-Moghaddam, Pejman; Hollestelle, Antoinette; Merajver, Sofia D; van den Ouweland, Ans; Klijn, Jan G M; Ethier, Stephen P; Schutte, Mieke
2009-01-01
The CHEK2 protein plays a major role in the regulation of DNA damage response pathways. Mutations in the CHEK2 gene, in particular 1100delC, have been associated with increased cancer risks, but the precise function of CHEK2 mutations in carcinogenesis is not known. Human cancer cell lines with CHEK2 mutations are therefore of main interest. Here, we have sequenced 38 breast cancer cell lines for mutations in the CHEK2 gene and identified two cell lines with deleterious CHEK2 mutations. Cell line UACC812 has a nonsense truncating mutation in the CHEK2 kinase domain (L303X) and cell line SUM102PT has the well-known oncogenic CHEK2 1100delC founder mutation. Immunohistochemical analysis revealed that the two CHEK2 mutant cell lines expressed neither CHEK2 nor P-Thr(68) CHEK2 proteins, implying abrogation of normal CHEK2 DNA repair functions. Cell lines UACC812 and SUM102PT thus are the first human CHEK2 null cell lines reported and should therefore be a major help in further unraveling the function of CHEK2 mutations in carcinogenesis.
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Tan, Xiang-Wen; Xia, Hong; Xu, Jin-Hua; Cao, Jian-Guo
2009-05-14
To investigate the effect of 5-allyl-7-gen-difluoromethylenechrysin (ADFMChR) on apoptosis of human liver carcinoma HepG2 cell line and the molecular mechanisms involved. HepG2 cells and L-02 cells were cultured in vitro and the inhibitory effect of ADFMChR on their proliferation was measured by MTT assay. The apoptosis of HepG2 cells was determined by flow cytometry (FCM) using propidium iodide (PI) fluorescence staining. DNA ladder bands were observed by DNA agarose gel electrophoresis. The influence of ADFMChR on the proxisome proliferator-activated receptor gamma (PPARgamma), NF-kappaB, Bcl-2 and Bax protein expression of HepG2 cells were analyzed by Western blotting. MTT assay showed that ADFMChR significantly inhibited proliferation of HepG2 cells in a dose-dependent manner, with little effect on growth of L-02 cells, and when IC(50) was measured as 8.45 micromol/L and 191.55 micromol/L respectively, the potency of ADFMChR to HepG2 cells, was found to be similar to 5-fluorouracil (5-FU, IC(50) was 9.27 micromol/L). The selective index of ADFMChR cytotoxicity to HepG2 cells was 22.67 (191.55/8.45), higher than 5-FU (SI was 7.05 (65.37/9.27). FCM with PI staining demonstrated that the apoptosis rates of HepG2 cells treated with 3.0, 10.0 and 30.0 micromol/L ADFMChR for 48 h were 5.79%, 9.29% and 37.8%, respectively, and were significantly higher when treated with 30.0 micromol/L ADFMChR than when treated with 30.0 micromol/L ChR (16.0%) (P < 0.05) and were similar to those obtained with 30.0 micromol/L 5-FU (41.0%). DNA agarose gel electrophoresis showed that treatment of HepG2 cells with 10.0 micromol/L ADFMChR for 48 h and 72 h resulted in typical DNA ladders which could be reversed by 10.00 micromol/L GW9662, a blocker of PPARgamma. Western blotting analysis revealed that after 24 h of treatment with 3.0, 10.0, 30.0 micromol/L ADFMChR, PPARgamma and Bax protein expression in HepG2 cells increased but Bcl-2 and NF-kappaB expression decreased; however, pre
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Vacancies. 929.26 Section 929.26 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Vacancies. 929.26 Section 929.26 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Separability. 929.76 Section 929.76 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Acceptance. 929.25 Section 929.25 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Acceptance. 929.25 Section 929.25 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Act. 929.2 Section 929.2 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Agents. 929.73 Section 929.73 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Separability. 929.76 Section 929.76 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Act. 929.2 Section 929.2 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Act. 929.2 Section 929.2 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Derogation. 929.74 Section 929.74 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Vacancies. 929.26 Section 929.26 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Vacancies. 929.26 Section 929.26 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Person. 929.3 Section 929.3 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Derogation. 929.74 Section 929.74 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Procedure. 929.32 Section 929.32 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Derogation. 929.74 Section 929.74 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Accounting. 929.42 Section 929.42 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Acceptance. 929.25 Section 929.25 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Agents. 929.73 Section 929.73 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Powers. 929.30 Section 929.30 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Derogation. 929.74 Section 929.74 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Powers. 929.30 Section 929.30 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Procedure. 929.32 Section 929.32 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Expenses. 929.40 Section 929.40 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Person. 929.3 Section 929.3 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Person. 929.3 Section 929.3 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Separability. 929.76 Section 929.76 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Expenses. 929.40 Section 929.40 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Act. 929.2 Section 929.2 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Accounting. 929.42 Section 929.42 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Procedure. 929.32 Section 929.32 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Acceptance. 929.25 Section 929.25 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Accounting. 929.42 Section 929.42 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Procedure. 929.32 Section 929.32 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Person. 929.3 Section 929.3 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Agents. 929.73 Section 929.73 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Expenses. 929.40 Section 929.40 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Acceptance. 929.25 Section 929.25 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Expenses. 929.40 Section 929.40 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Act. 929.2 Section 929.2 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Derogation. 929.74 Section 929.74 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Accounting. 929.42 Section 929.42 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Separability. 929.76 Section 929.76 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Agents. 929.73 Section 929.73 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Person. 929.3 Section 929.3 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Separability. 929.76 Section 929.76 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Expenses. 929.40 Section 929.40 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
-
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Agents. 929.73 Section 929.73 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Powers. 929.30 Section 929.30 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Powers. 929.30 Section 929.30 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS...
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7 CFR 929.28 - Redistricting and Reapportionment.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Redistricting and Reapportionment. 929.28 Section 929.28 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING... Body § 929.28 Redistricting and Reapportionment. (a) The committee, with the approval of the Secretary...
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7 CFR 929.28 - Redistricting and Reapportionment.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Redistricting and Reapportionment. 929.28 Section 929.28 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING... Body § 929.28 Redistricting and Reapportionment. (a) The committee, with the approval of the Secretary...
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7 CFR 929.45 - Research and development.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Research and development. 929.45 Section 929.45... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Research § 929.45 Research and... establishment of production research, marketing research, and market development projects, including paid...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Accounting. 929.42 Section 929.42 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... Accounting. (a) If, at the end of a fiscal period, the assessments collected are in excess of expenses...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Compliance. 929.66 Section 929.66 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... Compliance. Except as provided in this part, no person shall handle cranberries, the handling of which has...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Compliance. 929.66 Section 929.66 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... Compliance. Except as provided in this part, no person shall handle cranberries, the handling of which has...
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7 CFR 929.51 - Recommendations for regulation.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Recommendations for regulation. 929.51 Section 929.51... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.51 Recommendations for... August 31. Such recommendation shall include the free and restricted percentages for the crop year; (3...
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7 CFR 929.51 - Recommendations for regulation.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Recommendations for regulation. 929.51 Section 929.51... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.51 Recommendations for... August 31. Such recommendation shall include the free and restricted percentages for the crop year; (3...
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7 CFR 929.125 - Committee review procedures.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Committee review procedures. 929.125 Section 929.125... LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.125 Committee review procedures. Growers may request, and the Committee may grant, a review of determinations made by the Committee...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... assessments on all cranberries acquired as the first handler thereof during such period, except as provided in § 929.55: Provided, That no handler shall pay assessments on excess cranberries as provided in § 929.57...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... assessments on all cranberries acquired as the first handler thereof during such period, except as provided in § 929.55: Provided, That no handler shall pay assessments on excess cranberries as provided in § 929.57...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... assessments on all cranberries acquired as the first handler thereof during such period, except as provided in § 929.55: Provided, That no handler shall pay assessments on excess cranberries as provided in § 929.57...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... assessments on all cranberries acquired as the first handler thereof during such period, except as provided in § 929.55: Provided, That no handler shall pay assessments on excess cranberries as provided in § 929.57...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... assessments on all cranberries acquired as the first handler thereof during such period, except as provided in § 929.55: Provided, That no handler shall pay assessments on excess cranberries as provided in § 929.57...
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LINE-1 Cultured Cell Retrotransposition Assay
Kopera, Huira C.; Larson, Peter A.; Moldovan, John B.; Richardson, Sandra R.; Liu, Ying; Moran, John V.
2016-01-01
Summary The Long INterspersed Element-1 (LINE-1 or L1) retrotransposition assay has facilitated the discovery and characterization of active (i.e., retrotransposition-competent) LINE-1 sequences from mammalian genomes. In this assay, an engineered LINE-1 containing a retrotransposition reporter cassette is transiently transfected into a cultured cell line. Expression of the reporter cassette, which occurs only after a successful round of retrotransposition, allows the detection and quantification of the LINE-1 retrotransposition efficiency. This assay has yielded insight into the mechanism of LINE-1 retrotransposition. It also has provided a greater understanding of how the cell regulates LINE-1 retrotransposition and how LINE-1 retrotransposition impacts the structure of mammalian genomes. Below, we provide a brief introduction to LINE-1 biology and then detail how the LINE-1 retrotransposition assay is performed in cultured mammalian cells. PMID:26895052
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Lee, Kyeong-Eun; Kang, Young-Sook
2017-05-10
L -Citrulline is a neutral amino acid and a major precursor of L -arginine in the nitric oxide (NO) cycle. Recently it has been reported that L -citrulline prevents neuronal cell death and protects cerebrovascular injury, therefore, L -citrulline may have a neuroprotective effect to improve cerebrovascular dysfunction. Therefore, we aimed to clarify the brain transport mechanism of L -citrulline through blood-brain barrier (BBB) using the conditionally immortalized rat brain capillary endothelial cell line (TR-BBB cells), as an in vitro model of the BBB. The uptake study of [ 14 C] L-citrulline, quantitative real-time polymerase chain reaction (PCR) analysis, and rLAT1, system b 0,+ , and CAT1 small interfering RNA study were performed in TR-BBB cells. The uptake of [ 14 C] L -citrulline was a time-dependent, but ion-independent manner in TR-BBB cells. The transport process involved two saturable components with a Michaelis-Menten constant of 30.9 ± 1.0 μM (Km 1 ) and 1.69 ± 0.43 mM (Km 2 ). The uptake of [ 14 C] L -citrulline in TR-BBB cells was significantly inhibited by neutral and cationic amino acids, but not by anionic amino acids. In addition, [ 14 C] L -citrulline uptake in the cells was markedly inhibited by 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), which is the inhibitor of the large neutral amino acid transporter 1 (LAT1), B 0 , B 0,+ and harmaline, the inhibitor of system b 0,+ . Gabapentin and L -dopa as the substrates of LAT1 competitively inhibited the uptake of [ 14 C] L -citrulline. IC 50 values for L -dopa, gabapentin, L -phenylalanine and L -arginine were 501 μM, 223 μM, 68.9 μM and 33.4 mM, respectively. The expression of mRNA for LAT1 was predominantly increased 187-fold in comparison with that of system b 0,+ in TR-BBB cells. In the studies of LAT1, system b 0,+ and CAT1 knockdown via siRNA transfection into TR-BBB cells, the transcript level of LAT1 and [ 14 C] L -citrulline uptake by LAT1 siRNA were
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Arsenakis, M; Tomasi, L F; Speziali, V; Roizman, B; Campadelli-Fiume, G
1986-01-01
Ltk- cells were transfected with a plasmid containing the entire domain of glycoprotein C (gC), a true gamma or gamma 2 gene of herpes simplex virus 1 (HSV-1) and the methotrexate-resistant mouse dihydrofolate reductase mutant gene. The resulting methotrexate-resistant cell line was cloned; of the 39 clonal lines tested only 1, L3153(28), expressed gC after infection with HSV-1(MP), a gC- mutant, and none expressed gC constitutively. The induction of gC was optimal at multiplicities ranging between 0.5 and 2 PFU per cell, and the quantities produced were equivalent to or higher than those made by methotrexate-resistant gC- L cells infected with wild-type (gC+) virus. The gC gene resident in the L3153(28) cells was regulated as a beta gene inasmuch as the amounts of gC made in infected L3153(28) cells exposed to concentrations of phosphonoacetate that inhibited viral DNA synthesis were higher than those made in the absence of the drug, gC was induced at both permissive and nonpermissive temperatures by the DNA- mutant tsHA1 carrying a lesion in the gene specifying the major DNA-binding protein and which does not express gamma 2 genes at the nonpermissive temperature, and gC was induced only at the permissive temperature in cells infected with ts502 containing a mutation in the alpha 4 gene. The gC induced in L3153(28) cells was made earlier and processed faster to the mature form than that induced in a gC- clone of methotrexate-resistant cells infected with wild-type virus. Unlike virus stocks made in gC- cells, HSV-1(MP) made in L3153(28) cells was susceptible to neutralization by anti-gC monoclonal antibody. Images PMID:3009854
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Salazar-Aguilar, Sandra; Ruiz-Posadas, Lucero del Mar; Cadena-Iñiguez, Jorge; Santiago-Osorio, Edelmiro; Aguiñiga-Sánchez, Itzen; Rivera-Martínez, Ana Rocío; Aguirre-Medina, Juan Francisco
2017-01-01
The Sechium edule Perla Negra cultivar is a recently-obtained biological material whose progenitors are S. edule var. nigrum minor and S. edule var. amarus silvestrys, the latter of which has been reported to have antiproliferative activity against the HeLa P-388 and L-929 cancer cell lines. The present study aimed to determine if the methanolic extract of the fruit of the Perla Negra cultivar had the same biological activity. The methanolic extract was phytochemically characterized by thin layer chromatography (TLC) and column chromatography (CC), identifying the terpenes and flavonoids. The compounds identified via high performance liquid chromatography (HPLC) were Cucurbitacins B, D, E, and I for the terpene fractions, and Rutin, Phlorizidin, Myricetin, Quercetin, Naringenin, Phloretin, Apigenin, and Galangin for the flavonoid fractions). Biological activity was evaluated with different concentrations of the methanolic extract in the HeLa cell line and normal lymphocytes. The methanolic extract inhibited the proliferation of HeLa cells (IC50 1.85 µg·mL−1), but the lymphocytes were affected by the extract (IC50 30.04 µg·mL−1). Some fractions, and the pool of all of them, showed inhibition higher than 80% at a concentration of 2.11 µg·mL−1. Therefore, the biological effect shown by the methanolic extract of the Perla Negra has some specificity in inhibiting tumor cells and not normal cells; an unusual feature among molecules investigated as potential biomedical agents. PMID:28757593
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Kagami, Yoshitoyo; Uchiyama, Susumu; Kato, Harumi; Okada, Yasutaka; Seto, Masao; Kinoshita, Tomohiro
2017-07-05
Growing adult T-cell leukemia/lymphoma (ATLL) cells in vitro is difficult. Here, we examined the effects of static electricity in the culture medium on the proliferation of ATLL cells. Six out of 10 ATLL cells did not proliferate in vitro and thus had to be cultured in a medium containing negatively charged polymers. In the presence of poly-γ-glutamic acid (PGA) or chondroitin sulfate (CDR), cell lines (HKOX3-PGA, HKOX3-CDR) were established from the same single ATLL case using interleukin (IL)-2, IL-4, and feeder cells expressing OX40L (OX40L + HK). Dextran sulfate inhibited growth in both HKOX3 cell lines. Both PGA and OX40L + HK were indispensable for HKOX3-PGA growth, but HKOX3-CDR could proliferate in the presence of CDR or OX40L + HK alone. Thus, the specific action of each negatively charged polymer promoted the growth of specific ATLL cells in vitro.
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Cytoglobin inhibits migration through PI3K/AKT/mTOR pathway in fibroblast cells.
Demirci, Selami; Doğan, Ayşegül; Apdik, Hüseyin; Tuysuz, Emre Can; Gulluoglu, Sukru; Bayrak, Omer Faruk; Şahin, Fikrettin
2018-01-01
Cell proliferation and migration are crucial in many physiological processes including development, cancer, tissue repair, and wound healing. Cell migration is regulated by several signaling molecules. Identification of genes related to cell migration is required to understand molecular mechanism of non-healing chronic wounds which is a major concern in clinics. In the current study, the role of cytoglobin (CYGB) gene in fıbroblast cell migration and proliferation was described. L929 mouse fibroblast cells were transduced with lentiviral particles for CYGB and GFP, and analyzed for cell proliferation and migration ability. Fibroblast cells overexpressing CYGB displayed decreased cell proliferation, colony formation capacity, and cell migration. Phosphorylation levels of mTOR and two downstream effectors S6 and 4E-BP1 which take part in PI3K/AKT/mTOR signaling declined in CYGB-overexpressing cells. Microarray analysis indicated that CYGB overexpression leads to downregulation of cell proliferation, migration, and tumor growth associated genes in L929 cell line. This study demonstrated the role of CYGB in fibroblast cell motility and proliferation. CYGB could be a promising candidate for further studies as a potential target for diseases related to cell migration such as cancer and chronic wound treatment.
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7 CFR 929.4 - Production area.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Production area. 929.4 Section 929.4 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.65 - Confidential information.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Confidential information. 929.65 Section 929.65 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.75 - Personal liability.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Personal liability. 929.75 Section 929.75 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Fiscal period. 929.6 Section 929.6 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.106 - Fiscal period.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Fiscal period. 929.106 Section 929.106 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.27 - Alternate members.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Alternate members. 929.27 Section 929.27 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.4 - Production area.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Production area. 929.4 Section 929.4 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.27 - Alternate members.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Alternate members. 929.27 Section 929.27 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.4 - Production area.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Production area. 929.4 Section 929.4 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.4 - Production area.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Production area. 929.4 Section 929.4 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.4 - Production area.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Production area. 929.4 Section 929.4 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.65 - Confidential information.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Confidential information. 929.65 Section 929.65 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.106 - Fiscal period.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Fiscal period. 929.106 Section 929.106 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Fiscal period. 929.6 Section 929.6 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.75 - Personal liability.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Personal liability. 929.75 Section 929.75 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.106 - Fiscal period.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Fiscal period. 929.106 Section 929.106 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.75 - Personal liability.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Personal liability. 929.75 Section 929.75 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.27 - Alternate members.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Alternate members. 929.27 Section 929.27 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.65 - Confidential information.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Confidential information. 929.65 Section 929.65 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.27 - Alternate members.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Alternate members. 929.27 Section 929.27 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.75 - Personal liability.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Personal liability. 929.75 Section 929.75 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.27 - Alternate members.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Alternate members. 929.27 Section 929.27 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.106 - Fiscal period.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Fiscal period. 929.106 Section 929.106 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.65 - Confidential information.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Confidential information. 929.65 Section 929.65 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Fiscal period. 929.6 Section 929.6 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.57 - Outlets for restricted cranberries.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Outlets for restricted cranberries. 929.57 Section 929... SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES....57 Outlets for restricted cranberries. (a) Except as provided in this section and in § 929.56...
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7 CFR 929.57 - Outlets for restricted cranberries.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Outlets for restricted cranberries. 929.57 Section 929... SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES....57 Outlets for restricted cranberries. (a) Except as provided in this section and in § 929.56...
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7 CFR 929.57 - Outlets for restricted cranberries.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Outlets for restricted cranberries. 929.57 Section 929... SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES....57 Outlets for restricted cranberries. (a) Except as provided in this section and in § 929.56...
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7 CFR 929.57 - Outlets for restricted cranberries.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Outlets for restricted cranberries. 929.57 Section 929... SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES....57 Outlets for restricted cranberries. (a) Except as provided in this section and in § 929.56...
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7 CFR 929.57 - Outlets for restricted cranberries.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Outlets for restricted cranberries. 929.57 Section 929... SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES....57 Outlets for restricted cranberries. (a) Except as provided in this section and in § 929.56...
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Gardell, Alison M.; Qin, Qin; Rice, Robert H.; Li, Johnathan; Kültz, Dietmar
2014-01-01
Fish cell cultures are becoming more widely used models for investigating molecular mechanisms of physiological response to environmental challenge. In this study, we derived two immortalized Mozambique tilapia (Oreochromis mossambicus) cell lines from brain (OmB) and lip epithelium (OmL), and compared them to a previously immortalized bulbus arteriosus (TmB) cell line. The OmB and OmL cell lines were generated without or with Rho-associated kinase (ROCK) inhibitor/3T3 feeder layer supplementation. Although both approaches were successful, ROCK inhibitor/feeder layer supplementation was found to offer the advantages of selecting for epithelial-like cell type and decreasing time to immortalization. After immortalization (≥ passage 5), we characterized the proteomes of the newly derived cell lines (OmB and OmL) using LCMS and identified several unique cell markers for each line. Subsequently, osmotolerance for each of the three cell lines following acute exposure to elevated sodium chloride was evaluated. The acute maximum osmotolerance of these tilapia cell lines (>700 mOsm/kg) was markedly higher than that of any other known vertebrate cell line, but was significantly higher in the epithelial-like OmL cell line. To validate the physiological relevance of these tilapia cell lines, we quantified the effects of acute hyperosmotic challenge (450 mOsm/kg and 700 mOsm/kg) on the transcriptional regulation of two enzymes involved in biosynthesis of the compatible organic osmolyte, myo-inositol. Both enzymes were found to be robustly upregulated in all three tilapia cell lines. Therefore, the newly established tilapia cells lines represent valuable tools for studying molecular mechanisms involved in the osmotic stress response of euryhaline fish. PMID:24797371
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7 CFR 929.104 - Outlets for excess cranberries.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Outlets for excess cranberries. 929.104 Section 929... SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES... excess cranberries. (a) In accordance with § 929.61, excess cranberries may be disposed of only in the...
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7 CFR 929.104 - Outlets for excess cranberries.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Outlets for excess cranberries. 929.104 Section 929... SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES... excess cranberries. (a) In accordance with § 929.61, excess cranberries may be disposed of only in the...
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7 CFR 929.104 - Outlets for excess cranberries.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Outlets for excess cranberries. 929.104 Section 929... SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES... excess cranberries. (a) In accordance with § 929.61, excess cranberries may be disposed of only in the...
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7 CFR 929.104 - Outlets for excess cranberries.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Outlets for excess cranberries. 929.104 Section 929... SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES... excess cranberries. (a) In accordance with § 929.61, excess cranberries may be disposed of only in the...
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7 CFR 929.75 - Personal liability.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Personal liability. 929.75 Section 929.75 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... committee shall be held personally responsible, either individually or jointly with others, in any way...
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Label-free ferrohydrodynamic cell separation of circulating tumor cells.
Zhao, Wujun; Cheng, Rui; Jenkins, Brittany D; Zhu, Taotao; Okonkwo, Nneoma E; Jones, Courtney E; Davis, Melissa B; Kavuri, Sravan K; Hao, Zhonglin; Schroeder, Carsten; Mao, Leidong
2017-09-12
Circulating tumor cells (CTCs) have significant implications in both basic cancer research and clinical applications. To address the limited availability of viable CTCs for fundamental and clinical investigations, effective separation of extremely rare CTCs from blood is critical. Ferrohydrodynamic cell separation (FCS), a label-free method that conducted cell sorting based on cell size difference in biocompatible ferrofluids, has thus far not been able to enrich low-concentration CTCs from cancer patients' blood because of technical challenges associated with processing clinical samples. In this study, we demonstrated the development of a laminar-flow microfluidic FCS device that was capable of enriching rare CTCs from patients' blood in a biocompatible manner with a high throughput (6 mL h -1 ) and a high rate of recovery (92.9%). Systematic optimization of the FCS devices through a validated analytical model was performed to determine optimal magnetic field and its gradient, ferrofluid properties, and cell throughput that could process clinically relevant amount of blood. We first validated the capability of the FCS devices by successfully separating low-concentration (∼100 cells per mL) cancer cells using six cultured cell lines from undiluted white blood cells (WBCs), with an average 92.9% cancer cell recovery rate and an average 11.7% purity of separated cancer cells, at a throughput of 6 mL per hour. Specifically, at ∼100 cancer cells per mL spike ratio, the recovery rates of cancer cells were 92.3 ± 3.6% (H1299 lung cancer), 88.3 ± 5.5% (A549 lung cancer), 93.7 ± 5.5% (H3122 lung cancer), 95.3 ± 6.0% (PC-3 prostate cancer), 94.7 ± 4.0% (MCF-7 breast cancer), and 93.0 ± 5.3% (HCC1806 breast cancer), and the corresponding purities of separated cancer cells were 11.1 ± 1.2% (H1299 lung cancer), 10.1 ± 1.7% (A549 lung cancer), 12.1 ± 2.1% (H3122 lung cancer), 12.8 ± 1.6% (PC-3 prostate cancer), 11.9 ± 1.8% (MCF-7 breast cancer), and 12.2 ± 1
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The inhibitory effects of fluoroquinolones on L-carnitine transport in placental cell line BeWo.
Hirano, Takeshi; Yasuda, Satoru; Osaka, Yuki; Asari, Masaru; Kobayashi, Masaki; Itagaki, Shirou; Iseki, Ken
2008-03-03
L-Carnitine plays an important role in lipid metabolism by facilitating the transport of long-chain fatty acids across the mitochondrial inner membrane followed by fatty acid beta-oxidation. It is known that members of the OCTN family play an important role in L-carnitine transport in the placenta. Investigation of drug-drug or drug-nutrient interaction in the placenta is important for establishment of safety drug medication during pregnancy. The aim of this study was to determine the effects of fluoroquinolones, inhibitors of OCTN2, on L-carnitine transport in the placenta which is known to have a high expression level of OCTN2. We investigated the inhibitory effect of five fluoroquinolones, ciprofloxacin (CPFX), gatifloxacin (GFLX), ofloxacin (OFLX), levofloxacin (LVFX) and grepafloxacin (GPFX), on L-carnitine transport mediated by OCTN2 in placental cell line BeWo cells. We found that all of the fluoroquinolones inhibited L-carnitine transport, GPFX being the strongest inhibitor. We also found that the inhibitory effects of LVFX and GPFX depended on their existence ratio of zwitterionic forms as, we reported previously. Furthermore, we elucidated the LVFX transport mechanism in BeWo cells. LVFX was transported actively by transporters. However, we found that LVFX transport was Na+-independent and l-carnitine had no inhibitory effect on LVFX transport, suggesting that LVFX acts as inhibitor of OCTN2, not as a substrate for OCTN2.
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Mohammadi, A; Mansoori, B; Goldar, S; Shanehbandi, D; Khaze, V; Mohammadnejad, L; Baghbani, E; Baradaran, B
2016-02-29
Breast cancer is the most common cancer among women in worldwide, especially in developing countries. Therefore, a large number of anticancer agents with herbal origins have been reported against this deadly disease. This study is the first to examine the cytotoxic and apoptotic effects of Urtica dioica in MDA-MB-468, human breast adenocarcinoma cells. The 3-(4,5-dimethylethiazol-2 yl)-2,5- diphenyltetrazolium (MTT) reduction and trypan-blue exclusion assay were performed in MDA-MB-468 cells as well as control cell line L929 to analyze the cytotoxic activity of the dichloromethane extract. In addition, Apoptosis induction of Urtica dioica on the MDA-MB-468 cells was assessed using TUNEL (terminal deoxy transferase (TdT)-mediated dUTP nick- end labeling) assay and DNA fragmentation analysis and real-time polymerase chain reaction (PCR). The results showed that the extract significantly inhibited cell growth and viability without inducing damage to normal control cells. Nuclei Staining in TUNEL and DNA fragments in DNA fragmentation assay and increase in the mRNA expression levels of caspase-3, caspase-9, decrease in the bcl2 and no significant change in the caspase-8 mRNA expression level, showed that the induction of apoptosis was the main mechanism of cell death that induce by Urtica dioica extract. Our results suggest that urtica dioica dichloromethane extract may contain potential bioactive compound(s) for the treatment of breast adenocarcinoma.
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10 CFR 9.29 - Appeal from initial determination.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 10 Energy 1 2013-01-01 2013-01-01 false Appeal from initial determination. 9.29 Section 9.29 Energy NUCLEAR REGULATORY COMMISSION PUBLIC RECORDS Freedom of Information Act Regulations § 9.29 Appeal... determination on the appeal within 20 working days after the receipt of the appeal. If the Inspector General...
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10 CFR 9.29 - Appeal from initial determination.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 10 Energy 1 2012-01-01 2012-01-01 false Appeal from initial determination. 9.29 Section 9.29 Energy NUCLEAR REGULATORY COMMISSION PUBLIC RECORDS Freedom of Information Act Regulations § 9.29 Appeal... determination on the appeal within 20 working days after the receipt of the appeal. If the Inspector General...
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10 CFR 9.29 - Appeal from initial determination.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 10 Energy 1 2011-01-01 2011-01-01 false Appeal from initial determination. 9.29 Section 9.29 Energy NUCLEAR REGULATORY COMMISSION PUBLIC RECORDS Freedom of Information Act Regulations § 9.29 Appeal... determination on the appeal within 20 working days after the receipt of the appeal. If the Inspector General...
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Gregorczyk, Karolina P.; Wyżewski, Zbigniew; Szczepanowska, Joanna; Mielcarska, Matylda B.; Bossowska-Nowicka, Magdalena; Gieryńska, Małgorzata; Boratyńska-Jasińska, Anna; Niemiałtowski, Marek G.
2018-01-01
Mitochondria are multifunctional organelles that participate in numerous processes in response to viral infection, but they are also a target for viruses. The aim of this study was to define subcellular events leading to alterations in mitochondrial morphology and function during infection with ectromelia virus (ECTV). We used two different cell lines and a combination of immunofluorescence techniques, confocal and electron microscopy, and flow cytometry to address subcellular changes following infection. Early in infection of L929 fibroblasts and RAW 264.7 macrophages, mitochondria gathered around viral factories. Later, the mitochondrial network became fragmented, forming punctate mitochondria that co-localized with the progeny virions. ECTV-co-localized mitochondria associated with the cytoskeleton components. Mitochondrial membrane potential, mitochondrial fission–fusion, mitochondrial mass, and generation of reactive oxygen species (ROS) were severely altered later in ECTV infection leading to damage of mitochondria. These results suggest an important role of mitochondria in supplying energy for virus replication and morphogenesis. Presumably, mitochondria participate in transport of viral particles inside and outside of the cell and/or they are a source of membranes for viral envelope formation. We speculate that the observed changes in the mitochondrial network organization and physiology in ECTV-infected cells provide suitable conditions for viral replication and morphogenesis. PMID:29772718
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Gregorczyk, Karolina P; Wyżewski, Zbigniew; Szczepanowska, Joanna; Toka, Felix N; Mielcarska, Matylda B; Bossowska-Nowicka, Magdalena; Gieryńska, Małgorzata; Boratyńska-Jasińska, Anna; Struzik, Justyna; Niemiałtowski, Marek G; Szulc-Dąbrowska, Lidia
2018-05-16
Mitochondria are multifunctional organelles that participate in numerous processes in response to viral infection, but they are also a target for viruses. The aim of this study was to define subcellular events leading to alterations in mitochondrial morphology and function during infection with ectromelia virus (ECTV). We used two different cell lines and a combination of immunofluorescence techniques, confocal and electron microscopy, and flow cytometry to address subcellular changes following infection. Early in infection of L929 fibroblasts and RAW 264.7 macrophages, mitochondria gathered around viral factories. Later, the mitochondrial network became fragmented, forming punctate mitochondria that co-localized with the progeny virions. ECTV-co-localized mitochondria associated with the cytoskeleton components. Mitochondrial membrane potential, mitochondrial fission⁻fusion, mitochondrial mass, and generation of reactive oxygen species (ROS) were severely altered later in ECTV infection leading to damage of mitochondria. These results suggest an important role of mitochondria in supplying energy for virus replication and morphogenesis. Presumably, mitochondria participate in transport of viral particles inside and outside of the cell and/or they are a source of membranes for viral envelope formation. We speculate that the observed changes in the mitochondrial network organization and physiology in ECTV-infected cells provide suitable conditions for viral replication and morphogenesis.
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Harada, Kazuki; Sada, Shoko; Sakaguchi, Hidekazu; Takizawa, Mai; Ishida, Rika; Tsuboi, Takashi
2018-07-02
S-equol is one of gut bacterial metabolites produced from soybean isoflavone daizein. While S-equol is known to promote glucose-induced insulin secretion from pancreatic β cells, whether S-equol affects glucagon-like peptide-1 (GLP-1) secretion from enteroendoceine L cells remains unclear. Here we assessed the effect of S-equol on GLP-1 secretion from mouse enteroendocrine L cell line GLUTag cells. GLUTag cells expressed GPR30 and estrogen receptors, which are putative S-equol receptors. Application of S-equol induced an increase in intracellular Ca 2+ levels via GPR30. However, S-equol did not enhance GLP-1 exocytosis, and long-term treatment of S-equol suppressed GLP-1 secretion. Moreover, immunocytochemistry revealed that S-equol increased the density of cortical actin filaments via G 12/13 signaling under GPR30. These data suggest that S-equol prevents GLP-1 secretion as a result of competing regulation between Ca 2+ mobilization and actin reorganization. Copyright © 2018 Elsevier Inc. All rights reserved.
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7 CFR 929.55 - Interhandler transfer.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Interhandler transfer. 929.55 Section 929.55 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing... form provided by the committee four times a year or at other such times as may be recommended by the...
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7 CFR 929.65 - Confidential information.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Confidential information. 929.65 Section 929.65... Confidential information. All reports and records furnished or submitted by handlers to the committee and its authorized agents which include data or information constituting a trade secret or disclosing the trade...
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Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S
2009-10-29
Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding alphavbeta5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain.
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Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S
2009-01-01
Background Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. Results L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding αvβ5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Conclusion Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain. PMID:19874583
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Shirasaki, Hideaki; Kanaizumi, Etsuko; Himi, Tetsuo
2017-11-01
Numerous reports have shown that cysteinyl leukotrienes (CysLTs) contribute to tissue accumulation of eosinophils in allergic airway inflammation. To date, only a few studies have reported that CysLTs promote chemotactic activity of human eosinophils in vitro. The purpose of this study was to investigate whether CysLTs promote chemotaxis in the human eosinophilic cell line, EoL-1. EoL-1 cells were induced to differentiate into mature eosinophil-like cells via incubation with butyric acid and cytokines (IL-3, IL-5 and GM-CSF). The chemotactic activity of the differentiated EoL-1 cells was assessed using the commercial cell migration assay kit. LTD 4 elicited dose-related chemotactic activity in the differntiated EoL-1 cells in the range of 1-100 nM. A typical bell-shaped dose-response curve was observed with optimal activity at 10 nM. The chemotactic activity elicited by LTD 4 (10 nM) was significantly inhibited by montelukast (control, 345 ± 19.2 × 10 3 RFU; LTD 4 10 nM alone, 511 ± 39.2 × 10 3 RFU; LTD 4 10 nM plus montelukast 100 nM, 387 ± 28.2 × 10 3 RFU). LTD 4 induces migration in eosinophilic cells via activation of CysLT1 receptor. The present in vitro model may be useful for elucidation of the mechanism underlying CysLT-induced tissue eosinophilia.
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7 CFR 929.68 - Effective time.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Effective time. 929.68 Section 929.68 Agriculture... Effective time. The provisions of this part, and of any amendment thereto, shall become effective at such time as the Secretary may declare above his signature and shall continue in force until terminated in...
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7 CFR 929.68 - Effective time.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Effective time. 929.68 Section 929.68 Agriculture... Effective time. The provisions of this part, and of any amendment thereto, shall become effective at such time as the Secretary may declare above his signature and shall continue in force until terminated in...
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7 CFR 929.68 - Effective time.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Effective time. 929.68 Section 929.68 Agriculture... Effective time. The provisions of this part, and of any amendment thereto, shall become effective at such time as the Secretary may declare above his signature and shall continue in force until terminated in...
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7 CFR 929.68 - Effective time.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Effective time. 929.68 Section 929.68 Agriculture... Effective time. The provisions of this part, and of any amendment thereto, shall become effective at such time as the Secretary may declare above his signature and shall continue in force until terminated in...
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7 CFR 929.68 - Effective time.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Effective time. 929.68 Section 929.68 Agriculture... Effective time. The provisions of this part, and of any amendment thereto, shall become effective at such time as the Secretary may declare above his signature and shall continue in force until terminated in...
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7 CFR 929.55 - Interhandler transfer.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Interhandler transfer. 929.55 Section 929.55... transfer. (a) Transfer of cranberries from one handler to another may be made without prior notice to the committee, except during a period when a volume regulation has been established. If such transfer is made...
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Vaarala, M H; Porvari, K S; Kyllönen, A P; Mustonen, M V; Lukkarinen, O; Vihko, P T
1998-09-25
A cDNA library specific for mRNA over-expressed in prostate cancer was generated by subtractive hybridization of transcripts originating from prostatic hyperplasia and cancer tissues. cDNA encoding ribosomal proteins L4, L5, L7a, L23a, L30, L37, S14 and S18 was found to be present among 100 analyzed clones. Levels of ribosomal mRNA were significantly higher at least in one of the prostate-cancer cell lines, LNCaP, DU-145 and PC-3, than in hyperplastic tissue, as determined by slot-blot hybridization. Furthermore, L23a- and S14-transcript levels were significantly elevated in PC-3 cells as compared with those in the normal prostate epithelial cell line PrEC. Generally, dramatic changes in the mRNA content of the ribosomal proteins were not detected, the most evident over-expression being that of L37 mRNA, which was 3.4 times more abundant in LNCaP cells than in hyperplastic prostate tissue. The over-expression of L7a and L37 mRNA was confirmed in prostate-cancer tissue samples by in situ hybridization. Elevated cancer-related expression of L4 and L30 has not been reported, but levels of the other ribosomal proteins are known to be increased in several types of cancers. These results therefore suggest that prostate cancer is comparable with other types of cancers, in that a larger pool of some ribosomal proteins is gained during the transformation process, by an unknown mechanism.
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NASA Technical Reports Server (NTRS)
Kraemer, S. M.; Kronenberg, A.; Ueno, A.; Waldren, C. A.; Chatterjee, A. (Principal Investigator)
2000-01-01
Astronauts can be exposed to charged particles, including protons, alpha particles and heavier ions, during space flights. Therefore, studying the biological effectiveness of these sparsely and densely ionizing radiations is important to understanding the potential health effects for astronauts. We evaluated the mutagenic effectiveness of sparsely ionizing 55 MeV protons and densely ionizing 32 MeV/nucleon nitrogen ions using cells of two human-hamster cell lines, A(L) and A(L)C. We have previously characterized a spectrum of mutations, including megabase deletions, in human chromosome 11, the sole human chromosome in the human-hamster hybrid cell lines A(L)C and A(L). CD59(-) mutants have lost expression of a human cell surface antigen encoded by the CD59 gene located at 11p13. Deletion of genes located on the tip of the short arm of 11 (11p15.5) is lethal to the A(L) hybrid, so that CD59 mutants that lose the entire chromosome 11 die and escape detection. In contrast, deletion of the 11p15.5 region is not lethal in the hybrid A(L)C, allowing for the detection of chromosome loss or other chromosomal mutations involving 11p15.5. The 55 MeV protons and 32 MeV/nucleon nitrogen ions were each about 10 times more mutagenic per unit dose at the CD59 locus in A(L)C cells than in A(L) cells. In the case of nitrogen ions, the mutations observed in A(L)C cells were predominantly due to chromosome loss events or 11p deletions, often containing a breakpoint in the pericentromeric region. The increase in the CD59(-) mutant fraction for A(L)C cells exposed to protons was associated with either translocation of portions of 11q onto a hamster chromosome, or discontinuous or "skipping" mutations. We demonstrate here that A(L)C cells are a powerful tool that will aid in the understanding of the mutagenic effects of different types of ionizing radiation.
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Heggendorn, Fabiano Luiz; Silva, Gabriela Cristina de Carvalho; Cardoso, Elisama Azevedo; Castro, Helena Carla; Gonçalves, Lúcio Souza; Dias, Eliane Pedra; Lione, Viviane de Oliveira Freitas; Lutterbach, Márcia Teresa Soares
2016-01-01
This study assessed the cell viability of the inoculation vehicle of BACCOR (a combination of sulfate-reducing bacteria plus a culture media for bacteria), a biopharmaceutical product under development for dental use as aid in fractured endodontic file removal from the root canal. Different culture media for bacteria were evaluated: modified Postgate E (MCP-E mod), Modified Postgate E without Agar-agar (MCP-E w/Ag), Postgate C with Agar-agar (MCP-C Ag) and Postgate C without Agar-agar (MCP-C w/Ag). Cytotoxicity was quantified by the MTT test, exposing L929 and Vero cell lines to the vehicles over 24 h. The exposure of L929 cell line to MCP-E w/Ag resulted in biocompatibility (52% cell viability), while the exposure of the Vero kidney line revealed only MCP-E mod as cytotoxic. When diluted, all the vehicles showed biocompatibility with both cell lines. MCP-E w/Ag was the vehicle chosen for BACCOR, because of its biocompatibility with the cells used.
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Identification of a novel rhabdovirus in Spodoptera frugiperda cell lines.
Ma, Hailun; Galvin, Teresa A; Glasner, Dustin R; Shaheduzzaman, Syed; Khan, Arifa S
2014-06-01
The Sf9 cell line, derived from Spodoptera frugiperda, is used as a cell substrate for biological products, and no viruses have been reported in this cell line after extensive testing. We used degenerate PCR assays and massively parallel sequencing (MPS) to identify a novel RNA virus belonging to the order Mononegavirales in Sf9 cells. Sequence analysis of the assembled virus genome showed the presence of five open reading frames (ORFs) corresponding to the genes for the N, P, M, G, and L proteins in other rhabdoviruses and an unknown ORF of 111 amino acids located between the G- and L-protein genes. BLAST searches indicated that the S. frugiperda rhabdovirus (Sf-rhabdovirus) was related in a limited region of the L-protein gene to Taastrup virus, a newly discovered member of the Mononegavirales from a leafhopper (Hemiptera), and also to plant rhabdoviruses, particularly in the genus Cytorhabdovirus. Phylogenetic analysis of sequences in the L-protein gene indicated that Sf-rhabdovirus is a novel virus that branched with Taastrup virus. Rhabdovirus morphology was confirmed by transmission electron microscopy of filtered supernatant samples from Sf9 cells. Infectivity studies indicated potential transient infection by Sf-rhabdovirus in other insect cell lines, but there was no evidence of entry or virus replication in human cell lines. Sf-rhabdovirus sequences were also found in the Sf21 parental cell line of Sf9 cells but not in other insect cell lines, such as BT1-TN-5B1-4 (Tn5; High Five) cells and Schneider's Drosophila line 2 [D.Mel.(2); SL2] cells, indicating a species-specific infection. The results indicate that conventional methods may be complemented by state-of-the-art technologies with extensive bioinformatics analysis for identification of novel viruses. The Spodoptera frugiperda Sf9 cell line is used as a cell substrate for the development and manufacture of biological products. Extensive testing has not previously identified any viruses in this cell
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Identification of a Novel Rhabdovirus in Spodoptera frugiperda Cell Lines
Ma, Hailun; Galvin, Teresa A.; Glasner, Dustin R.; Shaheduzzaman, Syed
2014-01-01
ABSTRACT The Sf9 cell line, derived from Spodoptera frugiperda, is used as a cell substrate for biological products, and no viruses have been reported in this cell line after extensive testing. We used degenerate PCR assays and massively parallel sequencing (MPS) to identify a novel RNA virus belonging to the order Mononegavirales in Sf9 cells. Sequence analysis of the assembled virus genome showed the presence of five open reading frames (ORFs) corresponding to the genes for the N, P, M, G, and L proteins in other rhabdoviruses and an unknown ORF of 111 amino acids located between the G- and L-protein genes. BLAST searches indicated that the S. frugiperda rhabdovirus (Sf-rhabdovirus) was related in a limited region of the L-protein gene to Taastrup virus, a newly discovered member of the Mononegavirales from a leafhopper (Hemiptera), and also to plant rhabdoviruses, particularly in the genus Cytorhabdovirus. Phylogenetic analysis of sequences in the L-protein gene indicated that Sf-rhabdovirus is a novel virus that branched with Taastrup virus. Rhabdovirus morphology was confirmed by transmission electron microscopy of filtered supernatant samples from Sf9 cells. Infectivity studies indicated potential transient infection by Sf-rhabdovirus in other insect cell lines, but there was no evidence of entry or virus replication in human cell lines. Sf-rhabdovirus sequences were also found in the Sf21 parental cell line of Sf9 cells but not in other insect cell lines, such as BT1-TN-5B1-4 (Tn5; High Five) cells and Schneider's Drosophila line 2 [D.Mel.(2); SL2] cells, indicating a species-specific infection. The results indicate that conventional methods may be complemented by state-of-the-art technologies with extensive bioinformatics analysis for identification of novel viruses. IMPORTANCE The Spodoptera frugiperda Sf9 cell line is used as a cell substrate for the development and manufacture of biological products. Extensive testing has not previously identified any
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7 CFR 929.45 - Research and development.
Code of Federal Regulations, 2010 CFR
2010-01-01
... establishment of production research, marketing research, and market development projects, including paid... 7 Agriculture 8 2010-01-01 2010-01-01 false Research and development. 929.45 Section 929.45 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing...
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Lung, H L; Ip, W K; Wong, C K; Mak, N K; Chen, Z Y; Leung, K N
2002-12-06
A novel approach for the treatment of leukemia is the differentiation therapy in which immature leukemia cells are induced to attain a mature phenotype when exposed to differentiation inducers, either alone or in combinations with other chemotherapeutic or chemopreventive drugs. Over the past decade, numerous studies indicated that green tea catechins (GTC) could suppress the growth and induce apoptosis on a number of human cancer cell lines. However, the differentiation-inducing activity of GTC on human tumors remains poorly understood. In the present study, the effect of the major GTC epigallocatechin-3-gallate (EGCG) on the proliferation and differentiation of a human eosinophilc leukemic cell line, EoL-1, was examined. Our results showed that EGCG suppressed the proliferation of the EoL-1 cells in a dose-dependent manner, with an estimated IC(50) value of 31.5 microM. On the other hand, EGCG at a concentration of 40 microM could trigger the EoL-1 cells to undergo morphological differentiation into mature eosinophil-like cells. Using RT-PCR and flow cytometry, it was found that EGCG upregulated the gene and protein expression of two eosinophil-specific granule proteins, the major basic protein (MBP) and eosinophil peroxidase (EPO), in EoL-1 cells. Taken together, our findings suggest that EGCG can exhibit anti-leukemic activity on a human eosinophilic cell line EoL-1 by suppressing the proliferation and by inducing the differentiation of the leukemia cells.
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7 CFR 929.72 - Duration of immunities.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Duration of immunities. 929.72 Section 929.72 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing... Duration of immunities. The benefits, privileges, and immunities conferred upon any person by virtue of...
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7 CFR 929.72 - Duration of immunities.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Duration of immunities. 929.72 Section 929.72 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing... Duration of immunities. The benefits, privileges, and immunities conferred upon any person by virtue of...
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7 CFR 929.72 - Duration of immunities.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Duration of immunities. 929.72 Section 929.72 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing... Duration of immunities. The benefits, privileges, and immunities conferred upon any person by virtue of...
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7 CFR 929.72 - Duration of immunities.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Duration of immunities. 929.72 Section 929.72 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING... Duration of immunities. The benefits, privileges, and immunities conferred upon any person by virtue of...
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7 CFR 929.72 - Duration of immunities.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Duration of immunities. 929.72 Section 929.72 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING... Duration of immunities. The benefits, privileges, and immunities conferred upon any person by virtue of...
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7 CFR 929.24 - Failure to nominate.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Failure to nominate. 929.24 Section 929.24 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.125 - Committee review procedures.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Committee review procedures. 929.125 Section 929.125 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.24 - Failure to nominate.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Failure to nominate. 929.24 Section 929.24 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.33 - Expenses and compensation.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Expenses and compensation. 929.33 Section 929.33 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.21 - Term of office.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Term of office. 929.21 Section 929.21 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.24 - Failure to nominate.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Failure to nominate. 929.24 Section 929.24 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.21 - Term of office.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Term of office. 929.21 Section 929.21 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.70 - Proceedings after termination.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Proceedings after termination. 929.70 Section 929.70 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.24 - Failure to nominate.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Failure to nominate. 929.24 Section 929.24 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.21 - Term of office.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Term of office. 929.21 Section 929.21 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.33 - Expenses and compensation.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Expenses and compensation. 929.33 Section 929.33 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.24 - Failure to nominate.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Failure to nominate. 929.24 Section 929.24 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.125 - Committee review procedures.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Committee review procedures. 929.125 Section 929.125 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.125 - Committee review procedures.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Committee review procedures. 929.125 Section 929.125 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.21 - Term of office.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Term of office. 929.21 Section 929.21 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.21 - Term of office.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Term of office. 929.21 Section 929.21 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.125 - Committee review procedures.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Committee review procedures. 929.125 Section 929.125 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.70 - Proceedings after termination.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Proceedings after termination. 929.70 Section 929.70 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.33 - Expenses and compensation.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Expenses and compensation. 929.33 Section 929.33 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.70 - Proceedings after termination.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Proceedings after termination. 929.70 Section 929.70 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.33 - Expenses and compensation.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Expenses and compensation. 929.33 Section 929.33 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.33 - Expenses and compensation.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Expenses and compensation. 929.33 Section 929.33 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.70 - Proceedings after termination.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Proceedings after termination. 929.70 Section 929.70 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.70 - Proceedings after termination.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Proceedings after termination. 929.70 Section 929.70 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Targeted delivery of melittin to cancer cells by AS1411 anti-nucleolin aptamer.
Rajabnejad, Seyed Hossein; Mokhtarzadeh, Ahad; Abnous, Khalil; Taghdisi, Seyed Mohammad; Ramezani, Mohammad; Razavi, Bibi Marjan
2018-06-01
Melittin, a small water-soluble cationic amphipathic α-helical linear peptide, consisted of 26 amino acids, is the honeybee venom major constituent. Several reports have proved the lytic and apoptotic effects of melittin in several cancerous cell lines. In this study, we aimed to fabricate an AS1411 aptamer-melittin to specifically deliver melittin to nucleolin positive cells (A549). Melittin was covalently attached to antinucleolin aptamer (AS1411) and its toxicity in A549 (nucleolin positive) and L929 (nucleolin negative) was studied using MTT and Annexin V flow cytometry methods. Aptamer-melittin conjugate formation was confirmed by gel electrophoresis. Hemolytic effect of aptamer-melittin conjugate was compared to melittin alone. The aptamer-melittin conjugate showed efficient cell uptake and was more cytotoxic in A549 cells than melittin (p < .001). This complex was less toxic in control cells. Competitive inhibition assay confirmed that aptamer-melittin complex delivery occurred through receptor-ligand interaction on the cell surface. Moreover, aptamer-melittin showed a significantly less hemolytic activity as compared with free melittin. This study showed that melittin could be specifically delivered to A549 cells when it was covalently conjugated to antinucleolin aptamer (AS1411) in vitro. This system can reduce the cytotoxic effects of melittin on cells with no nucleolin receptor overexpression which comprise most of normal cells such as L929 cells.
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Abdel-Lateef, Ezzat; Mahmoud, Faten; Hammam, Olfat; El-Ahwany, Eman; El-Wakil, Eman; Kandil, Sherihan; Abu Taleb, Hoda; El-Sayed, Mortada; Hassenein, Hanaa
2016-09-01
The present study was designed to identify the chemical constituents of the methanolic extract of Curcuma longa L. rhizomes and their inhibitory effect on a hepatoma cell line. The methanolic extract was subjected to GC-MS analysis to identify the volatile constituents and the other part of the same extract was subjected to liquid column chromatographic separation to isolate curcumin. The inhibition of cell growth in the hepatoma cell line and the cytopathological changes were studied. GC-MS analysis showed the presence of fifty compounds in the methanolic extract of C. longa. The major compounds were ar-turmerone (20.50 %), β-sesquiphellandrene (5.20 %) and curcumenol (5.11 %). Curcumin was identified using IR, 1H and 13C NMR. The inhibition of cell growth by curcumin (IC50 = 41.69 ± 2.87 μg mL-1) was much more effective than that of methanolic extract (IC50 = 196.12 ± 5.25 μg mL-1). Degenerative and apoptotic changes were more evident in curcumin- treated hepatoma cells than in those treated with the methanol extract. Antitumor potential of the methanolic extract may be attributed to the presence of sesquiterpenes and phenolic constituents including curcumin (0.051 %, 511.39 μg g-1 dried methanol extract) in C. longa rhizomes.
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7 CFR 929.11 - To can, freeze, or dehydrate.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false To can, freeze, or dehydrate. 929.11 Section 929.11... LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Definitions § 929.11 To can, freeze, or dehydrate. To can, freeze, or dehydrate means to convert cranberries into canned, frozen, or dehydrated...
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7 CFR 929.45 - Research and development.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Research and development. 929.45 Section 929.45... development. (a) The committee, with the approval of the Secretary, may establish or provide for the establishment of production research, marketing research, and market development projects, including paid...
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Arul, Melanie; Roslani, April Camilla; Cheah, Swee Hung
2017-05-01
Tumor heterogeneity may give rise to differential responses to chemotherapy drugs. Therefore, unraveling tumor heterogeneity has an implication for biomarker discovery and cancer therapeutics. To test this phenomenon, we investigated the differential responses of three secondary colorectal cancer cell lines of different origins (HCT116, HT29, and SW620 cells) and four novel primary cell lines obtained from different colorectal cancer patients to 5-fluorouracil (5-FU) and oxaliplatin (L-OHP) and explored the differences in gene expression among the primary cell lines in response to exposure to cytotoxic drugs. Cells were exposed to different doses of 5-FU and L-OHP separately or in combinations of equitoxic drug or equimolar drug ratios (median effect of Chou-Talalay principle). Cell viability was assessed using MTT assay and the respective IC 50 values were determined. Changes in gene expression in primary cell lines after exposure to the same drug doses were compared using real-time PCR array. The sensitivities (IC 50 ) of different cell lines, both secondary and primary, to 5-FU and L-OHP were significantly different, whether in monotherapy or combined treatment. Primary cell lines needed higher doses to reach IC 50 . There were variations in gene expression among the primary cell lines of different chemosensitivities to the challenge of the same combined dose of 5-FU and L-OHP. The results confirm the heterogeneous nature of colorectal cancer cells from different patient tumors. Studies using primary cancer cells established from patient's tumors rather than secondary cell lines will more closely reflect the actual character of the disease.
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Kibler, C; Schermutzki, F; Waller, H D; Timpl, R; Müller, C A; Klein, G
1998-06-01
Multiple myeloma represents a human B cell malignancy which is characterized by a predominant localization of the malignant cell clone within the bone marrow. With the exception of the terminal stage of the disease the myeloma tumor cells do not circulate in the peripheral blood. The bone marrow microenvironment is believed to play an important role in homing, proliferation and terminal differentiation of myeloma cells. Here we have studied the expression of several extracellular matrix (ECM) molecules in the bone marrow of multiple myeloma patients and analyzed their adhesive capacities with four different human myeloma-derived cell lines. All ECM molecules analyzed (tenascin, laminin, fibronectin, collagen types I, III, V and VI) could be detected in bone marrow cryostat sections of multiple myeloma patients. Adhesion assays showed that only laminin, the microfibrillar collagen type VI and fibronectin were strong adhesive components for the myeloma cell lines U266, IM-9, OPM-2 and NCI-H929. Tenascin and collagen type I were only weak adhesive substrates for these myeloma cells. Adhesion to laminin and fibronectin was beta 1-integrin-mediated since addition of anti-beta 1-integrin antibodies could inhibit the binding of the four different cell types to both matrix molecules. In contrast, integrins do not seem to be involved in binding of the myeloma cells to collagen type VI. Instead, inhibition of binding by heparin suggested that membrane-bound heparan sulfate proteoglycans are responsible ligands for binding to collagen type VI. Adhesion assays with several B-cell lines resembling earlier differentiation stages revealed only weak interactions with tenascin and no interactions with collagen type VI, laminin or fibronectin. In summary, the interactions of human myeloma cells with the extracellular matrix may explain the specific retention of the plasma cells within the bone marrow.
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Characterization of a Merkel Cell Polyomavirus-Positive Merkel Cell Carcinoma Cell Line CVG-1.
Velásquez, Celestino; Amako, Yutaka; Harold, Alexis; Toptan, Tuna; Chang, Yuan; Shuda, Masahiro
2018-01-01
Merkel cell polyomavirus (MCV) plays a causal role in ∼80% of Merkel cell carcinomas (MCC). MCV is clonally integrated into the MCC tumor genome, which results in persistent expression of large T (LT) and small T (sT) antigen oncoproteins encoded by the early locus. In MCV-positive MCC tumors, LT is truncated by premature stop codons or deletions that lead to loss of the C-terminal origin binding (OBD) and helicase domains important for replication. The N-terminal Rb binding domain remains intact. MCV-positive cell lines derived from MCC explants have been valuable tools to study the molecular mechanism of MCV-induced Merkel cell carcinogenesis. Although all cell lines have integrated MCV and express truncated LT antigens, the molecular sizes of the LT proteins differ between cell lines. The copy number of integrated viral genome also varies across cell lines, leading to significantly different levels of viral protein expression. Nevertheless, these cell lines share phenotypic similarities in cell morphology, growth characteristics, and neuroendocrine marker expression. Several low-passage MCV-positive MCC cell lines have been established since the identification of MCV. We describe a new MCV-positive MCV cell line, CVG-1, with features distinct from previously reported cell lines. CVG-1 tumor cells grow in more discohesive clusters in loose round cell suspension, and individual cells show dramatic size heterogeneity. It is the first cell line to encode an MCV sT polymorphism resulting in a unique leucine (L) to proline (P) substitution mutation at amino acid 144. CVG-1 possesses a LT truncation pattern near identical to that of MKL-1 cells differing by the last two C-terminal amino acids and also shows an LT protein expression level similar to MKL-1. Viral T antigen knockdown reveals that, like other MCV-positive MCC cell lines, CVG-1 requires T antigen expression for cell proliferation.
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Pereira, Joana M; Peixoto, Vanessa; Teixeira, Alexandra; Sousa, Diana; Barros, Lillian; Ferreira, Isabel C F R; Vasconcelos, M Helena
2018-06-05
The cell growth inhibitory activity of the hydroethanolic extract of Achillea millefolium was studied in human tumor cell lines (NCI-H460 and HCT-15) and its mechanism of action was investigated. The GI 50 concentration was determined with the sulforhodamine B assay and cell cycle and apoptosis were analyzed by flow cytometry following incubation with PI or Annexin V FITC/PI, respectively. The expression levels of proteins involved in cell cycle and apoptosis were analyzed by Western blot. The extracts were characterized regarding their phenolic composition by LC-DAD-ESI/MS. 3,5-O-Dicaffeoylquinic acid, followed by 5-O-caffeoylquinic acid, were the main phenolic acids, while, luteolin-O-acetylhexoside and apigenin-O-acetylhexoside were the main flavonoids. This extract decreased the growth of the tested cell lines, being more potent in HCT-15 and then in NCI-H460 cells. Two different concentrations of the extract (75 and 100 μg/mL) caused alterations in cell cycle profile and increased apoptosis levels in HCT-15 and NCI-H460 cells. Moreover, the extract caused an increase in p53 and p21 expression in NCI-H460 cells (which have wt p53), and reduced XIAP levels in HCT-15 cells (with mutant p53). This work enhances the importance of A. millefolium as source of bioactive phenolic compounds, particularly of XIAP inhibitors. Copyright © 2018 Elsevier Ltd. All rights reserved.
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Cell-Cycle Inhibition by Helicobacter pylori L-Asparaginase
Scotti, Claudia; Sommi, Patrizia; Pasquetto, Maria Valentina; Cappelletti, Donata; Stivala, Simona; Mignosi, Paola; Savio, Monica; Chiarelli, Laurent Roberto; Valentini, Giovanna; Bolanos-Garcia, Victor M.; Merrell, Douglas Scott; Franchini, Silvia; Verona, Maria Luisa; Bolis, Cristina; Solcia, Enrico; Manca, Rachele; Franciotta, Diego; Casasco, Andrea; Filipazzi, Paola; Zardini, Elisabetta; Vannini, Vanio
2010-01-01
Helicobacter pylori (H. pylori) is a major human pathogen causing chronic gastritis, peptic ulcer, gastric cancer, and mucosa-associated lymphoid tissue lymphoma. One of the mechanisms whereby it induces damage depends on its interference with proliferation of host tissues. We here describe the discovery of a novel bacterial factor able to inhibit the cell-cycle of exposed cells, both of gastric and non-gastric origin. An integrated approach was adopted to isolate and characterise the molecule from the bacterial culture filtrate produced in a protein-free medium: size-exclusion chromatography, non-reducing gel electrophoresis, mass spectrometry, mutant analysis, recombinant protein expression and enzymatic assays. L-asparaginase was identified as the factor responsible for cell-cycle inhibition of fibroblasts and gastric cell lines. Its effect on cell-cycle was confirmed by inhibitors, a knockout strain and the action of recombinant L-asparaginase on cell lines. Interference with cell-cycle in vitro depended on cell genotype and was related to the expression levels of the concurrent enzyme asparagine synthetase. Bacterial subcellular distribution of L-asparaginase was also analysed along with its immunogenicity. H. pylori L-asparaginase is a novel antigen that functions as a cell-cycle inhibitor of fibroblasts and gastric cell lines. We give evidence supporting a role in the pathogenesis of H. pylori-related diseases and discuss its potential diagnostic application. PMID:21085483
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Mahasti, Sahabi; Sattari, Mandana; Romoozi, Elham; Akbar-Zadeh Baghban, Alireza
2011-01-01
Resin cements, regardless of their biocompatibility, have been widely used in restorative dentistry during the recent years. These cements contain hydroxy ethyl methacrylate (HEMA) molecules which are claimed to penetrate into dentinal tubules and may affect dental pulp. Since tooth preparation for metal ceramic restorations involves a large surface of the tooth, cytotoxicity of these cements would be more important in fixed prosthodontic treatments. The purpose of this study was to compare the cytotoxicity of two resin cements (Panavia F2 and Rely X Plus) versus zinc phosphate cement (Harvard) using rat L929-fibroblasts in vitro. In this experimental study, ninety hollow glass cylinders (internal diameter 5-mm, height 2-mm) were made and divided into three groups. Each group was filled with one of three experimental cements; Harvard Zinc Phosphate cement, Panavia F2 resin cement and Rely X Plus resin cement. L929- Fibroblast were passaged and subsequently cultured in 6-well plates of 5×10(5) cells each. The culture medium was RPMI_ 1640. All samples were incubated in CO2. Using enzyme-linked immune-sorbent assay (ELISA) and (3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) (MTT) assay, the cytotoxicity of the cements was investigated at 1 hour, 24 hours and one week post exposure. Statistical analyses were performed via two-way ANOVA and honestly significant difference (HSD) Tukey tests. This study revealed significant differences between the three cements at the different time intervals. Harvard cement displayed the greatest cytotoxicity at all three intervals. After 1 hour Panavia F2 showed the next greatest cytotoxicity, but after 24-hours and oneweek intervals Rely X Plus showed the next greatest cytotoxicity. The results further showed that cytotoxicity decreased significantly in the Panavia F2 group with time (p<0.005), cytotoxicity increased significantly in the Rely X Plus group with time (p<0.001), and the Harvard cement group failed to
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Fares, Mona; Abedi-Valugerdi, Manuchehr; Hassan, Moustapha; Potácová, Zuzana
2015-07-31
We investigated mechanisms of cytotoxicity induced by doxycycline (doxy) and minocycline (mino) in the chronic myeloid leukemia K562 cell line. Doxy and mino induced cell death in exposure-dependent manner. While annexin V/propidium iodide staining was consistent with apoptosis, the morphological changes in Giemsa staining were more equivocal. A pancaspase inhibitor Z-VAD-FMK partially reverted cell death morphology, but concurrently completely prevented PARP cleavage. Mitochondrial involvement was detected as dissipation of mitochondrial membrane potential and cytochrome C release. DNA double strand breaks detected with γH2AX antibody and caspase-2 activation were found early after the treatment start, but caspase-3 activation was a late event. Decrement of Bcl-xL protein levels and electrophoretic shift of Bcl-xL molecule were induced by both drugs. Phosphorylation of Bcl-xL at serine 62 was ruled out. Similarly, Bcr/Abl tyrosine kinase levels were decreased. Lysosomal inhibitor chloroquine restored Bcl-xL and Bcr/Abl protein levels and inhibited caspase-3 activation. Thus, the cytotoxicity of doxy and mino in K562 cells is mediated by DNA damage, Bcl-xL deamidation and lysosomal degradation with activation of mitochondrial pathway of apoptosis. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
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7 CFR 929.67 - Right of the Secretary.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Right of the Secretary. 929.67 Section 929.67 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.67 - Right of the Secretary.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Right of the Secretary. 929.67 Section 929.67 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.67 - Right of the Secretary.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Right of the Secretary. 929.67 Section 929.67 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.67 - Right of the Secretary.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Right of the Secretary. 929.67 Section 929.67 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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7 CFR 929.67 - Right of the Secretary.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Right of the Secretary. 929.67 Section 929.67 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF...
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Zhao, Wujun; Cheng, Rui; Lim, So Hyun; Miller, Joshua R; Zhang, Weizhong; Tang, Wei; Xie, Jin; Mao, Leidong
2017-06-27
This paper reports a biocompatible and label-free cell separation method using ferrofluids that can separate a variety of low-concentration cancer cells from cell culture lines (∼100 cancer cells per mL) from undiluted white blood cells, with a throughput of 1.2 mL h -1 and an average separation efficiency of 82.2%. The separation is based on the size difference of the cancer cells and white blood cells, and is conducted in a custom-made biocompatible ferrofluid that retains not only excellent short-term viabilities but also normal proliferations of 7 commonly used cancer cell lines. A microfluidic device is designed and optimized specifically to shorten the time of live cells' exposure to ferrofluids from hours to seconds, by eliminating time-consuming off-chip sample preparation and extraction steps and integrating them on-chip to achieve a one-step process. As a proof-of-concept demonstration, a ferrofluid with 0.26% volume fraction was used in this microfluidic device to separate spiked cancer cells from cell lines at a concentration of ∼100 cells per mL from white blood cells with a throughput of 1.2 mL h -1 . The separation efficiencies were 80 ± 3%, 81 ± 5%, 82 ± 5%, 82 ± 4%, and 86 ± 6% for A549 lung cancer, H1299 lung cancer, MCF-7 breast cancer, MDA-MB-231 breast cancer, and PC-3 prostate cancer cell lines, respectively. The separated cancer cells' purity was between 25.3% and 28.8%. In addition, the separated cancer cells from this strategy showed an average short-term viability of 94.4 ± 1.3%, and these separated cells were cultured and demonstrated normal proliferation to confluence even after the separation process. Owing to its excellent biocompatibility and label-free operation and its ability to recover low concentrations of cancer cells from white blood cells, this method could lead to a promising tool for rare cell separation.
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7 CFR 929.61 - Outlets for excess cranberries.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Outlets for excess cranberries. 929.61 Section 929.61... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... excess cranberries. (a) Noncommercial outlets. Excess cranberries may be disposed of in noncommercial...
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7 CFR 929.61 - Outlets for excess cranberries.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Outlets for excess cranberries. 929.61 Section 929.61... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... excess cranberries. (a) Noncommercial outlets. Excess cranberries may be disposed of in noncommercial...
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7 CFR 929.61 - Outlets for excess cranberries.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Outlets for excess cranberries. 929.61 Section 929.61... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... excess cranberries. (a) Noncommercial outlets. Excess cranberries may be disposed of in noncommercial...
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7 CFR 929.61 - Outlets for excess cranberries.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Outlets for excess cranberries. 929.61 Section 929.61... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... excess cranberries. (a) Noncommercial outlets. Excess cranberries may be disposed of in noncommercial...
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Liu, Jun; Luo, Yan; Zheng, Liming; Liu, Qingqing; Yang, Zhongcai; Wang, Yongsheng; Su, Jianmin; Quan, Fusheng; Zhang, Yong
2013-10-01
This study was performed to qualify goat fetal fibroblast (GFF) cell lines for genetic modification and somatic cell nuclear transfer (SCNT) to produce human lysozyme (hLYZ) transgenic goats. Nine GFF cell lines were established from different fetuses, and the proliferative lifespan and chromosomal stability were analyzed. The results suggested that cell lines with a longer lifespan had stable chromosomes compared with those of cells lines with a shorter lifespan. According to the proliferative lifespan, we divided GFF cell lines into two groups: cell lines with a long lifespan (GFF1/2/7/8/9; group L) and cell lines with a short lifespan (GFF3/4/5/6; group S). Next, a hLYZ expression vector was introduced into these cell lines by electroporation. The efficiencies of colony formation, expansion in culture, and the quality of transgenic clonal cell lines were significant higher in group L than those in group S. The mean fusion rate and blastocyst rate in group L were higher than those in group S (80.3 ± 1.7 vs. 65.1 ± 4.2 % and 19.5 ± 0.6 vs. 15.1 ± 1.1 %, respectively, P < 0.05). After transferring cloned embryos into the oviducts of recipient goats, three live kids were born. PCR and Southern blot analyses confirmed integration of the transgene in cloned goats. In conclusion, the lifespan of GFF cell lines has a major effect on the efficiency to produce transgenic cloned goats. Therefore, the proliferative lifespan of primary cells may be used as a criterion to characterize the quality of cell lines for genetic modification and SCNT.
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47 CFR 22.929 - Application requirements for the Cellular Radiotelephone Service.
Code of Federal Regulations, 2010 CFR
2010-10-01
... 47 Telecommunication 2 2010-10-01 2010-10-01 false Application requirements for the Cellular Radiotelephone Service. 22.929 Section 22.929 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) COMMON CARRIER SERVICES PUBLIC MOBILE SERVICES Cellular Radiotelephone Service § 22.929 Application...
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47 CFR 22.929 - Application requirements for the Cellular Radiotelephone Service.
Code of Federal Regulations, 2013 CFR
2013-10-01
... 47 Telecommunication 2 2013-10-01 2013-10-01 false Application requirements for the Cellular Radiotelephone Service. 22.929 Section 22.929 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) COMMON CARRIER SERVICES PUBLIC MOBILE SERVICES Cellular Radiotelephone Service § 22.929 Application...
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47 CFR 22.929 - Application requirements for the Cellular Radiotelephone Service.
Code of Federal Regulations, 2014 CFR
2014-10-01
... 47 Telecommunication 2 2014-10-01 2014-10-01 false Application requirements for the Cellular Radiotelephone Service. 22.929 Section 22.929 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) COMMON CARRIER SERVICES PUBLIC MOBILE SERVICES Cellular Radiotelephone Service § 22.929 Application...
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Harada, Kazuki; Kitaguchi, Tetsuya; Tsuboi, Takashi
2015-05-15
Adrenaline reacts with three types of adrenergic receptors, α1, α2 and β-adrenergic receptors (ARs), inducing many physiological events including exocytosis. Although adrenaline has been shown to induce glucagon-like peptide-1 (GLP-1) secretion from intestinal L cells, the precise molecular mechanism by which adrenaline regulates GLP-1 secretion remains unknown. Here we show by live cell imaging that all types of adrenergic receptors are stimulated by adrenaline in enteroendocrine L cell line GLUTag cells and are involved in GLP-1 exocytosis. We performed RT-PCR analysis and found that α1B-, α2A-, α2B-, and β1-ARs were expressed in GLUTag cells. Application of adrenaline induced a significant increase of intracellular Ca(2+) and cAMP concentration ([Ca(2+)]i and [cAMP]i, respectively), and GLP-1 exocytosis in GLUTag cells. Blockade of α1-AR inhibited adrenaline-induced [Ca(2+)]i increase and exocytosis but not [cAMP]i increase, while blockade of β1-AR inhibited adrenaline-induced [cAMP]i increase and exocytosis but not [Ca(2+)]i increase. Furthermore, overexpression of α2A-AR suppressed the adrenaline-induced [cAMP]i increase and exocytosis. These results suggest that the fine-turning of GLP-1 secretion from enteroendocrine L cells is established by the balance between α1-, α2-, and β-ARs activation. Copyright © 2015 Elsevier Inc. All rights reserved.
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14 CFR 23.929 - Engine installation ice protection.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 14 Aeronautics and Space 1 2011-01-01 2011-01-01 false Engine installation ice protection. 23.929... General § 23.929 Engine installation ice protection. Propellers (except wooden propellers) and other components of complete engine installations must be protected against the accumulation of ice as necessary to...
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14 CFR 23.929 - Engine installation ice protection.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 14 Aeronautics and Space 1 2010-01-01 2010-01-01 false Engine installation ice protection. 23.929... General § 23.929 Engine installation ice protection. Propellers (except wooden propellers) and other components of complete engine installations must be protected against the accumulation of ice as necessary to...
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14 CFR 23.929 - Engine installation ice protection.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 14 Aeronautics and Space 1 2013-01-01 2013-01-01 false Engine installation ice protection. 23.929... General § 23.929 Engine installation ice protection. Propellers (except wooden propellers) and other components of complete engine installations must be protected against the accumulation of ice as necessary to...
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14 CFR 23.929 - Engine installation ice protection.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 14 Aeronautics and Space 1 2014-01-01 2014-01-01 false Engine installation ice protection. 23.929... General § 23.929 Engine installation ice protection. Propellers (except wooden propellers) and other components of complete engine installations must be protected against the accumulation of ice as necessary to...
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14 CFR 23.929 - Engine installation ice protection.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 14 Aeronautics and Space 1 2012-01-01 2012-01-01 false Engine installation ice protection. 23.929... General § 23.929 Engine installation ice protection. Propellers (except wooden propellers) and other components of complete engine installations must be protected against the accumulation of ice as necessary to...
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7 CFR 929.56 - Special provisions relating to withheld (restricted) cranberries.
Code of Federal Regulations, 2013 CFR
2013-01-01
...) cranberries. 929.56 Section 929.56 Agriculture Regulations of the Department of Agriculture (Continued... AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN... Regulations § 929.56 Special provisions relating to withheld (restricted) cranberries. (a) A handler shall...
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7 CFR 929.56 - Special provisions relating to withheld (restricted) cranberries.
Code of Federal Regulations, 2011 CFR
2011-01-01
...) cranberries. 929.56 Section 929.56 Agriculture Regulations of the Department of Agriculture (Continued... AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN... Regulations § 929.56 Special provisions relating to withheld (restricted) cranberries. (a) A handler shall...
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7 CFR 929.56 - Special provisions relating to withheld (restricted) cranberries.
Code of Federal Regulations, 2014 CFR
2014-01-01
...) cranberries. 929.56 Section 929.56 Agriculture Regulations of the Department of Agriculture (Continued... AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN... Regulations § 929.56 Special provisions relating to withheld (restricted) cranberries. (a) A handler shall...
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7 CFR 929.56 - Special provisions relating to withheld (restricted) cranberries.
Code of Federal Regulations, 2012 CFR
2012-01-01
...) cranberries. 929.56 Section 929.56 Agriculture Regulations of the Department of Agriculture (Continued... AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN... Regulations § 929.56 Special provisions relating to withheld (restricted) cranberries. (a) A handler shall...
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Cell-wall deficient L. monocytogenes L-forms feature abrogated pathogenicity
Schnell, Barbara; Staubli, Titu; Harris, Nicola L.; Rogler, Gerhard; Kopf, Manfred; Loessner, Martin J.; Schuppler, Markus
2014-01-01
Stable L-forms are cell wall-deficient bacteria which are able to multiply and propagate indefinitely, despite the absence of a rigid peptidoglycan cell wall. We investigated whether L-forms of the intracellular pathogen L. monocytogenes possibly retain pathogenicity, and if they could trigger an innate immune response. While phagocytosis of L. monocytogenes L-forms by non-activated macrophages sometimes resulted in an unexpected persistence of the bacteria in the phagocytes, they were effectively eliminated by IFN-γ preactivated or bone marrow-derived macrophages (BMM). These findings were in line with the observed down-regulation of virulence factors in the cell-wall deficient L. monocytogenes. Absence of Interferon-β (IFN-β) triggering indicated inability of L-forms to escape from the phagosome into the cytosol. Moreover, abrogated cytokine response in MyD88-deficient dendritic cells (DC) challenged with L. monocytogenes L-forms suggested an exclusive TLR-dependent host response. Taken together, our data demonstrate a strong attenuation of Listeria monocytogenes L-form pathogenicity, due to diminished expression of virulence factors and innate immunity recognition, eventually resulting in elimination of L-form bacteria from phagocytes. PMID:24904838
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Differential antimutagenicity of WR-1065 added after irradiation in L5178Y cell lines
NASA Technical Reports Server (NTRS)
Evans, H. H.; Horng, M. F.; Ricanati, M.; McCoy, E. C.
1999-01-01
The purpose of this study was to determine the antimutagenicity of WR-1065 added after irradiation of cells of cell lines differing in their ability to rejoin radiation-induced DNA double-strand breaks (DSBs). The postirradiation antimutagenicity of WR-1065 at the thymidine kinase locus was demonstrated for L5178Y (LY)-S1 cells that are deficient in repair of DNA DSBs. Less postirradiation antimutagenicity of WR-1065 was observed in LY-R16 and LY-SR1 cells, which are relatively efficient in DSB repair. Postirradiation treatment with WR-1065 had only a small stimulatory effect on DSB rejoining. A 3-h incubation of irradiated LY cells with WR-1065 caused slight changes in the distribution of cells in the phases of the cell cycle that differed between LY-S1 and LY-SR1 cells. Both LY-S1 and LY-SR1 cells were protected against the cytotoxic and mutagenic effects of radiation when WR-1065 was present 30 min before and during the irradiation. We conclude that the differential postirradiation effects of WR-1065 in the LY-S1 and LY-SR1 cells are not caused by differences in cellular uptake of the radioprotector or in its radical scavenging activity. Possible mechanisms for the postirradiation antimutagenicity of WR-1065 are discussed.
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Infectivity of Sf-rhabdovirus variants in insect and mammalian cell lines.
Maghodia, Ajay B; Jarvis, Donald L
2017-12-01
Sf-rhabdovirus was only recently identified as an adventitious agent of Spodoptera frugiperda (Sf) cell lines used as hosts for baculovirus vectors. As such, we still know little about its genetic variation, infectivity, and the potential impact of variation on the Sf-rhabdovirus-host interaction. Here, we characterized Sf-rhabdoviruses from two widely used Sf cell lines to confirm and extend information on Sf-rhabdovirus variation. We then used our novel Sf-rhabdovirus-negative (Sf-RVN) Sf cell line to assess the infectivity of variants with and without a 320bp X/L deletion and found both established productive persistent infections in Sf-RVN cells. We also assessed their infectivity using heterologous insect and mammalian cell lines and found neither established productive persistent infections in these cells. These results are the first to directly demonstrate Sf-rhabdoviruses are infectious for Sf cells, irrespective of the X/L deletion. They also confirm and extend previous results indicating Sf-rhabdoviruses have a narrow host range. Copyright © 2017 Elsevier Inc. All rights reserved.
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Establishment of an immortal chicken embryo liver-derived cell line.
Lee, Jeongyoon; Foster, Douglas N; Bottje, Walter G; Jang, Hyeon-Min; Chandra, Yohanna G; Gentles, Lauren E; Kong, Byung-Whi
2013-06-01
A continuously growing immortal cell substrate can be used for virus propagation, diagnostic purposes, and vaccine production. The aim of this study was to develop an immortal chicken cell line for efficient propagation of avian infectious viruses. From the various chicken embryo cells that were tested for life span extension, an immortalized chicken embryo liver (CEL) cell line, named CEL-im, was derived spontaneously without either oncogenic viruses or carcinogenic chemical treatment. Currently, CEL-im cells are growing 0.8 to 1.1 population doublings per day and have reached 120 passages. The CEL-im cell line is permissive for poultry infectious viruses, including avian metapneumovirus (AMPV), Marek's disease virus serotype 1 (MDV-1), and infectious laryngotracheitis virus. The CEL-im cells produced high AMPV titer (>10(5) pfu/mL), whereas very low titers (~10 pfu/mL) for MDV-1 and infectious laryngotracheitis virus were produced. To identify genetic alterations in the immortal CEL-im cell line, telomerase activity and mRNA expression for major cell cycle regulatory genes were determined during the immortalizing process. The CEL-im cell line has negative telomerase activity, and when compared with the primary passage 2 CEL cell counterpart, mRNA expression of tumor suppressor protein p53, mouse double minute 2 (Mdm2), cyclin dependent kinase (CDK) inhibitor p21 (p21(WAF)), and CDK inhibitor p16 (p16(INK4)) were downregulated in the CEL-im cell line, whereas retinoblastoma (Rb), transcription factor E2F, member 1 (E2F-1), and alternative reading frame of p16(INK4) (ARF) were upregulated. These results are similar to genetic alterations found previously in immortal chicken embryo fibroblast (CEF) cell lines that showed efficient propagation of MDV-1. Therefore, this newly established CEL-im cell line can serve as an alternative cell substrate for the propagation of poultry viruses, such as AMPV.
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SAHA-induced TRAIL-sensitisation of Multiple Myeloma cells is enhanced in 3D cell culture.
Arhoma, A; Chantry, A D; Haywood-Small, S L; Cross, N A
2017-11-15
Multiple Myeloma (MM) is currently incurable despite many novel therapies. Tumour Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) is a potential anti-tumour agent although effects as a single agent are limited. In this study, we investigated whether the Histone Deacetylase (HDAC) inhibitor SAHA can enhance TRAIL-induced apoptosis and target TRAIL resistance in both suspension culture, and 3D cell culture as a model of disseminated MM lesions that form in bone. The effects of SAHA and/or TRAIL in 6 Multiple Myeloma cell lines were assessed in both suspension cultures and in an Alginate-based 3D cell culture model. The effect of SAHA and/or TRAIL was assessed on apoptosis by assessment of nuclear morphology using Hoechst 33342/Propidium Iodide staining. Viable cell number was assessed by CellTiter-Glo luminescence assay, Caspase-8 and -9 activities were measured by Caspase-Glo™ assay kit. TRAIL-resistant cells were generated by culture of RPMI 8226 and NCI-H929 by acute exposure to TRAIL followed by selection of TRAIL-resistant cells. TRAIL significantly induced apoptosis in a dose-dependent manner in OPM-2, RPMI 8226, NCI-H929, U266, JJN-3 MM cell lines and ADC-1 plasma cell leukaemia cells. SAHA amplified TRAIL responses in all lines except OPM-2, and enhanced TRAIL responses were both via Caspase-8 and -9. SAHA treatment induced growth inhibition that further increased in the combination treatment with TRAIL in MM cells. The co-treatment of TRAIL and SAHA reduced viable cell numbers all cell lines. TRAIL responses were further potentiated by SAHA in 3D cell culture in NCI-H929, RPMI 8226 and U266 at lower TRAIL + SAHA doses than in suspension culture. However TRAIL responses in cells that had been selected for TRAIL resistance were not further enhanced by SAHA treatment. SAHA is a potent sensitizer of TRAIL responses in both TRAIL sensitive and resistant cell lines, in both suspension and 3D culture, however SAHA did not sensitise TRAIL-sensitive cell
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2012-01-01
Background Chondrosarcoma is the second most common primary sarcoma of bone. High-grade conventional chondrosarcoma and dedifferentiated chondrosarcoma have a poor outcome. In pre-clinical research aiming at the identification of novel treatment targets, the need for representative cell lines and model systems is high, but availability is scarce. Methods We developed and characterized three cell lines, derived from conventional grade III chondrosarcoma (L835), and dedifferentiated chondrosarcoma (L2975 and L3252) of bone. Proliferation and migration were studied and we used COBRA-FISH and array-CGH for karyotyping and genotyping. Immunohistochemistry for p16 and p53 was performed as well as TP53 and IDH mutation analysis. Cells were injected into nude mice to establish their tumorigenic potential. Results We show that the three cell lines have distinct migrative properties, L2975 had the highest migration rate and showed tumorigenic potential in mice. All cell lines showed chromosomal rearrangements with complex karyotypes and genotypic aberrations were conserved throughout late passaging of the cell lines. All cell lines showed loss of CDKN2A, while TP53 was wild type for exons 5–8. L835 has an IDH1 R132C mutation, L2975 an IDH2 R172W mutation and L3252 is IDH wild type. Conclusions Based on the stable culturing properties of these cell lines and their genotypic profile resembling the original tumors, these cell lines should provide useful functional models to further characterize chondrosarcoma and to evaluate new treatment strategies. PMID:22928481
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L1 Retrotransposon Heterogeneity in Ovarian Tumor Cell Evolution.
Nguyen, Thu H M; Carreira, Patricia E; Sanchez-Luque, Francisco J; Schauer, Stephanie N; Fagg, Allister C; Richardson, Sandra R; Davies, Claire M; Jesuadian, J Samuel; Kempen, Marie-Jeanne H C; Troskie, Robin-Lee; James, Cini; Beaven, Elizabeth A; Wallis, Tristan P; Coward, Jermaine I G; Chetty, Naven P; Crandon, Alexander J; Venter, Deon J; Armes, Jane E; Perrin, Lewis C; Hooper, John D; Ewing, Adam D; Upton, Kyle R; Faulkner, Geoffrey J
2018-06-26
LINE-1 (L1) retrotransposons are a source of insertional mutagenesis in tumor cells. However, the clinical significance of L1 mobilization during tumorigenesis remains unclear. Here, we applied retrotransposon capture sequencing (RC-seq) to multiple single-cell clones isolated from five ovarian cancer cell lines and HeLa cells and detected endogenous L1 retrotransposition in vitro. We then applied RC-seq to ovarian tumor and matched blood samples from 19 patients and identified 88 tumor-specific L1 insertions. In one tumor, an intronic de novo L1 insertion supplied a novel cis-enhancer to the putative chemoresistance gene STC1. Notably, the tumor subclone carrying the STC1 L1 mutation increased in prevalence after chemotherapy, further increasing STC1 expression. We also identified hypomethylated donor L1s responsible for new L1 insertions in tumors and cultivated cancer cells. These congruent in vitro and in vivo results highlight L1 insertional mutagenesis as a common component of ovarian tumorigenesis and cancer genome heterogeneity. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.
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Ye, Qing; Ding, Shao-Feng; Wang, Zhi-An; Feng, Jie; Tan, Wen-Bin
2014-01-01
Cancer constitutes a key pressure on public health regardless of the economy state in different countries. As a kind of highly malignant epithelial tumor, lacrimal gland adenoid cystic carcinoma can occur in any part of the body, such as salivary gland, submandibular gland, trachea, lung, breast, skin and lacrimal gland. Chemotherapy is one of the key treatment techniques, but drug resistance, especially MDR, seriously blunts its effects. As an element of the 60S large ribosomal subunit, the ribosomal protein L39-L gene appears to be documented specifically in the human testis and many human cancer samples of different origins. Total RNA of cultured drug-resistant and susceptible lacrimal gland adenoid cystic carcinoma cells was seperated, and real time quantitative RT-PCR were used to reveal transcription differences between amycin resistant and susceptible strains of lacrimal gland adenoid cystic carcinoma cells. Viability assays were used to present the amycin resistance difference in a RPL39-L transfected lacrimal gland adenoid cystic carcinoma cell line as compared to control vector and null-transfected lacrimal gland adenoid cystic carcinoma cell lines. The ribosomal protein L39-L transcription level was 6.5-fold higher in the drug-resistant human lacrimal gland adenoid cystic carcinoma cell line than in the susceptible cell line by quantitative RT-PCR analysis. The ribosomal protein L39-L transfected cells revealed enhanced drug resistance compared to plasmid vector-transfected or null-transfected cells as determined by methyl tritiated thymidine (3H-TdR) incorporation. The ribosomal protein L39-L gene could possibly have influence on the drug resistance mechanism of lacrimal gland adenoid cystic carcinoma cells.
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7 CFR 929.149 - Determination of sales history.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Determination of sales history. 929.149 Section 929... Determination of sales history. A sales history for each grower shall be computed by the Committee in the following manner. (a) For each grower with acreage with 7 or more years of sales history, a new sales...
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7 CFR 929.149 - Determination of sales history.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Determination of sales history. 929.149 Section 929... Determination of sales history. A sales history for each grower shall be computed by the Committee in the following manner. (a) For each grower with acreage with 7 or more years of sales history, a new sales...
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7 CFR 929.149 - Determination of sales history.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Determination of sales history. 929.149 Section 929... Determination of sales history. A sales history for each grower shall be computed by the Committee in the following manner. (a) For each grower with acreage with 7 or more years of sales history, a new sales...
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7 CFR 929.149 - Determination of sales history.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Determination of sales history. 929.149 Section 929... Determination of sales history. A sales history for each grower shall be computed by the Committee in the following manner. (a) For each grower with acreage with 7 or more years of sales history, a new sales...
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7 CFR 929.149 - Determination of sales history.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Determination of sales history. 929.149 Section 929... Determination of sales history. A sales history for each grower shall be computed by the Committee in the following manner. (a) For each grower with acreage with 6 or more years of sales history, a new sales...
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Szulc-Dabrowska, Lidia; Gregorczyk, Karolina P; Struzik, Justyna; Boratynska-Jasinska, Anna; Szczepanowska, Joanna; Wyzewski, Zbigniew; Toka, Felix N; Gierynska, Malgorzata; Ostrowska, Agnieszka; Niemialtowski, Marek G
2016-08-01
Ectromelia virus (ECTV, the causative agent of mousepox), which represents the same genus as variola virus (VARV, the agent responsible for smallpox in humans), has served for years as a model virus for studying mechanisms of poxvirus-induced disease. Despite increasing knowledge on the interaction between ECTV and its natural host-the mouse-surprisingly, still little is known about the cell biology of ECTV infection. Because pathogen interaction with the cytoskeleton is still a growing area of research in the virus-host cell interplay, the aim of the present study was to evaluate the consequences of ECTV infection on the cytoskeleton in a murine fibroblast cell line. The viral effect on the cytoskeleton was reflected by changes in migration of the cells and rearrangement of the architecture of tubulin, vimentin, and actin filaments. The virus-induced cytoskeletal rearrangements observed in these studies contributed to the efficient cell-to-cell spread of infection, which is an important feature of ECTV virulence. Additionally, during later stages of infection L929 cells produced two main types of actin-based cellular protrusions: short (actin tails and "dendrites") and long (cytoplasmic corridors). Due to diversity of filopodial extensions induced by the virus, we suggest that ECTV represents a valuable new model for studying processes and pathways that regulate the formation of cytoskeleton-based cellular structures. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.
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Bulera, S J; Sattler, C A; Gast, W L; Heath, S; Festerling, T A; Pitot, H C
1998-10-01
The hepatotoxicant thioacetamide (TH) has classically been used as a model to study hepatic necrosis; however, recent studies have shown that TH can also induce apoptosis. In this report we demonstrate that 2.68+/-0.54% of the albumin-SV40 T-antigen transgenic rat hepatocytes undergo TH-induced apoptosis, a level comparable to other in vivo models of liver apoptosis. In addition, TH could induce apoptosis and necrosis in the L37 albumin-SV40 T-antigen transgenic rat liver-derived cell line. Examination of dying L37 cells treated with 100 mM TH by electron microscopy revealed distinct morphological characteristics that could be attributed to apoptosis. Quantitation of apoptosis by FACS analysis 24 h after treatment with 100 mM TH revealed that 81.3+/-1.6% of the cells were undergoing apoptosis. In contrast, when L37 cells were treated with 250 mM TH, cells exhibited characteristics consistent with necrotic cell death. DNA fragmentation ladders were produced by growth factor withdrawal-induced apoptosis; however, in 100 mM TH-induced apoptosis, DNA fragmentation ladders were not observed. Analysis of endonuclease activity in L37 cells revealed that the enzymes were not inactivated in the presence of 100 mM TH. The data presented in this report indicate that the L37 cell line could be used to study the mechanism of TH-induced apoptosis that was not mediated through a mechanism requiring DNA fragmentation.
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Induction of cytochrome P450 (CYPIA) mRNA by polychlorinated dibenzo-p-dioxin (PCDD), polychlorinated dibenzofuran (PCDF) and polychlorinated biphenyl (PCB) congeners was measured in a zebrafish liver cell line (ZF-L). ZF-L cells were far less sensitive to PCDD, PCDF and PCB cong...
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7 CFR 929.64 - Verification of reports and records.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Verification of reports and records. 929.64 Section 929.64 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.71 - Effect of termination or amendment.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Effect of termination or amendment. 929.71 Section 929.71 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.71 - Effect of termination or amendment.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 8 2011-01-01 2011-01-01 false Effect of termination or amendment. 929.71 Section 929.71 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.71 - Effect of termination or amendment.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Effect of termination or amendment. 929.71 Section 929.71 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.64 - Verification of reports and records.
Code of Federal Regulations, 2010 CFR
2010-01-01
... 7 Agriculture 8 2010-01-01 2010-01-01 false Verification of reports and records. 929.64 Section 929.64 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.64 - Verification of reports and records.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Verification of reports and records. 929.64 Section 929.64 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.64 - Verification of reports and records.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Verification of reports and records. 929.64 Section 929.64 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.71 - Effect of termination or amendment.
Code of Federal Regulations, 2013 CFR
2013-01-01
... 7 Agriculture 8 2013-01-01 2013-01-01 false Effect of termination or amendment. 929.71 Section 929.71 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.64 - Verification of reports and records.
Code of Federal Regulations, 2012 CFR
2012-01-01
... 7 Agriculture 8 2012-01-01 2012-01-01 false Verification of reports and records. 929.64 Section 929.64 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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7 CFR 929.71 - Effect of termination or amendment.
Code of Federal Regulations, 2014 CFR
2014-01-01
... 7 Agriculture 8 2014-01-01 2014-01-01 false Effect of termination or amendment. 929.71 Section 929.71 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES...
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Borawska, Maria H; Naliwajko, Sylwia K; Moskwa, Justyna; Markiewicz-Żukowska, Renata; Puścion-Jakubik, Anna; Soroczyńska, Jolanta
2016-09-20
Propolis and Hypericum perforatum L. are natural products which contain many active compounds and have numerous beneficial effects, including an antitumor effect. Gliobmastoma multiforme (GBM) is a common primary brain tumor with poor prognosis and limited treatment options. In this study, the effect of propolis (EEP) combined with H. perforatum L. (HPE) on glioblastoma cell line U87MG was investigated for the first time. Anti-proliferative activity of EEP, HPE and their combination (EEP + HPE) was determined by a cytotoxicity test, DNA binding by [(3)H]-thymidine incorporation and cell migration assay. Anti-metastatic properties in U87MG treated with EEP, HPE and EEP + HPE were estimated on cells migration test (scratch assay) and metalloproteinases (MMP2 and MMP9) secretion (gelatin zymography). Combination of HPE and EEP extracts was found to have a time- and dose-dependent inhibitory effect on the viability of U87MG cells. This effect was significantly higher (p < 0.05) when compared to these two extracts applied separately, which was confirmed by the significant reduction of DNA synthesis and significantly higher mitochondrial membrane permeabilization. A significant decreasing in migration cells and in pro-MMP9 and pro-MMP2 secretion in U87MG cells were demonstrated after exposure to combination of EEP (30 μg/ml) with HPE (6.25 μg/ml). In this study, the combination of ethanolic extract from propolis and ethanolic extract of fresh-cut H. perforatum L. was proved the ability to reduce invasiveness of glioma cells through the inhibition of MMP2 and MMP9 secretion and suppression of cell migration. It has a more potent anti-proliferative effect on U87MG glioma cell line compared to using propolis and H. perforatum L. separately. Further studies are required to verify whether the examined extracts can activate apoptotic pathways.
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Control of cell proliferation by a porous chitosan scaffold with multiple releasing capabilities
NASA Astrophysics Data System (ADS)
Cai, Shu-Jyun; Li, Ching-Wen; Weihs, Daphne; Wang, Gou-Jen
2017-12-01
The aim of this study was to develop a porous chitosan scaffold with long-acting drug release as an artificial dressing to promote skin wound healing. The dressing was fabricated by pre-freezing at different temperatures (-20 and -80 °C) for different periods of time, followed by freeze-drying to form porous chitosan scaffolds with different pore sizes. The chitosan scaffolds were then used to investigate the effect of the controlled release of fibroblast growth factor-basic (bFGF) and transforming growth factor-β1 (TGFβ1) on mouse fibroblast cells (L929) and bovine carotid endothelial cells (BEC). The biocompatibility of the prepared chitosan scaffold was confirmed with WST-1 proliferation and viability assay, which demonstrated that the material is suitable for cell growth. The results of this study show that the pore sizes of the porous scaffolds prepared by freeze-drying can change depending on the pre-freezing temperature and time via the formation of ice crystals. In this study, the scaffolds with the largest pore size were found to be 153 ± 32 μm and scaffolds with the smallest pores to be 34 ± 9 μm. Through cell culture analysis, it was found that the concentration that increased proliferation of L929 cells for bFGF was 0.005 to 0.1 ng/mL, and the concentration for TGFβ1 was 0.005 to 1 ng/mL. The cell culture of the chitosan scaffold and growth factors shows that 3.75 ng of bFGF in scaffolds with pore sizes of 153 ± 32 μm can promote L929 cell proliferation, while 400 pg of TGFβ1 in scaffolds with pore size of 34 ± 9 μm can enhance the proliferation of L929 cells, but also inhibit BEC proliferation. It is proposed that the prepared chitosan scaffolds can form a multi-drug (bFGF and TGFβ1) release dressing that has the ability to control wound healing via regulating the proliferation of different cell types.
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24 CFR 200.929a - Fair Housing Accessibility Guidelines.
Code of Federal Regulations, 2010 CFR
2010-04-01
... Guidelines. 200.929a Section 200.929a Housing and Urban Development Regulations Relating to Housing and Urban... Fair Housing Accessibility Guidelines. Builders and developers may use the Department's Fair Housing Accessibility Guideline when designing or constructing covered multifamily dwelling units in order to comply...
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El-Schich, Zahra; Mölder, Anna; Tassidis, Helena; Härkönen, Pirkko; Falck Miniotis, Maria; Gjörloff Wingren, Anette
2015-03-01
We are using the label-free technique of holographic microscopy to analyze cellular parameters including cell number, confluence, cellular volume and area directly in the cell culture environment. We show that death-induced cells can be distinguished from untreated counterparts by the use of holographic microscopy, and we demonstrate its capability for cell death assessment. Morphological analysis of two representative cell lines (L929 and DU145) was performed in the culture flasks without any prior cell detachment. The two cell lines were treated with the anti-tumour agent etoposide for 1-3days. Measurements by holographic microscopy showed significant differences in average cell number, confluence, volume and area when comparing etoposide-treated with untreated cells. The cell volume of the treated cell lines was initially increased at early time-points. By time, cells decreased in volume, especially when treated with high doses of etoposide. In conclusion, we have shown that holographic microscopy allows label-free and completely non-invasive morphological measurements of cell growth, viability and death. Future applications could include real-time monitoring of these holographic microscopy parameters in cells in response to clinically relevant compounds. Copyright © 2015 Elsevier Inc. All rights reserved.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Walter, M.N.M.; School of Life and Health Science, Aston University, Aston Triangle, Birmingham, B4 7EJ; Wright, K.T.
2010-04-15
We have used in vitro scratch assays to examine the relative contribution of dermal fibroblasts and keratinocytes in the wound repair process and to test the influence of mesenchymal stem cell (MSC) secreted factors on both skin cell types. Scratch assays were established using single cell and co-cultures of L929 fibroblasts and HaCaT keratinocytes, with wound closure monitored via time-lapse microscopy. Both in serum supplemented and serum free conditions, wound closure was faster in L929 fibroblast than HaCaT keratinocyte scratch assays, and in co-culture the L929 fibroblasts lead the way in closing the scratches. MSC-CM generated under serum free conditionsmore » significantly enhanced the wound closure rate of both skin cell types separately and in co-culture, whereas conditioned medium from L929 or HaCaT cultures had no significant effect. This enhancement of wound closure in the presence of MSC-CM was due to accelerated cell migration rather than increased cell proliferation. A number of wound healing mediators were identified in MSC-CM, including TGF-{beta}1, the chemokines IL-6, IL-8, MCP-1 and RANTES, and collagen type I, fibronectin, SPARC and IGFBP-7. This study suggests that the trophic activity of MSC may play a role in skin wound closure by affecting both dermal fibroblast and keratinocyte migration, along with a contribution to the formation of extracellular matrix.« less
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Walter, M N M; Wright, K T; Fuller, H R; MacNeil, S; Johnson, W E B
2010-04-15
We have used in vitro scratch assays to examine the relative contribution of dermal fibroblasts and keratinocytes in the wound repair process and to test the influence of mesenchymal stem cell (MSC) secreted factors on both skin cell types. Scratch assays were established using single cell and co-cultures of L929 fibroblasts and HaCaT keratinocytes, with wound closure monitored via time-lapse microscopy. Both in serum supplemented and serum free conditions, wound closure was faster in L929 fibroblast than HaCaT keratinocyte scratch assays, and in co-culture the L929 fibroblasts lead the way in closing the scratches. MSC-CM generated under serum free conditions significantly enhanced the wound closure rate of both skin cell types separately and in co-culture, whereas conditioned medium from L929 or HaCaT cultures had no significant effect. This enhancement of wound closure in the presence of MSC-CM was due to accelerated cell migration rather than increased cell proliferation. A number of wound healing mediators were identified in MSC-CM, including TGF-beta1, the chemokines IL-6, IL-8, MCP-1 and RANTES, and collagen type I, fibronectin, SPARC and IGFBP-7. This study suggests that the trophic activity of MSC may play a role in skin wound closure by affecting both dermal fibroblast and keratinocyte migration, along with a contribution to the formation of extracellular matrix. Copyright 2010 Elsevier Inc. All rights reserved.
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Kirilyuk, Alexander; Tolstonog, Genrich V; Damert, Annette; Held, Ulrike; Hahn, Silvia; Löwer, Roswitha; Buschmann, Christian; Horn, Axel V; Traub, Peter; Schumann, Gerald G
2008-02-01
LINE-1 (L1) is a highly successful autonomous non-LTR retrotransposon and a major force shaping mammalian genomes. Although there are about 600 000 L1 copies covering 23% of the rat genome, full-length rat L1s (L1Rn) with intact open reading frames (ORFs) representing functional master copies for retrotransposition have not been identified yet. In conjunction with studies to elucidate the role of L1 retrotransposons in tumorigenesis, we isolated and characterized 10 different cDNAs from transcribed full-length L1Rn elements in rat chloroleukemia (RCL) cells, each encoding intact ORF1 proteins (ORF1p). We identified the first functional L1Rn retrotransposon from this pool of cDNAs, determined its activity in HeLa cells and in the RCL cell line the cDNAs originated from and demonstrate that it is mobilized in the tumor cell line in which it is expressed. Furthermore, we generated monoclonal antibodies directed against L1Rn ORF1 and ORF2-encoded recombinant proteins, analyzed the expression of L1-encoded proteins and found ORF1p predominantly in the nucleus. Our results support the hypothesis that the reported explosive amplification of genomic L1Rn sequences after their transcriptional activation in RCL cells is based on L1 retrotransposition. Therefore, L1 activity might be one cause for genomic instability observed during the progression of leukemia.
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Grenga, Italia; Donahue, Renee N; Gargulak, Morgan L; Lepone, Lauren M; Roselli, Mario; Bilusic, Marijo; Schlom, Jeffrey
2018-03-01
Avelumab has recently been approved by the Food and Drug Administration for the therapy of Merkel cell carcinoma and urothelial carcinoma. M7824 is a novel first-in-class bifunctional fusion protein comprising a monoclonal antibody against programmed death-ligand 1 (PD-L1, avelumab), fused to the extracellular domain of human transforming growth factor beta (TGFβ) receptor 2, which functions as a TGFβ "trap." Advanced urothelial tumors have been shown to express TGFβ, which possesses immunosuppressive properties that promote cancer progression and metastasis. The rationale for a combined molecule is to block the PD-1/PD-L1 interaction between tumor cells and immune cell infiltrate and simultaneously reduce or eliminate TGFβ from the tumor microenvironment. In this study, we explored the effect of M7824 on invasive urothelial carcinoma cell lines. Human urothelial (transitional cell) carcinoma cell lines HTB-4, HTB-1, and HTB-5 were treated with M7824, M7824mut (M7824 that is mutated in the anti-PD-L1 portion of the molecule and thus does not bind PD-L1), anti-PD-L1 (avelumab), or IgG1 isotype control monoclonal antibody, and were assessed for gene expression, cell-surface phenotype, and sensitivity to lysis by TRAIL, antigen-specific cytotoxic T lymphocytes and natural killer cells. M7824 retains the ability to mediate antibody-dependent cellular cytotoxicity of tumor cells, although in some cases to a lesser extent than anti-PD-L1. However, compared to anti-PD-L1, M7824 increases (A) gene expression of molecules involved in T-cell trafficking in the tumor (e.g., CXCL11), (B) TRAIL-mediated tumor cell lysis, and (C) antigen-specific CD8 + T-cell-mediated lysis of tumor cells. These studies demonstrate the immunomodulatory properties of M7824 on both tumor cell phenotype and immune-mediated lysis. Compared to anti-PD-L1 or M7824mut, M7824 induces immunogenic modulation of urothelial carcinoma cell lines, rendering them more susceptible to immune
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Chandipura virus growth kinetics in vertebrate cell lines, insect cell lines & embryonated eggs.
Jadi, R S; Sudeep, A B; Kumar, Satyendra; Arankalle, V A; Mishra, A C
2010-08-01
Since not much information on Chandipura virus is available, an attempt was made to study the growth kinetics of the virus in certain vertebrate, invertebrate cell lines and embryonated chicken eggs. Comparative study of Chandipura virus (CHPV) growth kinetics in three vertebrate cell lines [Vero E6, Rhabdo myosarcoma (RD), Porcine stable kidney (PS) cell lines], two insect cell lines [Aedes aegypti (AA) and Phlebotomus papatasi (PP-9) cell lines] and embryonated pathogen free chicken eggs was conducted, by tissue culture infective dose 50 per cent (TCID(50)) and indirect immunofluorescence assay (IFA). All the cell lines and embryonated egg supported the growth of CHPV and yielded high virus titre. The vertebrate cell lines showed distinct cytopathic effect (CPE) within 4-6 h post infection (PI), while no CPE was observed in insect cell lines. PP-9 cell line was the most sensitive system to CHPV as viral antigen could be detected at 1 h PI by IFA. Our results demonstrated that all the systems were susceptible to CHPV and achieved high yield of virus. However, the PP-9 cell line had an edge over the others due to its high sensitivity to the virus which might be useful for detection and isolation of the virus during epidemics.
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Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cells.
Fan, Lianchun; Kadura, Ibrahim; Krebs, Lara E; Hatfield, Christopher C; Shaw, Margaret M; Frye, Christopher C
2012-04-01
Although Chinese hamster ovary (CHO) cells, with their unique characteristics, have become a major workhorse for the manufacture of therapeutic recombinant proteins, one of the major challenges in CHO cell line generation (CLG) is how to efficiently identify those rare, high-producing clones among a large population of low- and non-productive clones. It is not unusual that several hundred individual clones need to be screened for the identification of a commercial clonal cell line with acceptable productivity and growth profile making the cell line appropriate for commercial application. This inefficiency makes the process of CLG both time consuming and laborious. Currently, there are two main CHO expression systems, dihydrofolate reductase (DHFR)-based methotrexate (MTX) selection and glutamine synthetase (GS)-based methionine sulfoximine (MSX) selection, that have been in wide industrial use. Since selection of recombinant cell lines in the GS-CHO system is based on the balance between the expression of the GS gene introduced by the expression plasmid and the addition of the GS inhibitor, L-MSX, the expression of GS from the endogenous GS gene in parental CHOK1SV cells will likely interfere with the selection process. To study endogenous GS expression's potential impact on selection efficiency, GS-knockout CHOK1SV cell lines were generated using the zinc finger nuclease (ZFN) technology designed to specifically target the endogenous CHO GS gene. The high efficiency (∼2%) of bi-allelic modification on the CHO GS gene supports the unique advantages of the ZFN technology, especially in CHO cells. GS enzyme function disruption was confirmed by the observation of glutamine-dependent growth of all GS-knockout cell lines. Full evaluation of the GS-knockout cell lines in a standard industrial cell culture process was performed. Bulk culture productivity improved two- to three-fold through the use of GS-knockout cells as parent cells. The selection stringency was
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49 CFR 236.929 - Training specific to roadway workers.
Code of Federal Regulations, 2011 CFR
2011-10-01
... 49 Transportation 4 2011-10-01 2011-10-01 false Training specific to roadway workers. 236.929... for Processor-Based Signal and Train Control Systems § 236.929 Training specific to roadway workers. (a) How is training for roadway workers to be coordinated with part 214? Training required under this...
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Liang Li; Tanaka, Michiko; Kawaguchi, Yasushi
2004-11-10
Previous results indicated that the herpes simplex virus 1 (HSV-1) U{sub L}31 gene is necessary and sufficient for localization of the U{sub L}34 protein exclusively to the nuclear membrane of infected Hep2 cells. In the current studies, a bacterial artificial chromosome containing the entire HSV-1 strain F genome was used to construct a recombinant viral genome in which a gene encoding kanamycin resistance was inserted in place of 262 codons of the 306 codon U{sub L}31 open reading frame. The deletion virus produced virus titers approximately 10- to 50-fold lower in rabbit skin cells, more than 2000-fold lower in Veromore » cells, and more than 1500-fold lower in CV1 cells, compared to a virus bearing a restored U{sub L}31 gene. The replication of the U{sub L}31 deletion virus was restored on U{sub L}31-complementing cell lines derived either from rabbit skin cells or CV1 cells. Confocal microscopy indicated that the majority of U{sub L}34 protein localized aberrantly in the cytoplasm and nucleoplasm of Vero cells and CV1 cells, whereas U{sub L}34 protein localized at the nuclear membrane in rabbit skin cells, and U{sub L}31 complementing CV1 cells infected with the U{sub L}31 deletion virus. We conclude that rabbit skin cells encode a function that allows proper localization of U{sub L}34 protein to the nuclear membrane. We speculate that this function partially complements that of U{sub L}31 and may explain why U{sub L}31 is less critical for replication in rabbit skin cells as opposed to Vero and CV1 cells.« less
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2014-01-01
Background Cell lines have been widely used in biomedical research. The community-based Cell Line Ontology (CLO) is a member of the OBO Foundry library that covers the domain of cell lines. Since its publication two years ago, significant updates have been made, including new groups joining the CLO consortium, new cell line cells, upper level alignment with the Cell Ontology (CL) and the Ontology for Biomedical Investigation, and logical extensions. Construction and content Collaboration among the CLO, CL, and OBI has established consensus definitions of cell line-specific terms such as ‘cell line’, ‘cell line cell’, ‘cell line culturing’, and ‘mortal’ vs. ‘immortal cell line cell’. A cell line is a genetically stable cultured cell population that contains individual cell line cells. The hierarchical structure of the CLO is built based on the hierarchy of the in vivo cell types defined in CL and tissue types (from which cell line cells are derived) defined in the UBERON cross-species anatomy ontology. The new hierarchical structure makes it easier to browse, query, and perform automated classification. We have recently added classes representing more than 2,000 cell line cells from the RIKEN BRC Cell Bank to CLO. Overall, the CLO now contains ~38,000 classes of specific cell line cells derived from over 200 in vivo cell types from various organisms. Utility and discussion The CLO has been applied to different biomedical research studies. Example case studies include annotation and analysis of EBI ArrayExpress data, bioassays, and host-vaccine/pathogen interaction. CLO’s utility goes beyond a catalogue of cell line types. The alignment of the CLO with related ontologies combined with the use of ontological reasoners will support sophisticated inferencing to advance translational informatics development. PMID:25852852
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Vascular endothelial growth factor expression and inhibition in uveal melanoma cell lines
Logan, Patrick; Burnier, Julia; Burnier, Miguel N.
2013-01-01
Background: Uveal melanoma (UM) is a disease that affects approximately five people per million in the United States. This disease metastasises predominantly to the liver, and treatment options following the clinical detection of these sequelae are limited. Vascular endothelial growth factor-A (VEGF-A) is the primary activator of tumour angiogenesis and functions by binding to VEGF-Receptor 2 (VEGF-R2) and is often required for tumour growth beyond 2–3 mm. The purpose of this study was to investigate the expression of VEGF-A and the primary VEGF-R2 in three UM cell lines. Furthermore, we investigated the effects of VEGF-A inhibition on receptor activation and production of other cytokines. Finally, the effects of VEGF-A inhibition on the proliferation, migration, and invasion in the cell lines were ascertained. Materials: Three UM cell lines (92.1, OCM-1, and UW-1) were incubated with and without the addition of 100 μg/mL of bevacizumab. VEGF-A expression under both conditions was determined by sandwich enzyme-linked immunosorbent assay (ELISA), and phosphorylated VEGF-R2 expression was determined using western blot. The effects of VEGF-A inhibition on 20 cytokines (IL-1a, IL-2, IL-5, IL-8, IL-12p70, GM-CSF, IFNy, CCL3, MMP-9, TNF-a, IL-1b, IL-4, IL-6, IL-10, IL-13, GRO, MCP-1, MIP-1b, and RANTES) were determined using a multiplex sandwich ELISA. Proliferation rates before and after treatment were evaluated via sulforhodamine B assay, and migration and invasion assays implementing the Boyden chamber technique, the latter with artificial extracellular matrix, were used to assess their respective abilities. The Student’s t-test was used to compare changes in cytokine expression following VEGF-A inhibition. Analysis of variance was used to compare changes in the functional abilities of three uveal melanoma cell lines following VEGF-A inhibition. A P-value < 0.05 was considered statistically significant. Results: All three cell lines produced copious amounts of
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Esmaeilbeig, Maryam; Kouhpayeh, Seyed Amin; Amirghofran, Zahra
2015-01-01
Background: Traditional herbal medicine is a valuable resource that provides new drugs for cancer treatment. Objectives: In this study we aim to screen and investigate the in vitro anti-tumor activities of ten species of plants commonly grown in Southern Iran. Materials and Methods: We used the MTT colorimetric assay to evaluate the cytotoxic activities of the methanol extracts of these plants on various tumor cell lines. The IC50 was calculated as a scale for this evaluation. Results: Satureja bachtiarica, Satureja hortensis, Thymus vulgaris, Thymus daenensis and Mentha lonigfolia showed the inhibitoriest effects on Jurkat cells with > 80% inhibition at 200 µg/mL. Satureja hortensis (IC50: 66.7 µg/mL) was the most effective. These plants also strongly inhibited K562 cell growth; Satureja bachtiarica (IC50: 28.3 µg/mL), Satureja hortensis (IC50: 52 µg/mL) and Thymus vulgaris (IC50: 87 µg/mL) were the most effective extracts. Cichorium intybus, Rheum ribes, Alhagi pseudalhagi and Glycyrrihza glabra also showed notable effects on the leukemia cell lines. The Raji cell line was mostly inhibited by Satureja bachtiarica and Thymus vulgaris with approximately 40% inhibition at 200µg/ml. The influence of these extracts on solid tumor cell lines was not strong. Fen cells were mostly affected by Glycyrrihza glabra (IC50: 182 µg/mL) and HeLa cells by Satureja hortensis (31.6% growth inhibitory effect at 200 µg/mL). Conclusions: Leukemic cell lines were more sensitive to the extracts than the solid tumor cell lines; Satureja hortensis, Satureja bachtiarica, Thymus vulgaris, Thymus daenensis and Mentha lonigfolia showed remarkable inhibitory potential. PMID:26634114
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Kishimoto, Mana; Nomoto, Ryohei; Mizuno, Masashi; Osawa, Ro
2017-01-01
Many probiotic lactobacilli and their extracellular polysaccharides (EPS) have beneficial immunological properties. However, it is unclear how they elicit the host immune response. We thus investigated the immunological properties of UV-killed Lactobacillus delbrueckii TU-1 and L. plantarum KM-9 cells as well as their extracellular polysaccharides (EPSs). High-performance liquid chromatography and ion exchange chromatography analyses showed that their EPSs differ in sugar composition and sugar fractionation. The immunological properties were evaluated in a semi-intestinal model using a Transwell co-culture system that employed human intestinal epithelial (Caco-2) cells on the apical side and murine macrophage (RAW264.7) cells on the basolateral side. The UV-killed cells and EPSs were added to the apical side to allow direct contact with Caco-2 cells and incubated for 6 hr. After incubation, the amounts of tumor necrosis factor-α and several cytokines released by RAW264.7 or Caco-2 cells were quantified by cytotoxic activity on L929 cells (murine fibrosarcoma cell line) and quantitative reverse-transcriptase PCR. We found that the UV-killed cells and their EPSs had immunological effects on RAW264.7 cells via Caco-2 cells. The RAW264.7 cells showed different cytokine production profiles when treated with UV-killed cells and EPSs. The UV-killed cells and EPSs promoted a Th1-type cellular response. Furthermore, we found that the UV-killed cells sent positive signals through Toll-like receptor (TLR) 2. Meanwhile, neither EPS sent a positive signal through TLR4 and TLR2. This evidence suggests that both UV-killed cells of the lactobacillus strains and their EPSs trigger a Th1-type immune response in a human host, with the former triggering the response via the TLRs expressed on its epithelium and the latter employing a mechanism yet to be determined, possibly involving a novel receptor that is designed to recognize specific patterns of repeating sugar in the EPSs.
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KISHIMOTO, Mana; NOMOTO, Ryohei; MIZUNO, Masashi; OSAWA, Ro
2017-01-01
Many probiotic lactobacilli and their extracellular polysaccharides (EPS) have beneficial immunological properties. However, it is unclear how they elicit the host immune response. We thus investigated the immunological properties of UV-killed Lactobacillus delbrueckii TU-1 and L. plantarum KM-9 cells as well as their extracellular polysaccharides (EPSs). High-performance liquid chromatography and ion exchange chromatography analyses showed that their EPSs differ in sugar composition and sugar fractionation. The immunological properties were evaluated in a semi-intestinal model using a Transwell co-culture system that employed human intestinal epithelial (Caco-2) cells on the apical side and murine macrophage (RAW264.7) cells on the basolateral side. The UV-killed cells and EPSs were added to the apical side to allow direct contact with Caco-2 cells and incubated for 6 hr. After incubation, the amounts of tumor necrosis factor-α and several cytokines released by RAW264.7 or Caco-2 cells were quantified by cytotoxic activity on L929 cells (murine fibrosarcoma cell line) and quantitative reverse-transcriptase PCR. We found that the UV-killed cells and their EPSs had immunological effects on RAW264.7 cells via Caco-2 cells. The RAW264.7 cells showed different cytokine production profiles when treated with UV-killed cells and EPSs. The UV-killed cells and EPSs promoted a Th1-type cellular response. Furthermore, we found that the UV-killed cells sent positive signals through Toll-like receptor (TLR) 2. Meanwhile, neither EPS sent a positive signal through TLR4 and TLR2. This evidence suggests that both UV-killed cells of the lactobacillus strains and their EPSs trigger a Th1-type immune response in a human host, with the former triggering the response via the TLRs expressed on its epithelium and the latter employing a mechanism yet to be determined, possibly involving a novel receptor that is designed to recognize specific patterns of repeating sugar in the EPSs. PMID
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In vitro cytotoxicity of Indonesian stingless bee products against human cancer cell lines
Kustiawan, Paula M.; Puthong, Songchan; Arung, Enos T.; Chanchao, Chanpen
2014-01-01
Objective To screen crude extracts of propolis, bee pollen and honey from four stingless bee species [Trigona incisa (T. incisa)], Timia apicalis, Trigona fusco-balteata and Trigona fuscibasis) native to East Kalimantan, Indonesia for cytotoxic activity against five human cancer cell lines (HepG2, SW620, ChaGo-I, KATO-III and BT474). Methods All samples were extracted with methanol, and then subpartitioned with n-hexane and ethyl acetate. Each crude extract was screened at 20 µg/mL for in vitro cytotoxicity against the cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In addition, four previously shown bioactive components from propolis (apigenin, caffeic acid phenyl ester, kaempferol and naringenin) and two chemotherapeutic drugs (doxorubicin and 5-fluorouracil) were used to evaluate the sensitivity of the cell lines. Results Overall, crude extracts from propolis and honey had higher cytotoxic activities than bee pollen, but the activity was dependent upon the extraction solvent, bee species and cell line. Propolis extracts from T. incisa and Timia apicalis showed the highest and lowest cytotoxic activity, respectively. Only the HepG2 cell line was broadly sensitive to the honey extracts. For pure compounds, doxorubicin was the most cytotoxic, the four propolis compounds the least, but the ChaGo-I cell line was sensitive to kaempferol at 10 µg/mL and KATO-III was sensitive to kaempferol and apigenin at 10 µg/mL. All pure compounds were effective against the BT474 cell line. Conclusions Propolis from T. incisa and Trigona fusco-balteata contain an in vitro cytotoxic activity against human cancer cell lines. Further study is required, including the isolation and characterization of the active antiproliferative agent(s). PMID:25183275
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In vitro cytotoxicity of Indonesian stingless bee products against human cancer cell lines.
Kustiawan, Paula M; Puthong, Songchan; Arung, Enos T; Chanchao, Chanpen
2014-07-01
To screen crude extracts of propolis, bee pollen and honey from four stingless bee species [Trigona incisa (T. incisa)], Timia apicalis, Trigona fusco-balteata and Trigona fuscibasis) native to East Kalimantan, Indonesia for cytotoxic activity against five human cancer cell lines (HepG2, SW620, ChaGo-I, KATO-III and BT474). All samples were extracted with methanol, and then subpartitioned with n-hexane and ethyl acetate. Each crude extract was screened at 20 µg/mL for in vitro cytotoxicity against the cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In addition, four previously shown bioactive components from propolis (apigenin, caffeic acid phenyl ester, kaempferol and naringenin) and two chemotherapeutic drugs (doxorubicin and 5-fluorouracil) were used to evaluate the sensitivity of the cell lines. Overall, crude extracts from propolis and honey had higher cytotoxic activities than bee pollen, but the activity was dependent upon the extraction solvent, bee species and cell line. Propolis extracts from T. incisa and Timia apicalis showed the highest and lowest cytotoxic activity, respectively. Only the HepG2 cell line was broadly sensitive to the honey extracts. For pure compounds, doxorubicin was the most cytotoxic, the four propolis compounds the least, but the ChaGo-I cell line was sensitive to kaempferol at 10 µg/mL and KATO-III was sensitive to kaempferol and apigenin at 10 µg/mL. All pure compounds were effective against the BT474 cell line. Propolis from T. incisa and Trigona fusco-balteata contain an in vitro cytotoxic activity against human cancer cell lines. Further study is required, including the isolation and characterization of the active antiproliferative agent(s).
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Povey, Jane F; O'Malley, Christopher J; Root, Tracy; Martin, Elaine B; Montague, Gary A; Feary, Marc; Trim, Carol; Lang, Dietmar A; Alldread, Richard; Racher, Andrew J; Smales, C Mark
2014-08-20
Despite many advances in the generation of high producing recombinant mammalian cell lines over the last few decades, cell line selection and development is often slowed by the inability to predict a cell line's phenotypic characteristics (e.g. growth or recombinant protein productivity) at larger scale (large volume bioreactors) using data from early cell line construction at small culture scale. Here we describe the development of an intact cell MALDI-ToF mass spectrometry fingerprinting method for mammalian cells early in the cell line construction process whereby the resulting mass spectrometry data are used to predict the phenotype of mammalian cell lines at larger culture scale using a Partial Least Squares Discriminant Analysis (PLS-DA) model. Using MALDI-ToF mass spectrometry, a library of mass spectrometry fingerprints was generated for individual cell lines at the 96 deep well plate stage of cell line development. The growth and productivity of these cell lines were evaluated in a 10L bioreactor model of Lonza's large-scale (up to 20,000L) fed-batch cell culture processes. Using the mass spectrometry information at the 96 deep well plate stage and phenotype information at the 10L bioreactor scale a PLS-DA model was developed to predict the productivity of unknown cell lines at the 10L scale based upon their MALDI-ToF fingerprint at the 96 deep well plate scale. This approach provides the basis for the very early prediction of cell lines' performance in cGMP manufacturing-scale bioreactors and the foundation for methods and models for predicting other mammalian cell phenotypes from rapid, intact-cell mass spectrometry based measurements. Copyright © 2014 Elsevier B.V. All rights reserved.
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Cell-based capacitance sensor for analysis of EGFR expression on cell membrane
NASA Astrophysics Data System (ADS)
Shin, Dong-Myeong; Shin, Yong-Cheol; Ha, Ji Hye; Lee, Jong-Ho; Han, Dong-Wook; Kim, Jong-Man; Kim, Hyung Kook; Hwang, Yoon-Hwae
2013-02-01
Cancer cells have many kinds of cancer biomarkers. Among them, the epidermal growth factor (EGF) receptors can show a possibility for a cancer marker because the over-expression of EGF receptor is related with fibrous, colorectal, cervical and gastric tumorigenesis. We fabricated the capacitance sensor with a gap area of 50 μm × 200 μm by using photolithography and lift-off method. Using the capacitance sensor, we investigated the time dependent capacitance changes of different kinds of fibrous cells, such as HT1080 fibrosarcoma, L-929 fibroblast cell line and nHDF dermal fibroblast primary cell. We found that when we put the EGF, the capacitance decreased due to the immobilization of EGF to EGF receptor on the cell membrane. The quantitative determination of EGF receptor level for various fibrous cells was carried out and the results showed good correlation with conventional method. Based on our results, we suggest that the capacitance sensor can measure the expression level of the EGF receptor on cell membrane and be a good candidate as a cancer diagnosis.
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Tima, Singkome; Ichikawa, Hideki; Ampasavate, Chadarat; Okonogi, Siriporn; Anuchapreeda, Songyot
2014-04-25
Leukemia is a hematologic malignancy with a frequent incidence and high mortality rate. Previous studies have shown that the FLT3 gene is overexpressed in leukemic blast cells, especially in acute myeloid leukemia. In this study, a commercially available curcuminoid mixture (1), pure curcumin (2), pure demethoxycurcumin (3), and pure bisdemethoxycurcumin (4) were investigated for their inhibitory effects on cell growth, FLT3 expression, and cell cycle progression in an FLT3-overexpressing EoL-1 leukemic cell line using an MTT assay, Western blotting, and flow cytometry, respectively. The mixture (1) and compounds 2-4 demonstrated cytotoxic effects with IC50 values ranging from 6.5 to 22.5 μM. A significant decrease in FLT3 protein levels was found after curcuminoid treatment with IC20 doses, especially with mixture 1 and compound 2. In addition, mixture 1 and curcumin (2) showed activity on cell cycle arrest at the G0/G1 phase and decreased the FLT3 and STAT5A protein levels in a dose-dependent manner. Compound 2 demonstrated the greatest potential for inhibiting cell growth, cell cycle progression, and FLT3 expression in EoL-1 cells. This investigation has provided new findings regarding the effect of turmeric curcuminoids on FLT3 expression in leukemic cells.
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Code of Federal Regulations, 2010 CFR
2010-01-01
... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.58 Exemptions. (a...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and..., RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Administrative Body § 929.30 Powers. The...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.54 Withholding. (a...
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Enhanced adherence of mouse fibroblast and vascular cells to plasma modified polyethylene.
Reznickova, Alena; Novotna, Zdenka; Kolska, Zdenka; Kasalkova, Nikola Slepickova; Rimpelova, Silvie; Svorcik, Vaclav
2015-01-01
Since the last decade, tissue engineering has shown a sensational promise in providing more viable alternatives to surgical procedures for harvested tissues, implants and prostheses. Biomedical polymers, such as low-density polyethylene (LDPE), high-density polyethylene (HDPE) and ultra-high molecular weight polyethylene (UHMWPE), were activated by Ar plasma discharge. Degradation of polymer chains was examined by determination of the thickness of ablated layer. The amount of an ablated polymer layer was measured by gravimetry. Contact angle, measured by goniometry, was studied as a function of plasma exposure and post-exposure aging times. Chemical structure of modified polymers was characterized by angle resolved X-ray photoelectron spectroscopy. Surface chemistry and polarity of the samples were investigated by electrokinetic analysis. Changes in surface morphology were followed using atomic force microscopy. Cytocompatibility of plasma activated polyethylene foils was studied using two distinct model cell lines; VSMCs (vascular smooth muscle cells) as a model for vascular graft testing and connective tissue cells L929 (mouse fibroblasts) approved for standardized material cytotoxicity testing. Specifically, the cell number, morphology, and metabolic activity of the adhered and proliferated cells on the polyethylene matrices were studied in vitro. It was found that the plasma treatment caused ablation of the polymers, resulting in dramatic changes in their surface morphology and roughness. ARXPS and electrokinetic measurements revealed oxidation of the polymer surface. It was found that plasma activation has a positive effect on the adhesion and proliferation of VSMCs and L929 cells. Copyright © 2015 Elsevier B.V. All rights reserved.
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Yang, Lingyun; Huang, Feng; Mei, Jiandong; Wang, Xun; Zhang, Qiuyang; Wang, Hongjing; Xi, Mingrong; You, Zongbing
2017-02-01
Estrogen is a well-known oncogenic driver in endometrial (ECs) and breast cancers (BCs). Programmed cell death protein 1 (PD-1) and its ligands PD-1 Ligand 1 (PD-L1) and PD-L2 have been shown to mediate immune evasion of the tumor cells. The purpose of the present study was to assess the effects of estrogen on PD-L1 and PD-L2 expression in EC and BC cell lines. 17β-Estradiol (E2)-induced expression of PD-L1 and PD-L2 and possible signaling pathway were investigated in EC and BC cells. Coculture of T cells and cancer cells with E2 stimulation was performed to assess the functions of T cells. We found that E2 increased expression of PD-L1, but not PD-L2, protein via activation of phosphoinositide 3-kinase (PI3K)/Akt pathway in Ishikawa and Michigan Cancer Foundation-7 (MCF-7) cells. Phosphoinositide 3-kinase and Akt inhibitors could block E2's effects. 17β-Estradiol did not increase PD-L1 mRNA transcription, but stabilized PD-L1 mRNA. 17β-Estradiol's effects were only observed in estrogen receptor α (ERα)-positive Ishikawa and MCF-7 cells, but not in ERα-negative MDA-MB-231 cells. Coculture of Ishikawa or MCF-7 cells with T cells inhibited expression of interferon-γ and interleukin-2 and increased BCL-2-interacting mediator of cell death expression in the presence of E2. This study provides the first evidence that estrogen upregulates PD-L1 protein expression in ERα-positive EC and BC cells to suppress immune functions of T cells in the tumor microenvironment, demonstrating a new mechanism of how estrogen drives cancer progression.
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Cell lines derived from the squash bug, Anasa tristis (Coreidae: Hemiptera).
Goodman, Cynthia L; Ringbauer, Joseph A; Li, Yao-Fa; Lincoln, Tamra Reall; Stanley, David
2017-05-01
The squash bug, Anasa tristis, is a pest of cucurbits that exerts direct damage on crops and is a vector of plant pathogens. We established cell lines from this insect to serve as tools for basic biology, including virology and immunology, as well as applied studies, such as insecticide development programs. We initiated 15 cell cultures, using nine media or combinations of media. The media yielding the best results were a modification of Kimura's medium and a combination of two commercially available cell culture media (EX-CELL 420 and L15). We designated the two cell lines as BCIRL-AtE-CLG11 and BCIRL-AtE-CLG15. From the AtE-CLG15 line, we isolated two sub-lines, A and B. Of these, the most consistently replicating line was AtE-CLG15A. We determined the doubling time of this line (190 h) and its mean cell diameter (14.5 ± 0.7 μm). We characterized the AtE-CLG15A line using DAF-PCR. The BCIRL-AtE-CLG15A cell line is now available for researchers world-wide.
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Goulart, T L S; Boyle, R T; Souza, M M
2015-10-01
The present study aimed to ascertain the cytotoxicity of pesticides commonly used in rice cultivation, through in vitro assays employing the ZF-L cell line. The in vitro analyses investigated three cellular targets (cell membrane integrity, mitochondrial activity and lysosomal stability) in cells exposed to concentrations of Roundup Transorb® (67.7 μg L(-1), 135.4 μg L(-1) and 270.8 μg L(-1)), Furadan 350 SC® (0.1 μg L(-1), 0.05 μg L(-1) and 0.02 μg L(-1)). We also tested these products in combination. We analyzed the defensive capacity of the cells by measuring the activity of xenobiotic extruder proteins, as well as the expression of these same proteins. Cytotoxic effects of both pesticides were observed individually, as well as with the mixture of both products; including an inhibitory effect on the activity of xenobiotic extrusion. When exposed to the insecticide Furadan, and also the mixture of Furadan and Roundup, there was an increase in the expression of P glycoproteins (P-gps). There was also a negative correlation with cytotoxicity, mainly exhibited by mitochondrial activity and lysosomal integrity, but also with respect to the activity of P-gps. We observed that concentrations below those allowed by law were toxic regarding all parameters tested in this study, with the exception of mitochondrial function. Taken together, our results suggest that toxicity may be due to the surfactants present in the commercial formulations. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.105 Reporting. (a) Each report... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and..., RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Reports and Records § 929.62 Reports. (a...
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Control of cell proliferation by a porous chitosan scaffold with multiple releasing capabilities
Cai, Shu-Jyun; Li, Ching-Wen; Weihs, Daphne; Wang, Gou-Jen
2017-01-01
Abstract The aim of this study was to develop a porous chitosan scaffold with long-acting drug release as an artificial dressing to promote skin wound healing. The dressing was fabricated by pre-freezing at different temperatures (−20 and −80 °C) for different periods of time, followed by freeze-drying to form porous chitosan scaffolds with different pore sizes. The chitosan scaffolds were then used to investigate the effect of the controlled release of fibroblast growth factor-basic (bFGF) and transforming growth factor-β1 (TGFβ1) on mouse fibroblast cells (L929) and bovine carotid endothelial cells (BEC). The biocompatibility of the prepared chitosan scaffold was confirmed with WST-1 proliferation and viability assay, which demonstrated that the material is suitable for cell growth. The results of this study show that the pore sizes of the porous scaffolds prepared by freeze-drying can change depending on the pre-freezing temperature and time via the formation of ice crystals. In this study, the scaffolds with the largest pore size were found to be 153 ± 32 μm and scaffolds with the smallest pores to be 34 ± 9 μm. Through cell culture analysis, it was found that the concentration that increased proliferation of L929 cells for bFGF was 0.005 to 0.1 ng/mL, and the concentration for TGFβ1 was 0.005 to 1 ng/mL. The cell culture of the chitosan scaffold and growth factors shows that 3.75 ng of bFGF in scaffolds with pore sizes of 153 ± 32 μm can promote L929 cell proliferation, while 400 pg of TGFβ1 in scaffolds with pore size of 34 ± 9 μm can enhance the proliferation of L929 cells, but also inhibit BEC proliferation. It is proposed that the prepared chitosan scaffolds can form a multi-drug (bFGF and TGFβ1) release dressing that has the ability to control wound healing via regulating the proliferation of different cell types. PMID:29230255
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A common deletion in two gamma ray induced rat pulmonary tumor cell lines.
Van Klaveren, P; De Bruijne, J; Van der Winden, H; Kal, H B; Bentvelzen, P
1994-01-01
Subtraction hybridization was performed on normal WAG/Rij rat DNA with DNA from a syngeneic Ir-192 induced pulmonary tumor cell line L37. The residual DNA was amplified by means of sequence-independent PCR. This procedure yielded a sequence, of which multiple copies are present in normal rat DNA. In the tumor line L37 two restriction fragments hybridizing with this repeat sequence are lacking. In another Ir-192 induced pulmonary tumor line, L33, one of these fragments was also lacking. This indicates a common deletion in the two tumor lines.
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CellLineNavigator: a workbench for cancer cell line analysis
Krupp, Markus; Itzel, Timo; Maass, Thorsten; Hildebrandt, Andreas; Galle, Peter R.; Teufel, Andreas
2013-01-01
The CellLineNavigator database, freely available at http://www.medicalgenomics.org/celllinenavigator, is a web-based workbench for large scale comparisons of a large collection of diverse cell lines. It aims to support experimental design in the fields of genomics, systems biology and translational biomedical research. Currently, this compendium holds genome wide expression profiles of 317 different cancer cell lines, categorized into 57 different pathological states and 28 individual tissues. To enlarge the scope of CellLineNavigator, the database was furthermore closely linked to commonly used bioinformatics databases and knowledge repositories. To ensure easy data access and search ability, a simple data and an intuitive querying interface were implemented. It allows the user to explore and filter gene expression, focusing on pathological or physiological conditions. For a more complex search, the advanced query interface may be used to query for (i) differentially expressed genes; (ii) pathological or physiological conditions; or (iii) gene names or functional attributes, such as Kyoto Encyclopaedia of Genes and Genomes pathway maps. These queries may also be combined. Finally, CellLineNavigator allows additional advanced analysis of differentially regulated genes by a direct link to the Database for Annotation, Visualization and Integrated Discovery (DAVID) Bioinformatics Resources. PMID:23118487
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Leong, Ooi Kheng; Muhammad, Tengku Sifzizul Tengku; Sulaiman, Shaida Fariza
2011-01-01
Physalis minima L. is reputed for having anticancer property. In this study, the chloroform extract of this plant exhibited remarkable cytotoxic activities on NCI-H23 (human lung adenocarcinoma) cell line at dose- and time-dependent manners (after 24, 48 and 72 h of incubation). Analysis of cell-death mechanism demonstrated that the extract exerted apoptotic programed cell death in NCI-H23 cells with typical DNA fragmentation, which is a biochemical hallmark of apoptosis. Morphological observation using transmission electron microscope (TEM) also displayed apoptotic characteristics in the treated cells, including clumping and margination of chromatins, followed by convolution of the nuclear and budding of the cells to produce membrane-bound apoptotic bodies. Different stages of apoptotic programed cell death as well as phosphatidylserine externalization were confirmed using annexin V and propidium iodide staining. Furthermore, acute exposure to the extract produced a significant regulation of c-myc, caspase-3 and p53 mRNA expression in this cell line. Due to its apoptotic effect on NCI-H23 cells, it is strongly suggested that the extract could be further developed as an anticancer drug. PMID:19541726
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Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.142 Reserve. (a) It is necessary and... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements...
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Code of Federal Regulations, 2010 CFR
2010-01-01
... marketing order districts as designated in § 929.20(c). (2) Six qualified persons for members and four... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... nominations in accordance with this section. (c) Whenever any cooperative marketing organization handles more...
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Code of Federal Regulations, 2012 CFR
2012-01-01
... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... in § 929.52(a), each handler shall withhold from handling a portion of the cranberries acquired... volume of marketable cranberries acquired. Such withholding requirements shall not apply to any lot of...
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Code of Federal Regulations, 2014 CFR
2014-01-01
... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... in § 929.52(a), each handler shall withhold from handling a portion of the cranberries acquired... volume of marketable cranberries acquired. Such withholding requirements shall not apply to any lot of...
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Code of Federal Regulations, 2013 CFR
2013-01-01
... AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... in § 929.52(a), each handler shall withhold from handling a portion of the cranberries acquired... volume of marketable cranberries acquired. Such withholding requirements shall not apply to any lot of...
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Code of Federal Regulations, 2011 CFR
2011-01-01
... and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... in § 929.52(a), each handler shall withhold from handling a portion of the cranberries acquired... volume of marketable cranberries acquired. Such withholding requirements shall not apply to any lot of...
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7 CFR 58.929 - Frequency of sampling for quality control.
Code of Federal Regulations, 2011 CFR
2011-01-01
... 7 Agriculture 3 2011-01-01 2011-01-01 false Frequency of sampling for quality control. 58.929... Service 1 Operations and Operating Procedures § 58.929 Frequency of sampling for quality control. (a... to control composition. On continuous production runs, enough samples shall be taken throughout the...
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Gnirss, Kerstin; Kühl, Annika; Karsten, Christina; Glowacka, Ilona; Bertram, Stephanie; Kaup, Franziska; Hofmann, Heike; Pöhlmann, Stefan
2012-03-01
Ebola (EBOV) and Marburg virus (MARV) cause severe hemorrhagic fever. The host cell proteases cathepsin B and L activate the Zaire ebolavirus glycoprotein (GP) for cellular entry and constitute potential targets for antiviral intervention. However, it is unclear if different EBOV species and MARV equally depend on cathepsin B/L activity for infection of cell lines and macrophages, important viral target cells. Here, we show that cathepsin B/L inhibitors markedly reduce 293T cell infection driven by the GPs of all EBOV species, independent of the type II transmembrane serine protease TMPRSS2, which cleaved but failed to activate EBOV-GPs. Similarly, a cathepsin B/L inhibitor blocked macrophage infection mediated by different EBOV-GPs. In contrast, MARV-GP-driven entry exhibited little dependence on cathepsin B/L activity. Still, MARV-GP-mediated entry was efficiently blocked by leupeptin. These results suggest that cathepsins B/L promote entry of EBOV while MARV might employ so far unidentified proteases for GP activation. Copyright © 2011 Elsevier Inc. All rights reserved.
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Reprogramming somatic cells into iPS cells activates LINE-1 retroelement mobility
Wissing, Silke; Muñoz-Lopez, Martin; Macia, Angela; Yang, Zhiyuan; Montano, Mauricio; Collins, William; Garcia-Perez, Jose Luis; Moran, John V.; Greene, Warner C.
2012-01-01
Long interspersed element-1 (LINE-1 or L1) retrotransposons account for nearly 17% of human genomic DNA and represent a major evolutionary force that has reshaped the structure and function of the human genome. However, questions remain concerning both the frequency and the developmental timing of L1 retrotransposition in vivo and whether the mobility of these retroelements commonly results in insertional and post-insertional mechanisms of genomic injury. Cells exhibiting high rates of L1 retrotransposition might be especially at risk for such injury. We assessed L1 mRNA expression and L1 retrotransposition in two biologically relevant cell types, human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs), as well as in control parental human dermal fibroblasts (HDFs). Full-length L1 mRNA and the L1 open reading frame 1-encoded protein (ORF1p) were readily detected in hESCs and iPSCs, but not in HDFs. Sequencing analysis proved the expression of human-specific L1 element mRNAs in iPSCs. Bisulfite sequencing revealed that the increased L1 expression observed in iPSCs correlates with an overall decrease in CpG methylation in the L1 promoter region. Finally, retrotransposition of an engineered human L1 element was ∼10-fold more efficient in iPSCs than in parental HDFs. These findings indicate that somatic cell reprogramming is associated with marked increases in L1 expression and perhaps increases in endogenous L1 retrotransposition, which could potentially impact the genomic integrity of the resultant iPSCs. PMID:21989055
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22 CFR 92.9 - Refusals of requests for notarial services.
Code of Federal Regulations, 2012 CFR
2012-04-01
... 22 Foreign Relations 1 2012-04-01 2012-04-01 false Refusals of requests for notarial services. 92.9 Section 92.9 Foreign Relations DEPARTMENT OF STATE LEGAL AND RELATED SERVICES NOTARIAL AND RELATED... services should be refused only after the most careful deliberation. [22 FR 10858, Dec. 27, 1957, as...
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Cheng, T-C; Lai, Y-S; Lin, I-Y; Wu, C-P; Chang, S-L; Chen, T-I; Su, M-S
2010-02-01
Establishment and characterization of two cobia, Rachycentron canadum, cell lines derived from cobia brain (CB) and cobia fin (CF) are described. Caudal fin and brain from juvenile cobia were dissociated for 30 and 10 min, respectively, in phosphate-buffered saline containing 0.25% trypsin at 25 degrees C. The optimal culture condition for both dissociated cells (primary cell culture) was at 28 degrees C in Leibovitz-15 medium containing 10% foetal bovine serum. The cells have been sub-cultured at a ratio of 1:2 for more than 160 passages over a period of 3 years. Origin of the cultured cells was verified by comparison of their sequences of mitochondrial cytochrome oxidase subunit I genes (cox I) with the cox 1 sequence from cobia muscle tissue. The cell lines showed polyploidy. No mycoplasma contamination was detected. Susceptibility to grouper iridovirus was observed for the CB cell line but not the CF cell line. Both cell lines expressed green fluorescent protein after being transfected with green fluorescent reporter gene driven by the cytomegalovirus promoter.
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Mansard, Sandrine; Papon, Janine; Moreau, Marie-France; Miot-Noirault, Elisabeth; Labarre, Pierre; Bayle, Martine; Veyre, Annie; Madelmont, Jean-Claude; Moins, Nicole
2005-07-01
N-(2-diethylaminoethyl)-2-iodobenzamide (BZA(2)) has been singled out as the most efficacious melanoma scintigraphy imaging agent. Our work was designed to assess the mechanisms of the specific affinity of the radioiodinated iodobenzamide for melanoma tissue. We studied the cellular uptake and retention of [(125)I]-BZA(2) on various cell lines. In vitro, cellular [(125)I]-BZA(2) uptake was related to the pigmentation status of the cells: higher in pigmented melanoma cell lines (M4 Beu, IPC 227, B 16) than in a nonpigmented one (M3 Dau) and nonmelanoma cell lines (MCF 7 and L 929). Two mechanisms were assessed: binding of the tracer to melanin or to sigma receptors of melanoma cells. First, the uptake of [(125)I]-BZA(2) after melanogenesis stimulation by alpha-melanocyte-stimulating hormone and l-tyrosine increased in the B 16 melanoma cell line both in vitro and in vivo according to melanin concentration. Moreover, the binding of [(125)I]-BZA(2) to synthetic melanin was dependent on melanin concentration and could be saturated. Second, no competition was evidenced on M4 Beu cells between [(125)I]-BZA(2) and haloperidol, a sigma ligand, at concentrations < or =10(-6) M. We show that the specificity and sensibility of BZA(2) as a melanoma scintigraphic imaging agent are mostly due to interactions with melanic pigments.
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NASA Astrophysics Data System (ADS)
Sinha, Rileen; Winer, Andrew G.; Chevinsky, Michael; Jakubowski, Christopher; Chen, Ying-Bei; Dong, Yiyu; Tickoo, Satish K.; Reuter, Victor E.; Russo, Paul; Coleman, Jonathan A.; Sander, Chris; Hsieh, James J.; Hakimi, A. Ari
2017-05-01
The utility of cancer cell lines is affected by the similarity to endogenous tumour cells. Here we compare genomic data from 65 kidney-derived cell lines from the Cancer Cell Line Encyclopedia and the COSMIC Cell Lines Project to three renal cancer subtypes from The Cancer Genome Atlas: clear cell renal cell carcinoma (ccRCC, also known as kidney renal clear cell carcinoma), papillary (pRCC, also known as kidney papillary) and chromophobe (chRCC, also known as kidney chromophobe) renal cell carcinoma. Clustering copy number alterations shows that most cell lines resemble ccRCC, a few (including some often used as models of ccRCC) resemble pRCC, and none resemble chRCC. Human ccRCC tumours clustering with cell lines display clinical and genomic features of more aggressive disease, suggesting that cell lines best represent aggressive tumours. We stratify mutations and copy number alterations for important kidney cancer genes by the consistency between databases, and classify cell lines into established gene expression-based indolent and aggressive subtypes. Our results could aid investigators in analysing appropriate renal cancer cell lines.
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Development and characterization of a cell line WAF from freshwater shark Wallago attu.
Dubey, Akhilesh; Goswami, Mukunda; Yadav, Kamalendra; Sharma, Bhagwati S
2014-02-01
A new epithelial cell line, WAF was developed from caudal fin of freshwater shark, Wallago attu. The cell line was optimally maintained at 28 °C in Leibovitz-15 (L-15) medium supplemented with 20 % fetal bovine serum. The cell line was characterized by various cytogenetic and molecular markers. The cytogenetic analysis revealed a diploid count of 86 chromosomes at different passages. The origin of the cell lines was confirmed by the amplification of 547 and 654 bp sequences of 16S rRNA and cytochrome oxidase subunit I genes of mitochondrial DNA, respectively. WAF cells were characterized for their growth characteristics at different temperature and serum concentration. Epithelial morphology of the cell line was confirmed using immunocytochemistry. Further cell plating efficiency, transfection efficiency and viability of cryopreserved WAF cells was also determined. Cytotoxicity and genotoxicity assessment of cadmium salts on WAF cells by MTT, NR and comet assay illustrated the utility of this cell line as an in vitro model for aquatic toxicological studies. The cell line will be further useful for studying oxidative stress markers against aquatic pollutants.
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Akay, Canan; Tanış, Merve Ç; Sevim, Handan
2017-10-13
The aim of this study was to evaluate the cytotoxic effects of 9 different soft denture liners on the viability of L-929 mouse fibroblast cells at different incubation periods by storing them in artificial saliva (AS) with different pH levels. 96 disk samples from each lining material were prepared and divided into 4 groups: GI: No treatment; GII: Stored in artificial saliva with pH 3 for 21 days; Group III: Stored in artificial saliva with pH 7 for 21 days; and Group IV: Stored in artificial saliva with pH 14 for 21 days. The cytotoxicity of the extracts to cultured mouse fibroblasts (L-929) was measured by MTT (tetrazolium salt 3-[4,5-dimethylthiazol-2-yl]-2,5-dipHnyltetrazolium bromide) assay. Data were analyzed using 1-way analysis of variation (ANOVA). It was found that for the pH 3 values of New Truliner, Trusoft, Mollosil Plus, Dentusil, TDV, and HydroCast®; for the pH 7 values of Ufi Gel P and Elite plus; and for the pH 14 values of HydroCast®, there was a noncytotoxic effect during both the 24-hour and 48-hour incubation periods. In the control group 48-hour incubation period, HydroCast®, TDV, Mollosil, 24-hour incubation period Elite plus, for pH 3 values; Elite Plus 24-hour incubation period, for pH 7 values Trusoft 48-hour incubation period there was a moderately cytotoxic effect. This in vitro study revealed that storage in artificial saliva with different pH levels can affect the cytotoxicity of soft lining materials.
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7 CFR 929.102 - Procedure to determine quantity of screened cranberries in unscreened lots.
Code of Federal Regulations, 2012 CFR
2012-01-01
... cranberries in unscreened lots. 929.102 Section 929.102 Agriculture Regulations of the Department of..., Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND... NEW YORK Rules and Regulations § 929.102 Procedure to determine quantity of screened cranberries in...
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7 CFR 929.102 - Procedure to determine quantity of screened cranberries in unscreened lots.
Code of Federal Regulations, 2014 CFR
2014-01-01
... cranberries in unscreened lots. 929.102 Section 929.102 Agriculture Regulations of the Department of..., NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND... NEW YORK Rules and Regulations § 929.102 Procedure to determine quantity of screened cranberries in...
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7 CFR 929.102 - Procedure to determine quantity of screened cranberries in unscreened lots.
Code of Federal Regulations, 2013 CFR
2013-01-01
... cranberries in unscreened lots. 929.102 Section 929.102 Agriculture Regulations of the Department of..., NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND... NEW YORK Rules and Regulations § 929.102 Procedure to determine quantity of screened cranberries in...
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7 CFR 929.102 - Procedure to determine quantity of screened cranberries in unscreened lots.
Code of Federal Regulations, 2011 CFR
2011-01-01
... cranberries in unscreened lots. 929.102 Section 929.102 Agriculture Regulations of the Department of..., Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF MASSACHUSETTS, RHODE ISLAND... NEW YORK Rules and Regulations § 929.102 Procedure to determine quantity of screened cranberries in...
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33 CFR 117.929 - Durham Creek.
Code of Federal Regulations, 2011 CFR
2011-07-01
... OPERATION REGULATIONS Specific Requirements South Carolina § 117.929 Durham Creek. The removable span of the... Charleston of an emergency in the Bushy Park Reservoir, the span shall be removed as soon as possible to...
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33 CFR 117.929 - Durham Creek.
Code of Federal Regulations, 2010 CFR
2010-07-01
... OPERATION REGULATIONS Specific Requirements South Carolina § 117.929 Durham Creek. The removable span of the... Charleston of an emergency in the Bushy Park Reservoir, the span shall be removed as soon as possible to...
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Yang, Lingyun; Huang, Feng; Mei, Jiandong; Wang, Xun; Zhang, Qiuyang; Wang, Hongjing; Xi, Mingrong; You, Zongbing
2016-01-01
Objective Estrogen is a well-known oncogenic driver in endometrial and breast cancers. Programmed cell death protein 1 (PD-1) and its ligands PD-L1 and PD-L2 have been shown to mediate immune evasion of the tumor cells. The purpose of the present study was to assess the effects of estrogen on PD-L1 and PD-L2 expression in endometrial and breast cancer cell lines. Methods 17β-estradiol (E2) -induced expression of PD-L1 and PD-L2 and possible signaling pathway were investigated in endometrial and breast cancer cells. Co-culture of T cells and cancer cells with E2 stimulation was performed to assess the functions of T cells. Results We found that 17β-estradiol (E2) increased expression of PD-L1, but not PD-L2 protein via activation of phosphoinositide 3-kinase (PI3K)/Akt pathway in Ishikawa and MCF-7 cells. PI3K and Akt inhibitors could block E2's effects. E2 did not increase PD-L1 mRNA transcription, but stabilized PD-L1 mRNA. E2's effects were only observed in estrogen receptor α (ERα) -positive Ishikawa and MCF-7 cells, but not in ERα-negative MDA-MB-231 cells. Co-culture of Ishikawa or MCF-7 cells with T cells inhibited expression of interferon-γ and interleukin-2 and increased Bim expression in the presence of E2. Conclusion This study provides the first evidence that estrogen up-regulates PD-L1 protein expression in ERα-positive endometrial and breast cancer cells to suppress immune functions of T cells in the tumor microenvironment, demonstrating a new mechanism of how estrogen drives cancer progression. PMID:27870715
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Zainal Ariffin, Shahrul Hisham; Wan Omar, Wan Haifa Haryani; Zainal Ariffin, Zaidah; Safian, Muhd Fauzi; Senafi, Sahidan; Megat Abdul Wahab, Rohaya
2009-01-01
Background Piper sarmentosum, locally known as kaduk is belonging to the family of Piperaceae. It is our interest to evaluate their effect on human hepatoma cell line (HepG2) for the potential of anticarcinogenic activity. Results The anticarcinogenic activity of an ethanolic extract from Piper sarmentosum in HepG2 and non-malignant Chang's liver cell lines has been previously determined using (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) (MTT) assays, where the IC50 value was used as a parameter for cytotoxicity. The ethanolic extract that showed anticarcinogenic properties in HepG2 cells had an IC50 of 12.5 μg mL-1, while IC50 values in the non-malignant Chang's liver cell line were greater than 30 μg mL-1. Apoptotic morphological changes in HepG2 cells were observed using an inverted microscope and showed chromatin condensation, cell shrinkage and apoptotic bodies following May-Grunwald-Giemsa's staining. The percentage of apoptotic cells in the overall population (apoptotic index) showed a continuously significant increase (p < 0.05) in 12.5 μg mL-1 ethanolic extract-treated cells at 24, 48 and 72 hours compared to controls (untreated cells). Following acridine orange and ethidium bromide staining, treatment with 10, 12 and 14 μg mL-1 of ethanolic extracts caused typical apoptotic morphological changes in HepG2 cells. Molecular analysis of DNA fragmentation was used to examine intrinsic apoptosis induced by the ethanolic extracts. These results showed a typical intrinsic apoptotic characterisation, which included fragmentation of nuclear DNA in ethanolic extract-treated HepG2 cells. However, the non-malignant Chang's liver cell line produced no DNA fragmentation. In addition, the DNA genome was similarly intact for both the untreated non-malignant Chang's liver and HepG2 cell lines. Conclusion Therefore, our results suggest that the ethanolic extract from P. sarmentosum induced anticarcinogenic activity through an intrinsic apoptosis
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Characteristics of cell lines established from human colorectal carcinoma.
Park, J G; Oie, H K; Sugarbaker, P H; Henslee, J G; Chen, T R; Johnson, B E; Gazdar, A
1987-12-15
We have characterized 14 human colorectal carcinoma cell lines established from primary and metastatic sites by us during the years 1982 to 1985. Five lines were established in fully defined ACL-4 medium and 9 in serum supplemented R10 medium. However, after establishment, cultures could be grown interchangeably in either medium. The lines grew as floating cell aggregates in ACL-4 medium, while most demonstrated substrate adherence in R10 medium. The lines had relatively long doubling times and low cloning efficiencies. Twelve were tumorigenic in athymic nude mice when injected s.c., and two grew i.p. as well. Based on culture, xenograft, and ultrastructural morphologies, the 14 lines could be subtyped as follows: 4 were well differentiated; 5 were moderately differentiated; 4 were poorly differentiated; and 1 was a mucinous carcinoma. Membrane associated antigens characteristic for gastrointestinal cells (carcinoembryonic antigen, CA 19-9, and TAG-72 antigens) were expressed by 50-71% of the lines. Lines expressing carcinoembryonic antigen and CA 19-9 actively secreted these antigens into the supernatant fluids while TAG-72 antigen was not secreted. Surprisingly, 5 of 7 of the original tumor samples tested and 13 of 14 cultured lines expressed L-dopa decarboxylase activity, which is a characteristic enzyme marker of neuroendocrine cells and tumors. In addition, one poorly differentiated cell line contained dense core granules, characteristic of endocrine secretion. Preliminary cytogenetic analyses indicated that 9 of 11 lines examined contained double minute chromosomes. In addition, 3 of the 9 lines with double minutes also had homogeneously staining regions. These findings indicate a high incidence of amplification of one or more as yet unidentified genes.
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Liu, Jia; Zhang, Wei; Jing, Hao; Popovich, David G
2010-04-01
Bog bilberry (Vaccinium uliginosum L.) is a blue-pigmented edible berry related to bilberry (Vaccinium myrtillus L.) and the common blueberry (Vaccinium corymbosum). The objective of this study was to investigate the effect of a bog bilberry anthocyanin extract (BBAE) on cell growth, membrane permeability, and cell cycle of 2 malignant cancer cell lines, Caco-2 and Hep-G2, and a nonmalignant murine 3T3-L1 cell line. BBAE contained 3 identified anthocyanins. The most abundant anthocyanin was cyanidin-3-glucoside (140.9 +/- 2.6 microg/mg of dry weight), followed by malvidin-3-glucoside (10.3 +/- 0.3 microg/mg) and malvidin-3-galactoside (8.1 +/- 0.4 microg/mg). Hep-G2 LC50 was calculated to be 0.563 +/- 0.04 mg/mL, Caco-2 LC50 was 0.390 +/- 0.30 mg/mL and 0.214 +/- 0.02 mg/mL for 3T3-L1 cells. LDH release, a marker of membrane permeability, was significantly increased in Hep-G2 cells and Caco-2 cells after 48 and 72 h compared to 24 h. The increase was 21% at 48 h and 57% at 72 h in Caco-2 cells and 66% and 139% in Hep-G2 cells compared to 24 h. However, 3T3-L1 cells showed an unexpected significant lower LDH activity (P < or = 0.05) after 72 h of exposure corresponding to a 21% reduction in LDH release. BBAE treatment increased sub-G1 in all 3 cell lines without influencing cells in the G2/M phase. BBAE treatment reduced the growth and increased the accumulation of sub-G1 cells in 2 malignant and 1 nonmalignant cell line; however, the effect on membrane permeability differs considerably between the malignant and nonmalignant cells and may in part be due to differences in cellular membrane composition.
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Jung, Young Ho; Lee, Doh Young; Cha, Wonjae; Kim, Bo Hae; Sung, Myung-Whun; Kim, Kwang Hyun; Ahn, Soon-Hyun
2016-10-01
A tumorigenic cell line (BHP10-3M) derived from nontumorigenic papillary thyroid carcinoma (PTC) cells (BHP10-3) having rearranged during transfection (RET)/PTC1 gene rearrangement might have a higher expression of CXCR4, either quantitatively or functionally. The authors also postulated that CXCR4-mediated invasion or tumorigenesis could be blocked by CXCR4 antagonists, including AMD3100. The expression of CXCR4 in BHP10-3 and BHP10-3M cells was assessed using immunoblot analysis, flow cytometry, and quantitative reverse-transcriptase polymerase chain reaction (RT-PCR). The effect of AMD3100 on BHP10-3 and BHP10-3M cell lines was evaluated using cell proliferation assay, invasion assay, and tumor growth experiment in nude mice. Immunoblotting, flow cytometry, and quantitative RT-PCR proved that BHP10-3M cells expressed a higher level of CXCR4 than BHP10-3 cells. Although blocking CXCR4 with AMD3100 did not suppress cell proliferation in both cell lines from 1 ng/mL to 100 ng/mL concentration, AMD3100 suppressed invasion of BHP10-3M cells in vitro in a dose-dependent manner. At higher concentrations from 10(3) ng/mL to 10(5) ng/mL, the proliferation of BHP10-3M cells was inhibited more strongly by AMD3100 than that of BHP10-3 cells. Intraperitoneal injection of AMD3100 inhibited tumor formation by BHP10-3M cells in the thyroid of nude mice. A tumorigenic cell line (BHP10-3M) of PTC showed higher expression of CXCR4 quantitatively and functionally than a nontumorigenic cell line (BHP10-3). The CXCR4 antagonist (AMD3100) showed a significant antitumor effect on the tumorigenic cell line of PTC BHP10-3 cells both in vitro and in vivo. CXCR4 antagonist can be expected to have an adjuvant role in the management of PTC. © 2016 Wiley Periodicals, Inc. Head Neck, 2016 © 2016 Wiley Periodicals, Inc. Head Neck 38: First-1486, 2016. © 2016 Wiley Periodicals, Inc.
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7 CFR 929.160 - Public member eligibility requirements and nomination procedures.
Code of Federal Regulations, 2010 CFR
2010-01-01
..., MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations... committee activities regularly and to familiarize themselves with the background and economies of the... procedures. 929.160 Section 929.160 Agriculture Regulations of the Department of Agriculture (Continued...
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Corral-Vázquez, C; Aguilar-Quesada, R; Catalina, P; Lucena-Aguilar, G; Ligero, G; Miranda, B; Carrillo-Ávila, J A
2017-06-01
Establishment of continuous cell lines from human normal and tumor tissues is an extended and useful methodology for molecular characterization of cancer pathophysiology and drug development in research laboratories. The exchange of these cell lines between different labs is a common practice that can compromise assays reliability due to contamination with microorganism such as mycoplasma or cells from different flasks that compromise experiment reproducibility and reliability. Great proportions of cell lines are contaminated with mycoplasma and/or are replaced by cells derived for a different origin during processing or distribution process. The scientific community has underestimated this problem and thousand of research experiment has been done with cell lines that are incorrectly identified and wrong scientific conclusions have been published. Regular contamination and authentication tests are necessary in order to avoid negative consequences of widespread misidentified and contaminated cell lines. Cell banks generate, store and distribute cell lines for research, being mandatory a consistent and continuous quality program. Methods implementation for guaranteeing both, the absence of mycoplasma and authentication in the supplied cell lines, has been performed in the Andalusian Health System Biobank. Specifically, precise results were obtained using real time PCR detection for mycoplasma and 10 STRs identification by capillary electrophoresis for cell line authentication. Advantages and disadvantages of these protocols are discussed.
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47 CFR 1.929 - Classification of filings as major or minor.
Code of Federal Regulations, 2011 CFR
2011-10-01
... 47 Telecommunication 1 2011-10-01 2011-10-01 false Classification of filings as major or minor. 1.929 Section 1.929 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL PRACTICE AND PROCEDURE... Classification of filings as major or minor. Applications and amendments to applications for stations in the...
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20 CFR 404.929 - Hearing before an administrative law judge-general.
Code of Federal Regulations, 2010 CFR
2010-04-01
... evidence used in making the determination or decision under review, and present and question witnesses. The... 20 Employees' Benefits 2 2010-04-01 2010-04-01 false Hearing before an administrative law judge-general. 404.929 Section 404.929 Employees' Benefits SOCIAL SECURITY ADMINISTRATION FEDERAL OLD-AGE...
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Jia, J G; Zhang, L G; Guo, C X; Wang, Y G; Chen, B L; Wang, Y M; Qian, J
2016-03-01
To evaluate the effects of artemisinin on proliferation, cell cycle and apoptosis of gallbladder cancer cells. Gallbladder carcinoma cell lines(GBC-SD and NOZ)were cultured in vitro. The effects of artemisinin in different concentration on proliferation of the two cell lines in vitro were examined using MTT assay. The cell cycle distribution of GBC-SD and NOZ cells 24 h after treatments with artemisinin(20 μmol/L) were examined using flow cytometry. The apoptosis of GBC-SD and NOZ cells 24 h after treatments with artemisinin (20 μmol/L) were examined using Annexin V/PI staining.The expressions of p-ERK1/2, CDK4, cyclin D1, p16, cytochrome C and caspase-3 were examined by Western blot assay. t-test and one way ANOVA were used to evaluate the differences between two groups and more than two groups, respectively. The cell proliferation was significantly inhibited by artemisinin, the IC50 of artemisinin against GBC-SD and NOZ cells were 14.05 μmol/L and 12.42 μmol/L, respectively.Artemisinin induced cycle arrest, and G1 population of GBC-SD and NOZ cells increased to 74.60% and 78.86%. Cell apoptosis and apoptotic population of GBC-SD and NOZ cells were increased to 15.67% and 16.51% after dealt with artemisinin, respectively. In addition, expression of p16 was increased, and expressions of p-ERK1/2, CDK4 and cyclin D1 were down-regulated by artemisinin(all P<0.05). Cytochrome C was released from mitochondria to cytoplasm leading to the activation of caspase-3 and PARP after dealt with artemisinin(P<0.05). The inhibition effect of artemisinin on the proliferation gallbladder cancer cells is accompanied by down-regulation of ERK1/2 signaling pathway, G1 phase arrest and triggering caspase-3-mediate apoptosis.
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Pitz, Heloisa da S; Pereira, Aline; Blasius, Mayara B; Voytena, Ana Paula L; Affonso, Regina C L; Fanan, Simone; Trevisan, Adriana C D; Ribeiro-do-Valle, Rosa M; Maraschin, Marcelo
2016-01-01
Jaboticaba is a fruit from a native tree to Brazil, Plinia peruviana. Jaboticaba peels are an important source of antioxidant molecules such as phenolic compounds. This study aimed to evaluate in vitro the activity of a hydroalcoholic extract of jaboticaba fruit peels (HEJFP) in wound healing processes and antioxidant activity in murine fibroblasts (L929 cell line). HEJFP concentrations (0.5, 1, 5, 10, 25, 50, 100, and 200 µg/mL) were tested in MTT assay and cell proliferation was verified at 100 µg/mL after 24 h and at 25, 50, and 100 µg/mL after 48 h of extract exposure. Evaluation of antioxidant activity was performed at 0.5, 5, 25, 50, and 100 µg/mL HEJFP concentrations. Cell treatment with HEJFP at 25, 50, and 100 µg/mL for 24 h followed by H2O2 exposure for 3 h showed a strong cytoprotective effect. In vitro scratch wound healing assay indicated that none of tested HEJFP concentrations (0.5, 5, 25, 50, and 100 µg/mL) were capable of increasing migration rate after 12 h of incubation. These results demonstrate a positive effect of HEJFP on the wound healing process on L929 fibroblasts cell line, probably due to the antioxidant activity exhibited by phytochemicals in the extract.
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7 CFR 929.152 - Delinquent assessments.
Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.152 Delinquent assessments. There... Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing...
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7 CFR 929.103 - Inspection procedure.
Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.103 Inspection procedure. (a... Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing...
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7 CFR 929.101 - Minimum exemption.
Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.101 Minimum exemption. The... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements...
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7 CFR 929.59 - Excess cranberries.
Code of Federal Regulations, 2010 CFR
2010-01-01
... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Regulations § 929.59 Excess cranberries...
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Kubásek, J; Vojtěch, D; Jablonská, E; Pospíšilová, I; Lipov, J; Ruml, T
2016-01-01
Zn-(0-1.6)Mg (in wt.%) alloys were prepared by hot extrusion at 300 °C. The structure, mechanical properties and in vitro biocompatibility of the alloys were investigated. The hot-extruded magnesium-based WE43 alloy was used as a control. Mechanical properties were evaluated by hardness, compressive and tensile testing. The cytotoxicity, genotoxicity (comet assay) and mutagenicity (Ames test) of the alloy extracts and ZnCl2 solutions were evaluated with the use of murine fibroblasts L929 and human osteosarcoma cell line U-2 OS. The microstructure of the Zn alloys consisted of recrystallized Zn grains of 12 μm in size and fine Mg2Zn11 particles arranged parallel to the hot extrusion direction. Mechanical tests revealed that the hardness and strength increased with increasing Mg concentration. The Zn-0.8 Mg alloys showed the best combination of tensile mechanical properties (tensile yield strength of 203 MPa, ultimate tensile strength of 301 MPa and elongation of 15%). At higher Mg concentrations the plasticity of Zn-Mg alloys was deteriorated. Cytotoxicity tests with alloy extracts and ZnCl2 solutions proved the maximum safe Zn(2+) concentrations of 120 μM and 80 μM for the U-2 OS and L929 cell lines, respectively. Ames test with extracts of alloys indicated that the extracts were not mutagenic. The comet assay demonstrated that 1-day extracts of alloys were not genotoxic for U-2 OS and L929 cell lines after 1-day incubation. Copyright © 2015 Elsevier B.V. All rights reserved.
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Ma, Li; Han, Xiao-hong; Wang, Shuai; Wang, Jian-fei; Shi, Yuan-kai
2012-09-25
To explore the effects of icotinib on the proliferation and apoptosis of various lung cancer cell lines. Human lung cancer cell lines HCC827, H1650, H1975, A549 and human epidermal cancer cell line A431 were treated in vitro with icotinib or gefitinib at a concentration gradient of 0 - 40 µmol/L. Their proliferation effects were analyzed by the thiazolyl blue (MTT) assay and the apoptotic effects detected by flow cytometer. The downstream signaling proteins were detected by Western blot. The median inhibitory concentrations (IC(50)) of icotinib for A431 and HCC827 cell lines were (0.04 ± 0.02) and (0.15 ± 0.06) µmol/L respectively. No significant differences existed between the inhibitions of gefitinib and icotinib on A431, HCC827, H1650, H1975 and A549 cell lines (all P > 0.05). Compared with H1650, H1975 and A549 cell lines, icotinib significantly inhibited A431 (P = 0.009, 0.005 and 0.000) and HCC827 (P = 0.001, 0.001 and 0.000) cell lines. And it lowered the expressions of p-AKT, p-ERK and survivin protein expression through the inhibited activity of p-EGFR protein. Icotinib can arrest the proliferation of lung adenocarcinoma cells with EGFR mutation or over-expression by inhibiting the signal pathways of AKT-ERK and survivin.
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7 CFR 929.106 - Fiscal period.
Code of Federal Regulations, 2010 CFR
2010-01-01
... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Rules and Regulations § 929.106 Fiscal period. The fiscal period... Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements...
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Xu, Jianlin; Yang, Haitang; Jin, Bo; Lou, Yuqing; Zhang, Yanwei; Zhang, Xueyan; Zhong, Hua; Wang, Huiming; Wu, Dan; Han, Baohui
2016-11-04
The efficacy of EGFR tyrosine kinase inhibitors (TKIs) varies among different EGFR mutations. Here, we directly compared the efficacy of first-line TKIs to chemotherapy for non-small cell lung cancer (NSCLC) patients with the L858R mutation. The progression-free survival (PFS) for patients receiving TKIs as first-line therapy was longer than those who received chemotherapy (hazard ratio [HR]: 0.44, P < 0.001). Subgroup analyses showed that first-line TKI therapy resulted in longer PFS among non-smokers (HR: 0.41, P < 0.001), male (HR: 0.49, P = 0.002), female (HR: 0.39, P < 0.001), and patients with adenocarcinoma histology (HR: 0.41, P < 0.001). However, among patients with non-adenocarcinoma histology (HR: 1.11, P = 0.824) and those who used to smoke (HR: 0.55, P = 0.093), first-line TKI therapy failed to demonstrate statistically longer PFS compared to chemotherapy. Our results demonstrated that for patients with L858R mutation, first-line TKI therapy provided better survival benefits. However, among non-adenocarcinoma patients and those who used to smoke, the PFS in cohorts receiving first-line chemotherapy or TKI were not significantly different. The results of the current study will be helpful for decision-making in the treatment of patients with L858R mutation.
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Molluscan cells in culture: primary cell cultures and cell lines
Yoshino, T. P.; Bickham, U.; Bayne, C. J.
2013-01-01
In vitro cell culture systems from molluscs have significantly contributed to our basic understanding of complex physiological processes occurring within or between tissue-specific cells, yielding information unattainable using intact animal models. In vitro cultures of neuronal cells from gastropods show how simplified cell models can inform our understanding of complex networks in intact organisms. Primary cell cultures from marine and freshwater bivalve and gastropod species are used as biomonitors for environmental contaminants, as models for gene transfer technologies, and for studies of innate immunity and neoplastic disease. Despite efforts to isolate proliferative cell lines from molluscs, the snail Biomphalaria glabrata Say, 1818 embryonic (Bge) cell line is the only existing cell line originating from any molluscan species. Taking an organ systems approach, this review summarizes efforts to establish molluscan cell cultures and describes the varied applications of primary cell cultures in research. Because of the unique status of the Bge cell line, an account is presented of the establishment of this cell line, and of how these cells have contributed to our understanding of snail host-parasite interactions. Finally, we detail the difficulties commonly encountered in efforts to establish cell lines from molluscs and discuss how these difficulties might be overcome. PMID:24198436
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Production of coagulation factor VII in human cell lines Sk-Hep-1 and HKB-11.
Corrêa de Freitas, Marcela Cristina; Bomfim, Aline de Sousa; Mizukami, Amanda; Picanço-Castro, Virgínia; Swiech, Kamilla; Covas, Dimas Tadeu
2017-09-01
Recombinant factor VII (rFVII) is the main therapeutic choice for hemophilia patients who have developed inhibitory antibodies against conventional treatments (FVIII and FIX). Because of the post-translational modifications, rFVII needs to be produced in mammalian cell lines. In this study, for the first time, we have shown efficient rFVII production in HepG2, Sk-Hep-1, and HKB-11 cell lines. Experiments in static conditions for a period of 96 h showed that HepG2-FVII produced the highest amounts of rhFVII, with an average of 1843 ng/mL. Sk-hep-1-FVII cells reached a maximum protein production of 1432 ng/mL and HKB-11-FVII cells reached 1468 ng/mL. Sk-Hep-1-rFVII and HKB-11-rFVII were selected for the first step of scale-up. Over 10 days of spinner flask culture, HKB-11 and SK-Hep-1 cells showed a cumulative production of rFVII of 152 μg and 202.6 μg in 50 mL, respectively. Thus, these human cell lines can be used for an efficient production of recombinant FVII. With more investment in basic research, human cell lines can be optimized for the commercial production of different bio therapeutic proteins. Copyright © 2017 Elsevier Inc. All rights reserved.
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Romano, Paolo; Manniello, Assunta; Aresu, Ottavia; Armento, Massimiliano; Cesaro, Michela; Parodi, Barbara
2009-01-01
The Cell Line Data Base (CLDB) is a well-known reference information source on human and animal cell lines including information on more than 6000 cell lines. Main biological features are coded according to controlled vocabularies derived from international lists and taxonomies. HyperCLDB (http://bioinformatics.istge.it/hypercldb/) is a hypertext version of CLDB that improves data accessibility by also allowing information retrieval through web spiders. Access to HyperCLDB is provided through indexes of biological characteristics and navigation in the hypertext is granted by many internal links. HyperCLDB also includes links to external resources. Recently, an interest was raised for a reference nomenclature for cell lines and CLDB was seen as an authoritative system. Furthermore, to overcome the cell line misidentification problem, molecular authentication methods, such as fingerprinting, single-locus short tandem repeat (STR) profile and single nucleotide polymorphisms validation, were proposed. Since this data is distributed, a reference portal on authentication of human cell lines is needed. We present here the architecture and contents of CLDB, its recent enhancements and perspectives. We also present a new related database, the Cell Line Integrated Molecular Authentication (CLIMA) database (http://bioinformatics.istge.it/clima/), that allows to link authentication data to actual cell lines. PMID:18927105
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Romano, Paolo; Manniello, Assunta; Aresu, Ottavia; Armento, Massimiliano; Cesaro, Michela; Parodi, Barbara
2009-01-01
The Cell Line Data Base (CLDB) is a well-known reference information source on human and animal cell lines including information on more than 6000 cell lines. Main biological features are coded according to controlled vocabularies derived from international lists and taxonomies. HyperCLDB (http://bioinformatics.istge.it/hypercldb/) is a hypertext version of CLDB that improves data accessibility by also allowing information retrieval through web spiders. Access to HyperCLDB is provided through indexes of biological characteristics and navigation in the hypertext is granted by many internal links. HyperCLDB also includes links to external resources. Recently, an interest was raised for a reference nomenclature for cell lines and CLDB was seen as an authoritative system. Furthermore, to overcome the cell line misidentification problem, molecular authentication methods, such as fingerprinting, single-locus short tandem repeat (STR) profile and single nucleotide polymorphisms validation, were proposed. Since this data is distributed, a reference portal on authentication of human cell lines is needed. We present here the architecture and contents of CLDB, its recent enhancements and perspectives. We also present a new related database, the Cell Line Integrated Molecular Authentication (CLIMA) database (http://bioinformatics.istge.it/clima/), that allows to link authentication data to actual cell lines.
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Akasov, Roman; Gileva, Anastasia; Zaytseva-Zotova, Daria; Burov, Sergey; Chevalot, Isabelle; Guedon, Emmanuel; Markvicheva, Elena
2017-01-01
To design novel 3D in vitro co-culture models based on the RGD-peptide-induced cell self-assembly technique. Multicellular spheroids from M-3 murine melanoma cells and L-929 murine fibroblasts were obtained directly from monolayer culture by addition of culture medium containing cyclic RGD-peptide. To reach reproducible architecture of co-culture spheroids, two novel 3D in vitro models with well pronounced core-shell structure from tumor spheroids and single mouse fibroblasts were developed based on this approach. The first was a combination of a RGD-peptide platform with the liquid overlay technique with further co-cultivation for 1-2 days. The second allowed co-culture spheroids to generate within polyelectrolyte microcapsules by cultivation for 2 weeks. M-3 cells (a core) and L-929 fibroblasts (a shell) were easily distinguished by confocal microscopy due to cell staining with DiO and DiI dyes, respectively. The 3D co-culture spheroids are proposed as a tool in tumor biology to study cell-cell interactions as well as for testing novel anticancer drugs and drug delivery vehicles.
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Goto, Hiroaki; Naruto, Takuya; Tanoshima, Reo; Kato, Hiromi; Yokosuka, Tomoko; Yanagimachi, Masakatsu; Fujii, Hisaki; Yokota, Shumpei; Komine, Hiromi
2009-10-01
Sensitivity to 10 anticancer drugs was evaluated in 6 childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cell lines. Authenticity of newly established cell lines was confirmed by genomic fingerprinting. The line YCUB-5R established at relapse was more resistant to 4-hydroperoxy-cyclophosphamide, cytarabine, L-asparaginase, topotecan, fludarabine, and etoposide than YCUB-5 from the same patient at diagnosis. Of the drugs tested, etoposide and SN-38 (irinotecan) showed highest efficacy in the panel, with 50% growth inhibition at 0.22-1.8 microg/ml and 0.57-3.6 ng/ml, respectively. This cell line panel offers an in vitro model for the development of new therapies for childhood BCP-ALL.
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7 CFR 929.45 - Research and development.
Code of Federal Regulations, 2012 CFR
2012-01-01
... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... production of cranberries. The expense of such projects shall be paid from funds collected pursuant to § 929...
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7 CFR 929.45 - Research and development.
Code of Federal Regulations, 2014 CFR
2014-01-01
... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... production of cranberries. The expense of such projects shall be paid from funds collected pursuant to § 929...
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Radiation sensitivity of Merkel cell carcinoma cell lines
DOE Office of Scientific and Technical Information (OSTI.GOV)
Leonard, J.H.; Ramsay, J.R.; Birrell, G.W.
1995-07-30
Merkel cell carcinoma (MCC), being a small cell carcinoma, would be expected to be sensitive to radiation. Clinical analysis of patients at our center, especially those with macroscopic disease, would suggest the response is quite variable. We have recently established a number of MCC cell lines from patients prior to radiotherapy, and for the first time are in a position to determine their sensitivity under controlled conditions. Some of the MCC lines grew as suspension cultures and could not be single cell cloned; therefore, it was not possible to use clonogenic survival for all cell lines. A tetrazolium based (MTT)more » assay was used for these lines, to estimate cell growth after {gamma} irradiation. Control experiments were conducted on lymphoblastoid cell lines (LCL) and the adherent MCC line, MCC13, to demonstrate that the two assays were comparable under the conditions used. We have examined cell lines from MCC, small cell lung cancer (SCLC), malignant melanomas, Epstein Barr virus (EBV) transformed lymphocytes (LCL), and skin fibroblasts for their sensitivity to {gamma} irradiation using both clonogenic cell survival and MTT assays. The results show that the tumor cell lines have a range of sensitivities, with melanoma being more resistant (surviving fraction at 2 Gy (SF2) 0.57 and 0.56) than the small cell carcinoma lines, MCC (SF2 range 0.21-0.45, mean SF2 0.30, n = 8) and SCLC (SF2 0.31). Fibroblasts were the most sensitive (SF2 0.13-0.20, mean 0.16, n = 5). The MTT assay, when compared to clonogenic assay for the MCC13 adherent line and the LCL, gave comparable results under the conditions used. Both assays gave a range of SF2 values for the MCC cell lines, suggesting that these cancers would give a heterogeneous response in vivo. The results with the two derivative clones of MCC14 (SF2 for MCC14/1 0.38, MCC14/2 0.45) would further suggest that some of them may develop resistance during clonogenic evolution. 25 refs., 3 figs., 1 tab.« less
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Inhibitory effect of emodin on human hepatoma cell line SMMC-7721 and its mechanism.
Zhang, Xia; Chen, Yingping; Zhang, Ting; Zhang, Yaming
2015-03-01
Da Huang (Radix et Rhizoma Rhei) is the dried root or rhizome of Rheum palmatum L., Rheum tanguticum Maxim ex Balf. or Rheum officinale Braill of family Polygonaceae. It has heat clearing, damp drying, fire purging and toxin removing effects. Because of its definite curative efficacy, it has been widely applied in clinical settings. To study the inhibitory effect of emodin on human hepatoma cell line SMMC-7721 and its mechanism. MTT assay, flow cytometry and electron microscopy were used to investigate the inhibitory effect of different concentrations of emodin on human hepatoma cell line SMMC-7721. 12 h, 24 h and 48 h after the action of 20, 40 and 80 umol/L emodin on SMMC-7721 cells, the proliferation of human hepatoma SMMC-7721 cells was inhibited; the inhibitory effects showed time-and concentration-dependence. 48 h after the action of different concentrations of emodin on SMMC-7721 cells, cells in G2/M phase increased significantly, while the proportion of S phase cells gradually declined. Emodin can inhibit human hepatoma cell line SMMC-7721.
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BHD Tumor Cell Line and Renal Cell Carcinoma Line | NCI Technology Transfer Center | TTC
Scientists at the National Cancer Institute have developed a novel renal cell carcinoma (RCC) cell line designated UOK257, which was derived from the surgical kidney tissue of a patient with hereditary Birt-Hogg-Dube''''(BHD) syndrome and companion cell line UOK257-2 in which FLCN expression has been restored by lentivirus infection. The NCI Urologic Oncology Branch seeks parties interested in licensing or collaborative research to co-develop, evaluate, or commercialize kidney cancer tumor cell lines.
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L1 stimulation of human glioma cell motility correlates with FAK activation
Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A.; Boulos, Magdy I.; Kappes, John C.
2011-01-01
The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass
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L1 stimulation of human glioma cell motility correlates with FAK activation.
Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A; Boulos, Magdy I; Kappes, John C; Galileo, Deni S
2011-10-01
The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass
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Korch, Christopher; Hall, Erin M.; Dirks, Wilhelm G.; Ewing, Margaret; Faries, Mark; Varella‐Garcia, Marileila; Robinson, Steven; Storts, Douglas; Turner, Jacqueline A.; Wang, Ying; Burnett, Edward C.; Healy, Lyn; Kniss, Douglas; Neve, Richard M.; Nims, Raymond W.; Reid, Yvonne A.; Robinson, William A.
2017-01-01
A variety of analytical approaches have indicated that melanoma cell line UCLA‐SO‐M14 (M14) and breast carcinoma cell line MDA‐MB‐435 originate from a common donor. This indicates that at some point in the past, one of these cell lines became misidentified, meaning that it ceased to correspond to the reported donor and instead became falsely identified (through cross‐contamination or other means) as a cell line from a different donor. Initial studies concluded that MDA‐MB‐435 was the misidentified cell line and M14 was the authentic cell line, although contradictory evidence has been published, resulting in further confusion. To address this question, we obtained early samples of the melanoma cell line (M14), a lymphoblastoid cell line from the same donor (ML14), and donor serum preserved at the originator's institution. M14 samples were cryopreserved in December 1975, before MDA‐MB‐435 cells were established in culture. Through a series of molecular characterizations, including short tandem repeat (STR) profiling and cytogenetic analysis, we demonstrated that later samples of M14 and MDA‐MB‐435 correspond to samples of M14 frozen in 1975, to the lymphoblastoid cell line ML14, and to the melanoma donor's STR profile, sex and blood type. This work demonstrates conclusively that M14 is the authentic cell line and MDA‐MB‐435 is misidentified. With clear provenance information and authentication testing of early samples, it is possible to resolve debates regarding the origins of problematic cell lines that are widely used in cancer research. PMID:28940260
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40 CFR 63.929 - Implementation and enforcement.
Code of Federal Regulations, 2014 CFR
2014-07-01
...) National Emission Standards for Containers § 63.929 Implementation and enforcement. (a) This subpart can be... § 63.90, and as required in this subpart. (3) Approval of major alternatives to monitoring under § 63.8...
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40 CFR 63.929 - Implementation and enforcement.
Code of Federal Regulations, 2011 CFR
2011-07-01
...) National Emission Standards for Containers § 63.929 Implementation and enforcement. (a) This subpart can be... § 63.90, and as required in this subpart. (3) Approval of major alternatives to monitoring under § 63.8...
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40 CFR 63.929 - Implementation and enforcement.
Code of Federal Regulations, 2010 CFR
2010-07-01
...) National Emission Standards for Containers § 63.929 Implementation and enforcement. (a) This subpart can be... § 63.90, and as required in this subpart. (3) Approval of major alternatives to monitoring under § 63.8...
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40 CFR 63.929 - Implementation and enforcement.
Code of Federal Regulations, 2012 CFR
2012-07-01
...) National Emission Standards for Containers § 63.929 Implementation and enforcement. (a) This subpart can be... § 63.90, and as required in this subpart. (3) Approval of major alternatives to monitoring under § 63.8...
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40 CFR 63.929 - Implementation and enforcement.
Code of Federal Regulations, 2013 CFR
2013-07-01
...) National Emission Standards for Containers § 63.929 Implementation and enforcement. (a) This subpart can be... § 63.90, and as required in this subpart. (3) Approval of major alternatives to monitoring under § 63.8...
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Al-Rabia, Mohammed W; Blaylock, Morgan G; Sexton, Darren W; Walsh, Garry M
2004-06-01
Caspases are key molecules in the control of apoptosis, but relatively little is known about their contribution to eosinophil apoptosis. We examined caspase-3, -8, and -9 activities in receptor ligation-dependent apoptosis induction in the differentiated human eosinophilic cell line EoL-1. Differentiated EoL-1 exhibited bi-lobed nuclei, eosinophil-associated membrane receptors, and basic granule proteins. Annexin-V fluorescein isothiocyanate binding to EoL-1 revealed significant (P<0.01) apoptosis induction in cells cultured for 20 h with monoclonal antibodies (mAb) specific for CD45 (71%+/-4.3), CD45RA (58%+/-2.3), CD45RB (68%+/-2.4), CD95 (47%+/-2.6), and CD69 (52%+/-2.1) compared with control (23%+/-1.6) or CD45RO mAb (27%+/-3.9). The pan-caspase inhibitor Z-Val-Ala-Asp-fluoromethylketone (fmk) and inhibitors of caspase-8 (Z-Ile-Glu-Thr-Asp-fmk) and caspase-9 (Z-Leu-Glu-His-Asp-fmk) significantly inhibited mAb-induced apoptosis of EoL-1 but had no effect on constitutive (baseline) apoptosis at 16 and 20 h. Caspase activity was analyzed using the novel CaspaTag trade mark technique and flow cytometry. EoL-1 treated with pan-CD45, CD45RA, CD45RB, and CD95 mAb exhibited caspase-3 and -9 activation at 12 h post-treatment, which increased at 16 and 20 h. Activated caspase-8 was detected 12 and 16 h after ligation with CD45, CD45RA, CD45RB, and CD95 mAb followed by a trend toward basal levels at 20 h. CD69 ligation resulted in caspase-3 activation, a modest but significant activation of caspase-8, and a loss in mitochondrial transmembrane potential but had no significant effect on activation of caspase-9. Thus, the intrinsic and extrinsic caspase pathways are involved in controlling receptor ligation-mediated apoptosis induction in human eosinophils, findings that may aid the development of a more targeted, anti-inflammatory therapy for asthma.
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Antigen-specific T-cell lines transfer protective immunity against Trichinella spiralis in vivo.
Riedlinger, J; Grencis, R K; Wakelin, D
1986-01-01
T-cell lines specific for infective muscle larvae antigens of the intestinal nematode Trichinella spiralis have been generated in vitro. These antigen-specific T-cell lines express the L3T4+ Ly2- phenotype and secrete the lymphokines IL-2, IL-3 and gamma-IFN. They are stable in culture for up to 15 weeks and are protective when adoptively transferred into naive recipients. As few as 2 x 10(5) T. spiralis-specific tract. In addition, intestinal mastocytosis and peripheral blood eosinophilia were accelerated after adoptive transfer of T. spiralis-specific T-cell lines. PMID:2423438
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Thyroid cell lines in research on goitrogenesis.
Gerber, H; Peter, H J; Asmis, L; Studer, H
1991-12-01
Thyroid cell lines have contributed a lot to the understanding of goitrogenesis. The cell lines mostly used in thyroid research are briefly discussed, namely the rat thyroid cell lines FRTL and FRTL-5, the porcine thyroid cell lines PORTHOS and ARTHOS, The sheep thyroid cell lines OVNIS 5H and 6H, the cat thyroid cell lines PETCAT 1 to 4 and ROMCAT, and the human thyroid cell lines FTC-133 and HTh 74. Chinese hamster ovary (CHO) cells and COS-7 cells, stably transfected with TSH receptor cDNA and expressing a functional TSH receptor, are discussed as examples for non-thyroidal cells, transfected with thyroid genes.
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Shin, Ji-Ae; Ryu, Mi Heon; Kwon, Ki-Han; Choi, BuYoung; Cho, Sung-Dae
2015-07-01
The apoptotic activity of methanol extracts of Impatiens balsamina L. (MEIB) and related mechanisms in human oral squamous cell carcinoma (OSCC) cells have been systematically investigated. The effects of MEIB on human OSCC cell lines were investigated using trypan blue exclusion assay, MTS assay, Western blot, 4'-6-diamidino-2-phenylindole (DAPI) staining, Live/Dead assay, Immunohistochemistry, reverse transcription-polymerase chain reaction, and promoter assay. MEIB decreased cell viability and induced apoptosis in HSC-4 cells. Higher levels of p-Akt expression were observed in OSCC than in normal oral mucosa (NOM), and it correlated with poor survival of the patients. MEIB dephosphorylated p-Akt and decreased Akt expression through proteasome-dependent degradation. LY294002 (PI3K inhibitor) decreased p-Akt and Akt, resulting in enhancing MEIB-induced apoptosis. MEIB down-regulated the expression level of survivin protein at the transcriptional level and YM155 (survivin inhibitor) decreased survivin, which facilitated MEIB-induced apoptosis. MEIB and LY294002 significantly increased Bax, thereby inducing the conformational change, mitochondrial translocation, and oligomerization. In addition, MEIB-induced growth inhibition and apoptosis in OSC-20, another human OSCC cells were mediated by regulating Akt and it downstream targets, survivin and Bax. These results suggest that MEIB may serve as a potential drug candidate for the treatment of human OSCC. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
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Philippe, Claude; Vargas-Landin, Dulce B; Doucet, Aurélien J; van Essen, Dominic; Vera-Otarola, Jorge; Kuciak, Monika; Corbin, Antoine; Nigumann, Pilvi; Cristofari, Gaël
2016-01-01
LINE-1 (L1) retrotransposons represent approximately one sixth of the human genome, but only the human-specific L1HS-Ta subfamily acts as an endogenous mutagen in modern humans, reshaping both somatic and germline genomes. Due to their high levels of sequence identity and the existence of many polymorphic insertions absent from the reference genome, the transcriptional activation of individual genomic L1HS-Ta copies remains poorly understood. Here we comprehensively mapped fixed and polymorphic L1HS-Ta copies in 12 commonly-used somatic cell lines, and identified transcriptional and epigenetic signatures allowing the unambiguous identification of active L1HS-Ta copies in their genomic context. Strikingly, only a very restricted subset of L1HS-Ta loci - some being polymorphic among individuals - significantly contributes to the bulk of L1 expression, and these loci are differentially regulated among distinct cell lines. Thus, our data support a local model of L1 transcriptional activation in somatic cells, governed by individual-, locus-, and cell-type-specific determinants. DOI: http://dx.doi.org/10.7554/eLife.13926.001 PMID:27016617
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Kuwae, Shinobu; Miyakawa, Ichiko; Doi, Tomohiro
2018-01-11
A chemically defined platform basal medium and feed media were developed using a single Chinese hamster ovary (CHO) cell line that produces a monoclonal antibody (mAb). Cell line A, which showed a peak viable cell density of 5.9 × 10 6 cells/mL and a final mAb titer of 0.5 g/L in batch culture, was selected for the platform media development. Stoichiometrically balanced feed media were developed using glucose as an indicator of cell metabolism to determine the feed rates of all other nutrients. A fed-batch culture of cell line A using the platform fed-batch medium yielded a 6.4 g/L mAb titer, which was 12-fold higher than that of the batch culture. To examine the applicability of the platform basal medium and feed media, three other cell lines (A16, B, and C) that produce mAbs were cultured using the platform fed-batch medium, and they yielded mAb titers of 8.4, 3.3, and 6.2 g/L, respectively. The peak viable cell densities of the three cell lines ranged from 1.3 × 10 7 to 1.8 × 10 7 cells/mL. These results show that the nutritionally balanced fed-batch medium and feeds worked well for other cell lines. During the medium development, we found that choline limitation caused a lower cell viability, a lower mAb titer, a higher mAb aggregate content, and a higher mannose-5 content. The optimal choline chloride to glucose ratio for the CHO cell fed-batch culture was determined. Our platform basal medium and feed media will shorten the medium-development time for mAb-producing cell lines.
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Fucoidan Does Not Exert Anti-Tumorigenic Effects on Uveal Melanoma Cell Lines
Dithmer, Michaela; Kirsch, Anna-Maria; Richert, Elisabeth; Fuchs, Sabine; Wang, Fanlu; Schmidt, Harald; Coupland, Sarah E.; Roider, Johann; Klettner, Alexa
2017-01-01
Background. The polysaccharide fucoidan is widely investigated as an anti-cancer agent. Here, we tested the effect of fucoidan on uveal melanoma cell lines. Methods. The effect of 100 µM fucoidan was investigated on five cell lines (92.1, Mel270 OMM1, OMM2.3, OMM2.5) and of 1 µg/mL–1 mg/mL fucoidan in two cell lines (OMM1, OMM2.3). Cell proliferation and viability were investigated with a WST-1 assay, migration in a wound healing (scratch) assay. Vascular Endothelial Growth Factor (VEGF) was measured in ELISA. Angiogenesis was evaluated in co-cultures with endothelial cells. Cell toxicity was induced by hydrogen-peroxide. Protein expression (Akt, ERK1/2, Bcl-2, Bax) was investigated in Western blot. Results. Fucoidan increased proliferation in two and reduced it in one cell line. Migration was reduced in three cell lines. The effect of fucoidan on VEGF was cell type and concentration dependent. In endothelial co-culture with 92.1, fucoidan significantly increased tubular structures. Moreover, fucoidan significantly protected all tested uveal melanoma cell lines from hydrogen-peroxide induced cell death. Under oxidative stress, fucoidan did not alter the expression of Bcl-2, Bax or ERK1/2, while inducing Akt expression in 92.1 cells but not in any other cell line. Conclusion. Fucoidan did not show anti-tumorigenic effects but displayed protective and pro-angiogenic properties, rendering fucoidan unsuitable as a potential new drug for the treatment of uveal melanoma. PMID:28640204
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Natural zwitterionic l-Carnitine as efficient cryoprotectant for solvent-free cell cryopreservation.
Zhai, Hongwen; Yang, Jing; Zhang, Jiamin; Pan, Chao; Cai, Nana; Zhu, Yingnan; Zhang, Lei
2017-07-15
Organic solvents, such as dimethyl sulfoxide (DMSO) and glycerol, have been commonly used as cryoprotectants (CPAs) in cell cryopreservation. However, their cytotoxicity and need of complex freezing protocols have impeded their applications especially in clinical cell therapy and regenerative medicine. Trehalose has been explored as a natural CPA to cryopreserve cells, but its poor cell permeability frequently results in low cryopreservation efficacy. In this work, we presented that a natural zwitterionic molecule-l-carnitine-could serve as a promising CPA for solvent-free cryopreservation. We demonstrated that l-carnitine possessed strong ability to depress water freezing point, and with ultrarapid freezing protocol, we studied the post-thaw survival efficiency of four cell lines (GLC-82 cells, MCF-7 cells, NIH-3T3 cells and Sheep Red Blood Cells) using l-carnitine without addition of any organic solvents. At the optimum l-carnitine concentration, all four cell lines could achieve above 80% survival efficiency, compared with the significantly lower efficiency using organic CPAs and trehalose. After cryopreservation, the recovered cell behaviors including cell attachment and proliferation were found to be similar to the normal cells, indicating that the cell functionalities were not affected. Moreover, l-carnitine showed no observable cytotoxicity, which was superior to the organic CPAs. This work offered an attractive alternative to traditional CPAs and held great promise to revolutionize current cryopreservation technologies, to benefit the patients in various cell-based clinical applications. Copyright © 2017 Elsevier Inc. All rights reserved.
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40 CFR 35.929-2 - General requirements for all user charge systems.
Code of Federal Regulations, 2014 CFR
2014-07-01
... 40 Protection of Environment 1 2014-07-01 2014-07-01 false General requirements for all user charge systems. 35.929-2 Section 35.929-2 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... than every 2 years the waste water contribution of users and user classes, the total costs of operation...
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40 CFR 35.929-2 - General requirements for all user charge systems.
Code of Federal Regulations, 2012 CFR
2012-07-01
... 40 Protection of Environment 1 2012-07-01 2012-07-01 false General requirements for all user charge systems. 35.929-2 Section 35.929-2 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... than every 2 years the waste water contribution of users and user classes, the total costs of operation...
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40 CFR 35.929-2 - General requirements for all user charge systems.
Code of Federal Regulations, 2013 CFR
2013-07-01
... 40 Protection of Environment 1 2013-07-01 2013-07-01 false General requirements for all user charge systems. 35.929-2 Section 35.929-2 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... than every 2 years the waste water contribution of users and user classes, the total costs of operation...
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Establishment and characterization of two cell lines from bluefin trevally Caranx melampygus.
Zhao, Zhengshan; Lu, Yuanan
2006-01-30
Bluefin trevally Caranx melampygus Cuvier is a popular game fish and highly valued sea food with potential importance for aquaculture. To help establish this marine animal as an important aquacultural species in Hawaii and the Pacific and develop in vitro cell culture systems for long-term management and control of potential viral diseases 2 cell lines were established from body muscle (bluefin trevally muscles, BTMS) and fins (bluefin trevally fins, BTF). Primary culture of these cells was conducted at 25 degrees C using L-15 medium supplemented with 20% fetal bovine serum and various antibiotics. These cells have been serially subcultured 37 to 41 times since their initiation in June 2002. Growth of the bluefin trevally cells was serum-dependent at the time of the experiments and their plating efficiencies ranged from 11 to 28.2%. Comparative analysis showed that these bluefin trevally cells grew equally well in the media L-15 (Leibovitz medium), RPMI 1640, M199 and MEM (minimum essential medium), which are commonly used for cell cultures derived from aquatic animals and mammalian species. Examination of the early passage cells stored at -196 degrees C revealed a large percent (nearly 98%) of cell viability following a 6 mo storage in liquid nitrogen. Karyotyping analysis indicated that these bluefin trevally derived cell lines remained diploid with a chromosome count of 48 at passage 7 and 12. These 2 cell lines shared a same pattern of viral susceptibility and they were sensitive to infectious hematopoietic necrosis virus (IHNV), infectious pancreatic necrosis (IPN), spring viremia carp virus (SVCV), viral hemorrhagic septicemia virus (VHSV), and snakehead rhabdovirus (SHRV) but refractory to channel catfish virus (CCV) infection. These newly established cell lines are currently being used to facilitate the diagnosis of viral disease affecting marine fish aquaculture in Hawaii, and will be available for future isolation and study of bluefin trevally fish
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Why are breast cancer stem cells resistant to radiation?
2013-03-01
of Human Hsp27 in Rodent Cells: Absence of Compensatory Regulation between Small Heat Shock Proteins. J. Therm. Biol., 21, 365-372, 1996. 69...Corry, P.M., and Lee, Y.J. Comparison of Tumor Growth between Hsp25 and Hsp27 Transfected Murine L929 Cells in Nude Mice. Int. J. Cancer, 72, 871
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7 CFR 929.20 - Establishment and membership.
Code of Federal Regulations, 2010 CFR
2010-01-01
... Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing... MASSACHUSETTS, RHODE ISLAND, CONNECTICUT, NEW JERSEY, WISCONSIN, MICHIGAN, MINNESOTA, OREGON, WASHINGTON, AND LONG ISLAND IN THE STATE OF NEW YORK Order Regulating Handling Administrative Body § 929.20...
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Khanna, Swati; Thomas, Anish; Abate-Daga, Daniel; Zhang, Jingli; Morrow, Betsy; Steinberg, Seth M.; Orlandi, Augusto; Ferroni, Patrizia; Schlom, Jeffrey; Guadagni, Fiorella; Hassan, Raffit
2016-01-01
INTRODUCTION The functional aspects of programmed death 1 (PD-1) and PD ligand 1 (PD-L1) immune checkpoints in malignant mesothelioma have not been studied. METHODS Tumor samples from 65 patients with mesothelioma were evaluated for PD-L1 expression by immunohistochemistry and its prognostic significance. Malignant effusions from patients with pleural and peritoneal mesothelioma were evaluated for PD-1+ and PD-L1+ infiltrating lymphocytes and their role in inducing tumor cell PD-L1 expression. Antibody dependent cellular cytotoxicity (ADCC) of avelumab, a fully humanized IgG1 anti PD-L1 antibody towards primary mesothelioma cell lines was evaluated in presence of autologous and allogeneic NK cells. RESULTS Of 65 pleural and peritoneal mesothelioma tumors examined, 41 (63%) were PD-L1 positive, which was associated with slightly inferior overall survival compared to patients with PD-L1 negative tumors (median 23.0 vs. 33.3 months; p=0.35). The frequency of PD-L1 expression was similar in pleural and peritoneal mesothelioma patients with 62% and 64% of samples positive, respectively. Of nine mesothelioma effusion samples evaluated, the fraction of cells expressing PD-L1 ranged from 12 to 83%. Of 7 patients with paired malignant effusion and peripheral blood mononuclear cells (PBMC) samples, PD-L1 expression was significantly higher on CD3+ T cells present in malignant effusions as compared with PBMC (p=0.016). In addition, CD14+PD-1+ cells were elevated in malignant effusions compared with PBMC (p=0.031). The lymphocytes present in malignant effusions recognized autologous tumor cells and induced IFN-γ-mediated PD-L1 expression on the tumor cell surface. Of the three primary mesothelioma cell lines tested, two were susceptible to avelumab mediated ADCC in presence of autologous NK cells. CONCLUSION The majority of pleural as well as peritoneal mesothelioma express PD-L1. Malignant effusions in this disease are characterized by presence of tumor cells and CD3+ T
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Cytotoxicity of titanium dioxide nanoparticles in mouse fibroblast cells.
Jin, Cheng-Yu; Zhu, Bang-Shang; Wang, Xue-Feng; Lu, Qing-Hua
2008-09-01
Nanotitanium dioxide (TiO2) is an important industrial material that is widely used as an additive in cosmetics, pharmaceuticals, and food colorants. Although the small size of the TiO2 nanoparticle is useful in various applications, the biosafety of this material needs to be evaluated. In this study, mouse fibroblast (L929) cells were used to evaluate the cytotoxicity of different concentrations (3-600 microg/mL) of homogeneous and weakly aggregated TiO2 nanoparticles in aqueous solution. The L929 cells became round and even shrank as the concentration of TiO2 nanoparticles increased. Moreover, TiO2 nanoparticle-treated cells had condensed fragmented chromatin or were directly necrosed, as observed by acridine orange (AO) staining. The transmission electron microscopy (TEM) analysis showed that in cells cultured in a medium containing 300 microg/mL TiO2, the number of lysosomes increased, and some cytoplasmic organelles were damaged. In addition, there was a significant increase in oxidative stress at higher TiO2 nanoparticle concentrations (>60 microg/mL). As the concentration of TiO2 nanoparticles increased in the culture medium, the levels of reactive oxygen species (ROS) and lactate dehydrogenase (LDH) increased, while those of methyl tetrazolium cytotoxicity (MTT), glutathione (GSH), and superoxide dismutase (SOD) decreased. A possible mechanism for the cytotoxicity of TiO2 nanoparticles is also discussed.
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Duan, Qiaonan; Wang, Zichen; Fernandez, Nicolas F; Rouillard, Andrew D; Tan, Christopher M; Benes, Cyril H; Ma'ayan, Avi
2014-11-15
Recently, several high profile studies collected cell viability data from panels of cancer cell lines treated with many drugs applied at different concentrations. Such drug sensitivity data for cancer cell lines provide suggestive treatments for different types and subtypes of cancer. Visualization of these datasets can reveal patterns that may not be obvious by examining the data without such efforts. Here we introduce Drug/Cell-line Browser (DCB), an online interactive HTML5 data visualization tool for interacting with three of the recently published datasets of cancer cell lines/drug-viability studies. DCB uses clustering and canvas visualization of the drugs and the cell lines, as well as a bar graph that summarizes drug effectiveness for the tissue of origin or the cancer subtypes for single or multiple drugs. DCB can help in understanding drug response patterns and prioritizing drug/cancer cell line interactions by tissue of origin or cancer subtype. DCB is an open source Web-based tool that is freely available at: http://www.maayanlab.net/LINCS/DCB CONTACT: avi.maayan@mssm.edu Supplementary data are available at Bioinformatics online. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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L-Dopa decarboxylase expression profile in human cancer cells.
Chalatsa, Ioanna; Nikolouzou, Eleftheria; Fragoulis, Emmanuel G; Vassilacopoulou, Dido
2011-02-01
L-Dopa decarboxylase (DDC) catalyses the decarboxylation of L-Dopa. It has been shown that the DDC gene undergoes alternative splicing within its 5'-untranslated region (UTR), in a tissue-specific manner, generating identical protein products. The employment of two alternative 5'UTRs is thought to be responsible for tissue-specific expression of the human DDC mRNA. In this study, we focused on the investigation of the nature of the mRNA expression in human cell lines of neural and non-neural origin. Our results show the expression of a neural-type DDC mRNA splice variant, lacking exon 3 in all cell lines studied. Co-expression of the full length non-neural DDC mRNA and the neural-type DDC splice variant lacking exon 3 was detected in all cell lines. The alternative DDC protein isoform, Alt-DDC, was detected in SH-SY5Y and HeLa cells. Our findings suggest that the human DDC gene undergoes complex processing, leading to the formation of multiple mRNA isoforms. The study of the significance of this phenomenon of multiple DDC mRNA isoforms could provide us with new information leading to the elucidation of the complex biological pathways that the human enzyme is involved in.
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Klawitter, Sabine; Fuchs, Nina V; Upton, Kyle R; Muñoz-Lopez, Martin; Shukla, Ruchi; Wang, Jichang; Garcia-Cañadas, Marta; Lopez-Ruiz, Cesar; Gerhardt, Daniel J; Sebe, Attila; Grabundzija, Ivana; Merkert, Sylvia; Gerdes, Patricia; Pulgarin, J Andres; Bock, Anja; Held, Ulrike; Witthuhn, Anett; Haase, Alexandra; Sarkadi, Balázs; Löwer, Johannes; Wolvetang, Ernst J; Martin, Ulrich; Ivics, Zoltán; Izsvák, Zsuzsanna; Garcia-Perez, Jose L; Faulkner, Geoffrey J; Schumann, Gerald G
2016-01-08
Human induced pluripotent stem cells (hiPSCs) are capable of unlimited proliferation and can differentiate in vitro to generate derivatives of the three primary germ layers. Genetic and epigenetic abnormalities have been reported by Wissing and colleagues to occur during hiPSC derivation, including mobilization of engineered LINE-1 (L1) retrotransposons. However, incidence and functional impact of endogenous retrotransposition in hiPSCs are yet to be established. Here we apply retrotransposon capture sequencing to eight hiPSC lines and three human embryonic stem cell (hESC) lines, revealing endogenous L1, Alu and SINE-VNTR-Alu (SVA) mobilization during reprogramming and pluripotent stem cell cultivation. Surprisingly, 4/7 de novo L1 insertions are full length and 6/11 retrotransposition events occurred in protein-coding genes expressed in pluripotent stem cells. We further demonstrate that an intronic L1 insertion in the CADPS2 gene is acquired during hiPSC cultivation and disrupts CADPS2 expression. These experiments elucidate endogenous retrotransposition, and its potential consequences, in hiPSCs and hESCs.
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Klawitter, Sabine; Fuchs, Nina V.; Upton, Kyle R.; Muñoz-Lopez, Martin; Shukla, Ruchi; Wang, Jichang; Garcia-Cañadas, Marta; Lopez-Ruiz, Cesar; Gerhardt, Daniel J.; Sebe, Attila; Grabundzija, Ivana; Merkert, Sylvia; Gerdes, Patricia; Pulgarin, J. Andres; Bock, Anja; Held, Ulrike; Witthuhn, Anett; Haase, Alexandra; Sarkadi, Balázs; Löwer, Johannes; Wolvetang, Ernst J.; Martin, Ulrich; Ivics, Zoltán; Izsvák, Zsuzsanna; Garcia-Perez, Jose L.; Faulkner, Geoffrey J.; Schumann, Gerald G.
2016-01-01
Human induced pluripotent stem cells (hiPSCs) are capable of unlimited proliferation and can differentiate in vitro to generate derivatives of the three primary germ layers. Genetic and epigenetic abnormalities have been reported by Wissing and colleagues to occur during hiPSC derivation, including mobilization of engineered LINE-1 (L1) retrotransposons. However, incidence and functional impact of endogenous retrotransposition in hiPSCs are yet to be established. Here we apply retrotransposon capture sequencing to eight hiPSC lines and three human embryonic stem cell (hESC) lines, revealing endogenous L1, Alu and SINE-VNTR-Alu (SVA) mobilization during reprogramming and pluripotent stem cell cultivation. Surprisingly, 4/7 de novo L1 insertions are full length and 6/11 retrotransposition events occurred in protein-coding genes expressed in pluripotent stem cells. We further demonstrate that an intronic L1 insertion in the CADPS2 gene is acquired during hiPSC cultivation and disrupts CADPS2 expression. These experiments elucidate endogenous retrotransposition, and its potential consequences, in hiPSCs and hESCs. PMID:26743714
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Hu, Zhilan; Guo, Donglin; Yip, Shirley S M; Zhan, Dejin; Misaghi, Shahram; Joly, John C; Snedecor, Bradley R; Shen, Amy Y
2013-01-01
Therapeutic monoclonal antibodies (mAb) are often produced in Chinese hamster ovary (CHO) cells. Three commonly used CHO host cells for generating stable cell lines to produce therapeutic proteins are dihydrofolate reductase (DHFR) positive CHOK1, DHFR-deficient DG44, and DUXB11-based DHFR deficient CHO. Current Genentech commercial full-length antibody products have all been produced in the DUXB11-derived DHFR-deficient CHO host. However, it has been challenging to develop stable cell lines producing an appreciable amount of antibody proteins in the DUXB11-derived DHFR-deficient CHO host for some antibody molecules and the CHOK1 host has been explored as an alternative approach. In this work, stable cell lines were developed for three antibody molecules in both DUXB11-based and CHOK1 hosts. Results have shown that the best CHOK1 clones produce about 1 g/l for an antibody mAb1 and about 4 g/l for an antibody mAb2 in 14-day fed batch cultures in shake flasks. In contrast, the DUXB11-based host produced ∼0.1 g/l for both antibodies in the same 14-day fed batch shake flask production experiments. For an antibody mAb3, both CHOK1 and DUXB11 host cells can generate stable cell lines with the best clone in each host producing ∼2.5 g/l. Additionally, studies have shown that the CHOK1 host cell has a larger endoplasmic reticulum and higher mitochondrial mass. © 2013 American Institute of Chemical Engineers.
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She, Zhen; Wang, Chunxia; Li, Jun; Sukhorukov, Gleb B; Antipina, Maria N
2012-07-09
Basic fibroblast growth factor (FGF2) is an important protein for cellular activity and highly vulnerable to environmental conditions. FGF2 protected by heparin and bovine serum albumin was loaded into the microcapsules by a coprecipitation-based layer-by-layer encapsulation method. Low cytotoxic and biodegradable polyelectrolytes dextran sulfate and poly-L-arginine were used for capsule shell assembly. The shell thickness-dependent encapsulation efficiency was measured by enzyme-linked immunosorbent assay. A maximum encapsulation efficiency of 42% could be achieved by microcapsules with a shell thickness of 14 layers. The effects of microcapsule concentration and shell thickness on cytotoxicity, FGF2 release kinetics, and L929 cell proliferation were evaluated in vitro. The advantage of using microcapsules as the carrier for FGF2 controlled release for enhancing L929 cell proliferation was analyzed.
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Zhou, Xu; Qin, Xianyan; Gong, Tao; Zhang, Zhi-Rong; Fu, Yao
2017-07-01
d-Fructose modified poly(ε-caprolactone)-polyethylene glycol (PCL-PEG-Fru) diblock amphiphile is synthesized via Cu(I)-catalyzed click chemistry, which self-assembles with D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) into PCL-PEG-Fru/TPGS mixed micelles (PPF MM). It has been proven that glucose transporter (GLUT)5 is overexpressed in MCF-7 cells other than L929 cells. In this study, PPF MM exhibit a significantly higher uptake efficiency than fructose-free PCL-PEG-N 3 /TPGS mixed micelles in both 2D MCF-7 cells and 3D tumor spheroids. Also, the presence of free d-fructose competitively inhibits the internalization of PPF MM in MCF-7 cells other than L929 cells. PPF MM show selective tumor accumulation in MCF-7 breast tumor bearing mice xenografts. Taken together, PPF MM represent a promising nanoscale carrier system to achieve GLUT5-mediated cell specific delivery in cancer therapy. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Rajendra, Yashas; Hougland, Maria D; Alam, Riazul; Morehead, Teresa A; Barnard, Gavin C
2015-05-01
Transient gene expression (TGE) is a rapid method for the production of recombinant proteins in mammalian cells. While the volumetric productivity of TGE has improved significantly over the past decade, most methods involve extensive cell line engineering and plasmid vector optimization in addition to long fed batch cultures lasting up to 21 days. Our colleagues have recently reported the development of a CHO K1SV GS-KO host cell line. By creating a bi-allelic glutamine synthetase knock out of the original CHOK1SV host cell line, they were able to improve the efficiency of generating high producing stable CHO lines for drug product manufacturing. We developed a TGE method using the same CHO K1SV GS-KO host cell line without any further cell line engineering. We also refrained from performing plasmid vector engineering. Our objective was to setup a TGE process to mimic protein quality attributes obtained from stable CHO cell line. Polyethyleneimine (PEI)-mediated transfections were performed at high cell density (4 × 10(6) cells/mL) followed by immediate growth arrest at 32 °C for 7 days. Optimizing DNA and PEI concentrations proved to be important. Interestingly, found the direct transfection method (where DNA and PEI were added sequentially) to be superior to the more common indirect method (where DNA and PEI are first pre-complexed). Moreover, the addition of a single feed solution and a polar solvent (N,N dimethylacetamide) significantly increased product titers. The scalability of process from 2 mL to 2 L was demonstrated using multiple proteins and multiple expression volumes. Using this simple, short, 7-day TGE process, we were able to successfully produce 54 unique proteins in a fraction of the time that would have been required to produce the respective stable CHO cell lines. The list of 54 unique proteins includes mAbs, bispecific antibodies, and Fc-fusion proteins. Antibody titers of up to 350 mg/L were achieved with the simple 7-day process. Titers
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Anticancer effects of oligomeric proanthocyanidins on human colorectal cancer cell line, SNU-C4
Kim, Youn-Jung; Park, Hae-Jeong; Yoon, Seo-Hyun; Kim, Mi-Ja; Leem, Kang-Hyun; Chung, Joo-Ho; Kim, Hye-Kyung
2005-01-01
AIM: Oligomeric proanthocyanidins (OPC), natural polyphenolic compounds found in plants, are known to have antioxidant and anti-cancer effects. We investigated whether the anti-cancer effects of the OPC are induced by apoptosis on human colorectal cancer cell line, SNU-C4. METHODS: Colorectal cancer cell line, SNU-C4 was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum. The cytotoxic effect of OPC was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenylt-etrazolium bromide (MTT) assay. To find out the apoptotic cell death, 4, 6-diamidino-2-phenylindole (DAPI) staining, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, reverse transcription-polymerase chain reaction (RT-PCR), and caspase-3 enzyme assay were performed. RESULTS: In this study, cytotoxic effect of OPC on SNU-C4 cells appeared in a dose-dependent manner. OPC treatment (100 µg/mL) revealed typical morphological apoptotic features. Additionally OPC treatment (100 µg/mL) increased level of BAX and CASPASE-3, and decreased level of BCL-2 mRNA expression. Caspase-3 enzyme activity was also significantly increased by treatment of OPC (100 µg/mL) compared with control. CONCLUSION: These data indicate that OPC caused cell death by apoptosis through caspase pathways on human colorectal cancer cell line, SNU-C4. PMID:16094708
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LINE-1 protein localization and functional dynamics during the cell cycle
Wudzinska, Aleksandra; Sun, Xiaoji; Andrade, Joshua; Nayak, Shruti; Kahler, David J; Badri, Sana; LaCava, John; Ueberheide, Beatrix; Yun, Chi Y; Fenyö, David
2018-01-01
LINE-1/L1 retrotransposon sequences comprise 17% of the human genome. Among the many classes of mobile genetic elements, L1 is the only autonomous retrotransposon that still drives human genomic plasticity today. Through its co-evolution with the human genome, L1 has intertwined itself with host cell biology. However, a clear understanding of L1’s lifecycle and the processes involved in restricting its insertion and intragenomic spread remains elusive. Here we identify modes of L1 proteins’ entrance into the nucleus, a necessary step for L1 proliferation. Using functional, biochemical, and imaging approaches, we also show a clear cell cycle bias for L1 retrotransposition that peaks during the S phase. Our observations provide a basis for novel interpretations about the nature of nuclear and cytoplasmic L1 ribonucleoproteins (RNPs) and the potential role of DNA replication in L1 retrotransposition. PMID:29309036
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Wissing, Silke; Montano, Mauricio; Garcia-Perez, Jose Luis; Moran, John V; Greene, Warner C
2011-10-21
Members of the APOBEC3 (A3) family of cytidine deaminase enzymes act as host defense mechanisms limiting both infections by exogenous retroviruses and mobilization of endogenous retrotransposons. Previous studies revealed that the overexpression of some A3 proteins could restrict engineered human Long INterspersed Element-1 (LINE-1 or L1) retrotransposition in HeLa cells. However, whether endogenous A3 proteins play a role in restricting L1 retrotransposition remains largely unexplored. Here, we show that HeLa cells express endogenous A3B and A3C, whereas human embryonic stem cells (hESCs) express A3B, A3C, A3DE, A3F, and A3G. To study the relative contribution of endogenous A3 proteins in restricting L1 retrotransposition, we first generated small hairpin RNAs (shRNAs) to suppress endogenous A3 mRNA expression, and then assessed L1 mobility using a cell-based L1 retrotransposition assay. We demonstrate that in both HeLa and hESCs, shRNA-based knockdown of A3B promotes a ∼2-3.7-fold increase in the retrotransposition efficiency of an engineered human L1. Knockdown of the other A3s produced no significant increase in L1 activity. Thus, A3B appears to restrict engineered L1 retrotransposition in a broad range of cell types, including pluripotent cells.
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Bulera, S J; Sattler, C A; Pitot, H C
1996-06-01
A transgenic rat line carrying the alb-SV40A transgene has been described by this laboratory. Several cell lines have been established from the livers of two of these rats. One of these cell lines, L37, exhibits a large nuclear/cytoplasmic ratio and a well-differentiated cytoplasm containing numerous organelles. When L37 cells are placed into culture medium lacking necessary growth factors, cellular proliferation continues for 48 hours after medium change. Subsequent to the initial 48 hours, cells begin to shrink and lose contact with adjacent cells, eventually sloughing off the culture plate surface, with most cell deaths occurring between 48 and 96 hours after medium change. Microscopic examination of sloughing cells indicates they possess highly convoluted and blebbed plasma membranes, a morphological characteristic of apoptosis. Ultrastructural studies demonstrate the ubiquitous presence of apoptotic bodies. When DNA isolated from growth factor-depleted cells is resolved on agarose gels, DNA fragmentation ladders are observed at times of maximum apoptotic change. Quantitative analysis of L37 cells between 48 and 96 hours after the removal of the culture medium shows that 59% +/- 2% of the cells undergo apoptosis. When cycloheximide, puromycin, or actinomycin D is added to the L37 cultures, only cycloheximide is able to repress apoptosis, indicating that the mechanism of apoptosis in the L37 liver-derived cell line requires a cycloheximide-sensitive translational event. The extremely high rate of apoptosis, together with the maintenance of hepatocellular characteristics, indicates the usefulness of this cell line as a model in which to study the mechanisms of hepatocellular apoptosis.
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History of leukemia-lymphoma cell lines.
Drexler, Hans G; Macleod, Roderick A F
2010-08-01
We outline the near 50-year history of leukemia-lymphoma (LL) cell lines - a key model system in biomedicine. Due to the detailed documentation of their oncogenomic and transcriptional alterations via recent advances in molecular medicine, LL cell lines may be fitted to parent tumors with a degree of precision unattainable in other cancers. We have surveyed the corpus of published LL cell lines and found 637 examples that meet minimum standards of authentication and characterization. Alarmingly, the rate of establishment of new LL cell lines has plummeted over the last decade. Although the main hematopoietic developmental cell types are represented by cell lines, some LL categories stubbornly resist establishment in vitro. The advent of engineering techniques for immortalizing primary human cells that maintain differentiation means the time is ripe for renewed search for in vitro models from un(der)represented hematologic entities. Given their manifold applications in biomedicine, there is little doubt that LL-derived cell lines will continue to play a vital part well into the next half-century as well. © 2010 The Authors. Human Cell © 2010 Japan Human Cell Society.
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Caffeine inhibition of GLUT1 is dependent on the activation state of the transporter.
Gunnink, Leesha K; Busscher, Brianna M; Wodarek, Jeremy A; Rosette, Kylee A; Strohbehn, Lauren E; Looyenga, Brendan D; Louters, Larry L
2017-06-01
Caffeine has been shown to be a robust uncompetitive inhibitor of glucose uptake in erythrocytes. It preferentially binds to the nucleotide-binding site on GLUT1 in its tetrameric form and mimics the inhibitory action of ATP. Here we demonstrate that caffeine is also a dose-dependent, uncompetitive inhibitor of 2-deoxyglucose (2DG) uptake in L929 fibroblasts. The inhibitory effect on 2DG uptake in these cells was reversible with a rapid onset and was additive to the competitive inhibitory effects of glucose itself, confirming that caffeine does not interfere with glucose binding. We also report for the first time that caffeine inhibition was additive to inhibition by curcumin, suggesting distinct binding sites for curcumin and caffeine. In contrast, caffeine inhibition was not additive to that of cytochalasin B, consistent with previous data that reported that these two inhibitors have overlapping binding sites. More importantly, we show that the magnitude of maximal caffeine inhibition in L929 cells is much lower than in erythrocytes (35% compared to 90%). Two epithelial cell lines, HCLE and HK2, have both higher concentrations of GLUT1 and increased basal 2DG uptake (3-4 fold) compared to L929 cells, and subsequently display greater maximal inhibition by caffeine (66-70%). Interestingly, activation of 2DG uptake (3-fold) in L929 cells by glucose deprivation shifted the responsiveness of these cells to caffeine inhibition (35%-70%) without a change in total GLUT1 concentration. These data indicate that the inhibition of caffeine is dependent on the activity state of GLUT1, not merely on the concentration. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.
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Khanna, Swati; Thomas, Anish; Abate-Daga, Daniel; Zhang, Jingli; Morrow, Betsy; Steinberg, Seth M; Orlandi, Augusto; Ferroni, Patrizia; Schlom, Jeffrey; Guadagni, Fiorella; Hassan, Raffit
2016-11-01
The functional aspects of programmed death 1 (PD-1) and PD ligand 1 (PD-L1) immune checkpoints in malignant mesothelioma have not been studied. Tumor samples from 65 patients with mesothelioma were evaluated for PD-L1 expression by immunohistochemistry, and its prognostic significance was examined. Malignant effusions from patients with pleural and peritoneal mesothelioma were evaluated for PD-1-positive and PD-L1-positive infiltrating lymphocytes and their role in inducing PD-L1 expression in tumor cells. Antibody-dependent cellular cytotoxicity (ADCC) of avelumab, a fully humanized immunoglobulin G1 anti PD-L1 antibody against primary mesothelioma cell lines, was evaluated in presence of autologous and allogeneic natural killer cells. Of 65 pleural and peritoneal mesothelioma tumors examined, 41 (63%) were PD-L1-positive, which was associated with slightly inferior overall survival compared to patients with PD-L1-negative tumors (median 23.0 versus 33.3 months, p = 0.35). The frequency of PD-L1 expression was similar in patients with pleural and peritoneal mesothelioma, with 62% and 64% of samples testing positive, respectively. In nine mesothelioma effusion samples evaluated, the fraction of cells expressing PD-L1 ranged from 12% to 83%. In seven patients with paired malignant effusion and peripheral blood mononuclear cell (PBMC) samples, PD-L1 expression was significantly higher on CD3-positive T cells present in malignant effusions as compared with PBMCs (p = 0.016). In addition, the numbers of CD14-positive PD-1-positive cells were increased in malignant effusions compared with PBMCs (p = 0.031). The lymphocytes present in malignant effusions recognized autologous tumor cells and induced interferon-γ-mediated PD-L1 expression on the tumor cell surface. Of the three primary mesothelioma cell lines tested, two were susceptible to avelumab-mediated ADCC in the presence of autologous natural killer cells. Most pleural as well as peritoneal mesotheliomas
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Code of Federal Regulations, 2011 CFR
2011-10-01
... Assessment for Stress Corrosion Cracking (SCCDA)? 192.929 Section 192.929 Transportation Other Regulations...: MINIMUM FEDERAL SAFETY STANDARDS Gas Transmission Pipeline Integrity Management § 192.929 What are the requirements for using Direct Assessment for Stress Corrosion Cracking (SCCDA)? (a) Definition. Stress...
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Code of Federal Regulations, 2012 CFR
2012-10-01
... Assessment for Stress Corrosion Cracking (SCCDA)? 192.929 Section 192.929 Transportation Other Regulations...: MINIMUM FEDERAL SAFETY STANDARDS Gas Transmission Pipeline Integrity Management § 192.929 What are the requirements for using Direct Assessment for Stress Corrosion Cracking (SCCDA)? (a) Definition. Stress...
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Code of Federal Regulations, 2014 CFR
2014-10-01
... Assessment for Stress Corrosion Cracking (SCCDA)? 192.929 Section 192.929 Transportation Other Regulations...: MINIMUM FEDERAL SAFETY STANDARDS Gas Transmission Pipeline Integrity Management § 192.929 What are the requirements for using Direct Assessment for Stress Corrosion Cracking (SCCDA)? (a) Definition. Stress...
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Code of Federal Regulations, 2013 CFR
2013-10-01
... Assessment for Stress Corrosion Cracking (SCCDA)? 192.929 Section 192.929 Transportation Other Regulations...: MINIMUM FEDERAL SAFETY STANDARDS Gas Transmission Pipeline Integrity Management § 192.929 What are the requirements for using Direct Assessment for Stress Corrosion Cracking (SCCDA)? (a) Definition. Stress...
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The neuroendocrine phenotype, genomic profile and therapeutic sensitivity of GEPNET cell lines
Hofving, Tobias; Arvidsson, Yvonne; Almobarak, Bilal; Inge, Linda; Pfragner, Roswitha; Persson, Marta; Stenman, Göran; Kristiansson, Erik; Johanson, Viktor; Nilsson, Ola
2018-01-01
Experimental models of neuroendocrine tumour disease are scarce, and no comprehensive characterisation of existing gastroenteropancreatic neuroendocrine tumour (GEPNET) cell lines has been reported. In this study, we aimed to define the molecular characteristics and therapeutic sensitivity of these cell lines. We therefore performed immunophenotyping, copy number profiling, whole-exome sequencing and a large-scale inhibitor screening of seven GEPNET cell lines. Four cell lines, GOT1, P-STS, BON-1 and QGP-1, displayed a neuroendocrine phenotype while three others, KRJ-I, L-STS and H-STS, did not. Instead, these three cell lines were identified as lymphoblastoid. Characterisation of remaining authentic GEPNET cell lines by copy number profiling showed that GOT1, among other chromosomal alterations, harboured losses on chromosome 18 encompassing the SMAD4 gene, while P-STS had a loss on 11q. BON-1 had a homozygous loss of CDKN2A and CDKN2B, and QGP-1 harboured amplifications of MDM2 and HMGA2. Whole-exome sequencing revealed both disease-characteristic mutations (e.g. ATRX mutation in QGP-1) and, for patient tumours, rare genetic events (e.g. TP53 mutation in P-STS, BON-1 and QGP-1). A large-scale inhibitor screening showed that cell lines from pancreatic NETs to a greater extent, when compared to small intestinal NETs, were sensitive to inhibitors of MEK. Similarly, neuroendocrine NET cells originating from the small intestine were considerably more sensitive to a group of HDAC inhibitors. Taken together, our results provide a comprehensive characterisation of GEPNET cell lines, demonstrate their relevance as neuroendocrine tumour models and explore their therapeutic sensitivity to a broad range of inhibitors. PMID:29444910
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Valiahdi, Seied Mojtaba; Heffeter, Petra; Jakupec, Michael A.; Marculescu, Rodrig; Berger, Walter; Rappersberger, Klemens; Keppler, Bernhard K.
2012-01-01
The antineoplastic properties of gallium are well documented. Owing to their robust accumulation of gallium, melanoma cells should be amenable to gallium-based anticancer drugs. With the aim of improving the disappointingly low activity of inorganic gallium salts, we have developed the orally bioavailable gallium complex KP46 [tris(8-quinolinolato)gallium(III)] that was already successfully studied in a phase I clinical trial. To assess its therapeutic potential in malignant melanoma, its antiproliferative effects were investigated in series of human cell lines and primary explanted melanoma samples by means of the MTT [3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] assay and the Human Tumor Cloning Assay, respectively. When compared with other cell lines, the majority of melanoma cells rank among the KP46-sensitive cell lines (50% inhibitory concentration values: 0.8–3.7 μmol/l). Clinically achievable concentrations of KP46 proved to be highly effective in melanoma cells from primary explants of cutaneous and lymph node metastases. Colony growth was inhibited in 10 of 10 specimens by 5 lmol/l KP46 (corresponding to the steady-state plasma concentration measured earlier in a study patient) and in four of 10 specimens by 0.5 μmol/l KP46. In-vitro potency of KP46 is higher than that of dacarbazine or fotemustine and comparable with that of cisplatin. The effects induced by KP46 in melanoma cell lines involve cell cycle perturbations (S-phase arrest) and apoptosis (activation of caspase-9, PARP [poly(ADP-ribose) polymerase] cleavage, formation of apoptotic bodies). No effects on DNA secondary structure could be observed in an electrophoretic mobility shift assay using double-stranded plasmid DNA. Thus, further studies on the therapeutic applicability of KP46 in malignant melanoma are warranted. PMID:19584767
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Vega, Mario I; Jazirehi, Ali R; Huerta-Yepez, Sara; Bonavida, Benjamin
2005-08-15
Rituximab treatment of B non-Hodgkin's lymphoma (NHL) cell lines inhibits the constitutive NF-kappaB activity and results in the sensitization of tumor cells to both chemotherapy and Fas-induced apoptosis. Cells expressing dominant active IkappaB or treated with NF-kappaB-specific inhibitors were sensitive to both drugs and Fas agonist mAb (CH-11)-induced apoptosis. Down-regulation of Bcl-xL expression via inhibition of NF-kappaB activity correlated with chemosensitivity. The direct role of Bcl-xL in chemoresistance was demonstrated by the use of Bcl-xL-overexpressing Ramos cells, Ramos hemagglutinin (HA)-Bcl-x, which were not sensitized by rituximab to drug-induced apoptosis. However, inhibition of Bcl-xL in Ramos HA-Bcl-x resulted in sensitization to drug-induced apoptosis. The role of Bcl-xL expression in the regulation of Fas resistance was not apparent; Ramos HA-Bcl-x cells were as sensitive as the wild type to CH-11-induced apoptosis. Several lines of evidence support the direct role of the transcription repressor yin-yang 1 (YY1) in the regulation of resistance to CH-11-induced apoptosis. Inhibition of YY1 activity by either rituximab or the NO donor DETANONOate or after transfection with YY1 small interfering RNA resulted in up-regulation of Fas expression and sensitization to CH-11-induced apoptosis. These findings suggest two mechanisms underlying the chemosensitization and immunosensitization of B-NHL cells by rituximab via inhibition of NF-kappaB. The regulation of chemoresistance by NF-kappaB is mediated via Bcl-xL expression, whereas the regulation of Fas resistance by NF-kappaB is mediated via YY1 expression and activity. The potential clinical significance of these findings is discussed.
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Cytotoxic effect of Argentine medicinal plant extracts on human hepatocellular carcinoma cell line.
Ruffa, M J; Ferraro, G; Wagner, M L; Calcagno, M L; Campos, R H; Cavallaro, L
2002-03-01
Methanolic extracts from Achyrocline satureioides (Dc.) Lam, Aristolochia macroura Gomez, Lithraea molleoides (Vell.) Engl., Schinus molle L., unlike those from Celtis spinosa Spreng, Chenopodium ambrosioides L., Petiveria alliacea L., and Plantago major L. showed cytotoxic activity against a human hepatocellular carcinoma cell line, Hep G2. Schinus molle L. was the most active (IC50=50+/-7 microg/ml). These results call for further studies of these extracts.
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Park, Jun Won; Um, Hyejin; Yang, Hanna; Ko, Woori; Kim, Dae-Yong; Kim, Hark Kyun
2017-03-11
To elucidate signaling pathways that regulate gastric cancer stem cell (CSC) phenotypes and immune checkpoint, we performed a proteogenomic analysis of NCC-S1M, which is a gastric cancer cell line with CSC-like characteristics and is the only syngeneic gastric tumor cell line transplant model created in the scientific community. We found that the NCC-S1M allograft was responsive to anti-PD-1 treatment, and overexpressed Cd274 encoding PD-L1. PD-L1 was transcriptionally activated by loss of the TGF-β signaling. Il1rl1 protein was overexpressed in NCC-S1M cells compared with NCC-S1 cells that are less tumorigenic and less chemoresistant. Il1rl1 knockdown in NCC-S1M cells reduced tumorigenic potential and in vivo chemoresistance. Our proteogenomic analysis demonstrates a role of Smad4 loss in the PD-L1 immune evasion, as well as Il1rl1's role in CSC-like properties of NCC-S1M. Copyright © 2017 Elsevier Inc. All rights reserved.
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Busson, Laurent; Crucitti, Tania; De Foor, Marc; Van den Wijngaert, Sigi; Vandenberg, Olivier
2013-08-01
This article reports the fortuitous recovery of nine Chlamydia trachomatis serovar L strains in cell cultures (Vero and LLC-MK(2) cell line) designed for viral culture. Nine ano-genital swabs were inoculated on confluent Vero, MRC5 and LLC-MK(2) cell cultures. They were collected from HIV-positive patients who were primarily men who have sex with men (MSM) presenting ulcerations that mimicked herpes simplex infections. A cytopathogenic effect was observed on Vero and LLC-MK(2) cells on day 14. The presence of C trachomatis serovar L in the cell lines was confirmed by Real Time-PCR. C trachomatis serovar L can grow on Vero and LLC-MK(2) cell lines designed for viral cultures. Lymphogranuloma venereum must be considered as a differential diagnosis for herpes-like lesions, particularly in MSM with high-risk behaviours.
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Aguiñiga-Sánchez, Itzen; Soto-Hernández, Marcos; Cadena-Iñiguez, Jorge; Ruíz-Posadas, Lucero del Mar; Cadena-Zamudio, Jorge David; González-Ugarte, Ana Karen; Steider, Benny Weiss; Santiago-Osorio, Edelmiro
2015-01-01
The antiproliferative potential of a crude extract from the chayote hybrid H-837-07-GISeM® and its potential for apoptosis induction were assessed in leukaemic cell lines and normal mouse bone marrow mononuclear cells (BM-MNCs). The extract strongly inhibited the proliferation of the P388, J774, and WEHI-3 cell lines (with an IC50 below 1.3 μg·mL(-1)), reduced cell viability, and induced apoptotic body production, phosphatidylserine translocation, and DNA fragmentation. However, the extract had no effect on BM-MNCs. We postulate that these properties make the extract a good candidate for an anti-tumour agent for clinical use.
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Abdullah, Al-Shwyeh Hussah; Mohammed, Abdulkarim Sabo; Abdullah, Rasedee; Mirghani, Mohamed Elwathig Saeed; Al-Qubaisi, Mothanna
2014-06-25
Waterlily Mango (Mangifera indica L.) is thought to be antioxidant-rich, conferred by its functional phytochemicals. The potential anticancer effects of the ethanolic kernel extract on breast cancer cells (MDA-MB-231 and MCF-7) using MTT, anti-proliferation, neutral red (NR) uptake and lactate dehydrogenase (LDH) release assays were evaluated. Cytological studies on the breast cancer cells were also conducted, and phytochemical analyses of the extract were carried out to determine the likely bioactive compounds responsible for such effects. Results showed the extract induced cytotoxicity in MDA-MB-231 cells and MCF-7 cells with IC50 values of 30 and 15 μg/mL, respectively. The extract showed significant toxicity towards both cell lines, with low toxicity to normal breast cells (MCF-10A). The cytotoxic effects on the cells were further confirmed by the NR uptake, antiproliferative and LDH release assays. Bioactive analyses revealed that many bioactives were present in the extract although butylated hydroxytoluene, a potent antioxidant, was the most abundant with 44.65%. M. indica extract appears to be more cytoxic to both estrogen positive and negative breast cancer cell lines than to normal breast cells. Synergistic effects of its antioxidant bioactives could have contributed to the cytotoxic effects of the extract. The extract of M. indica, therefore, has potential anticancer activity against breast cancer cells. This potential is worth studying further, and could have implications on future studies and eventually management of human breast cancers.
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Comparative Study of the Biological Activity of Allantoin and Aqueous Extract of the Comfrey Root.
Savić, Vesna Lj; Nikolić, Vesna D; Arsić, Ivana A; Stanojević, Ljiljana P; Najman, Stevo J; Stojanović, Sanja; Mladenović-Ranisavljević, Ivana I
2015-08-01
This study investigates the biological activity of pure allantoin (PA) and aqueous extract of the comfrey (Symphytum officinale L.) root (AECR) standardized to the allantoin content. Cell viability and proliferation of epithelial (MDCK) and fibroblastic (L929) cell line were studied by using MTT test. Anti-irritant potential was determined by measuring electrical capacitance, erythema index (EI) and transepidermal water loss of artificially irritated skin of young healthy volunteers, 3 and 7 days after application of creams and gels with PA or AECR. Pure allantoin showed mild inhibitory effect on proliferation of both cell lines at concentrations 40 and 100 µg/ml, but more pronounced on MDCK cells. Aqueous extract of the comfrey root effect on cell proliferation in concentrations higher than 40 µg/ml was significantly stimulatory for L929 but inhibitory for MDCK cells. Pharmaceutical preparations that contained AECR showed better anti-irritant potential compared with PA. Creams showed better effect on hydration and EI compared with the gels that contained the same components. Our results indicate that the biological activity of the comfrey root extract cannot be attributed only to allantoin but is also likely the result of the interaction of different compounds present in AECR. Topical preparations that contain comfrey extract may have a great application in the treatment of skin irritation. Copyright © 2015 John Wiley & Sons, Ltd.
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Cancer stem cell-like cells from a single cell of oral squamous carcinoma cell lines
DOE Office of Scientific and Technical Information (OSTI.GOV)
Felthaus, O.; Department of Oral and Maxillofacial Surgery, University of Regensburg; Ettl, T.
2011-04-01
Research highlights: {yields} Four oral squamous cancer cell lines (OSCCL) were analyzed for cancer stem cells (CSCs). {yields} Single cell derived colonies of OSCCL express CSC-marker CD133 differentially. {yields} Monoclonal cell lines showed reduced sensitivity for Paclitaxel. {yields} In situ CD133{sup +} cells are slow cycling (Ki67-) indicating a reduced drug sensitivity. {yields} CD133{sup +} and CSC-like cells can be obtained from single colony forming cells of OSCCL. -- Abstract: Resistance of oral squamous cell carcinomas (OSCC) to conventional chemotherapy or radiation therapy might be due to cancer stem cells (CSCs). The development of novel anticancer drugs requires a simplemore » method for the enrichment of CSCs. CSCs can be enriched from OSCC cell lines, for example, after cultivation in serum-free cell culture medium (SFM). In our study, we analyzed four OSCC cell lines for the presence of CSCs. CSC-like cells could not be enriched with SFM. However, cell lines obtained from holoclone colonies showed CSC-like properties such as a reduced rate of cell proliferation and a reduced sensitivity to Paclitaxel in comparison to cells from the parental lineage. Moreover, these cell lines differentially expressed the CSC-marker CD133, which is also upregulated in OSCC tissues. Interestingly, CD133{sup +} cells in OSCC tissues expressed little to no Ki67, the cell proliferation marker that also indicates reduced drug sensitivity. Our study shows a method for the isolation of CSC-like cell lines from OSCC cell lines. These CSC-like cell lines could be new targets for the development of anticancer drugs under in vitro conditions.« less
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Production of thrombopoietin (TPO) by rat hepatocytes and hepatoma cell lines.
Shimada, Y; Kato, T; Ogami, K; Horie, K; Kokubo, A; Kudo, Y; Maeda, E; Sohma, Y; Akahori, H; Kawamura, K
1995-12-01
Recently, we purified rat thrombopoietin (TPO) from plasma of irradiated rats (XRP) by measuring its activity that stimulated the production of megakaryocytes from megakaryocyte progenitor cells (CFU-MK) in vitro. We then cloned the cDNAs for rat and human TPO. In this study, we found the production of TPO by hepatocytes isolated with the collagenase perfusion method from both normal and thrombocytopenic rats, by a two-step fractionation of hepatocyte culture medium (CM). Subsequently, CM of rat hepatoma cell lines was screened for the presence of TPO; three cell lines, H4-II-E, McA-RH8994, and HTC, were found to produce TPO. According to the purification procedure for TPO from XRP, TPO was partially purified from 2 L CM of each of three cell lines with a six-step procedure. In the final reverse-phase column, TPO from each cell line was eluted with the same retention time as that from XRP, and the TPO fraction exhibited megakaryocyte colony-stimulating activity (Meg-CSA). TPO-active fraction eluted from the final reverse-phase column was separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), extracted from the gel, and assayed. TPO activity from each cell line was found in the respective molecular weight region, indicating the heterogeneity of the TPO molecule. Using reverse transcriptase-polymerase chain reaction (RT-PCR), we detected the expression of TPO mRNA in hepatocytes, three hepatoma cell lines, normal rat liver, and X-irradiated rat liver. Northern blot analysis showed that TPO mRNA was expressed mainly in liver among the various organs tested. These data demonstrate that TPO is produced by rat hepatocytes and hepatoma cell lines and suggest that liver may be the primary organ that produces TPO.
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Taju, G; Abdul Majeed, S; Nambi, K S N; Sahul Hameed, A S
2017-10-01
In the present study, we hypothesize that cytotoxicity, genotoxicity and oxidative stress play a key role in chromium induced toxicity in SISS, SISK, IEE, IEK, IEG, SICH and ICG cell lines after 24 h exposure. Three fish species namely Lates calcarifer, Etroplus suratensis and Catla catla were exposed to the concentrations of 0, 10, 20, 30, 40 and 50 mg/L of chromium for 96 h under static conditions for conducting acute toxicity tests. LC 50 was then calculated. The percentage cell survival was assessed by multiple endpoints such as MTT, NR, AB and CB assays in the seven fish cell lines exposed to different concentrations of chromium and EC 50 values of all the four endpoints were calculated. High significances were noted in the correlations between each in vitro cytotoxicity assays and in vivo mortality data. Cell shrinkage, cell detachment, vacuolations and cell swelling at the highest concentration of chromium (50 mg/L) were seen on microscopic examination of cell morphology. Comet assay and Hoechst staining were carried out to assess DNA damage and nuclear fragmentation in the seven fish lines exposed to chromium. The results of antioxidant parameters obtained indicate a significant reduction in the level of catalase, superoxide dismutase, glutathione S-transferase and Glutathione peroxidase, and increased level of lipid peroxidation in all the cell lines exposed to chromium. These results confirm that fish cell lines could be used as an alternative to whole fish for cytotoxicity, genotoxicity and oxidative stress assessment in chromium toxicity studies. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Wissing, Silke; Montano, Mauricio; Garcia-Perez, Jose Luis; Moran, John V.; Greene, Warner C.
2011-01-01
Members of the APOBEC3 (A3) family of cytidine deaminase enzymes act as host defense mechanisms limiting both infections by exogenous retroviruses and mobilization of endogenous retrotransposons. Previous studies revealed that the overexpression of some A3 proteins could restrict engineered human Long INterspersed Element-1 (LINE-1 or L1) retrotransposition in HeLa cells. However, whether endogenous A3 proteins play a role in restricting L1 retrotransposition remains largely unexplored. Here, we show that HeLa cells express endogenous A3B and A3C, whereas human embryonic stem cells (hESCs) express A3B, A3C, A3DE, A3F, and A3G. To study the relative contribution of endogenous A3 proteins in restricting L1 retrotransposition, we first generated small hairpin RNAs (shRNAs) to suppress endogenous A3 mRNA expression, and then assessed L1 mobility using a cell-based L1 retrotransposition assay. We demonstrate that in both HeLa and hESCs, shRNA-based knockdown of A3B promotes a ∼2–3.7-fold increase in the retrotransposition efficiency of an engineered human L1. Knockdown of the other A3s produced no significant increase in L1 activity. Thus, A3B appears to restrict engineered L1 retrotransposition in a broad range of cell types, including pluripotent cells. PMID:21878639
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Góes, Luiz Gustavo Bentim; de Freitas, Antonio Carlos; Ferraz, Oilita Pereira; Rieger, Tania Tassinari; dos Santos, José Ferreira; Pereira, Alexandre; Beçak, Willy; Lindsey, Charles J.; de Cassia Stocco, Rita
2008-01-01
The development of a bovine papillomavirus (BPV) vaccine is an outstanding challenge. BPV protein L1 gene transfection in the Drosophila melanogaster S2 cell expression system failed to produce L1 protein notwithstanding correct L1 gene insertion. Severe genetic inbalance in the host cell line, including cytogenetic alterations, may account for the lack of protein expression. PMID:24031166
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Development and characterization of a cell line TTCF from endangered mahseer Tor tor (Ham.).
Yadav, K; Lakra, W S; Sharma, J; Goswami, M; Singh, Akhilesh
2012-08-01
Tor tor is an important game and food fish of India with a distribution throughout Asia from the trans-Himalayan region to the Mekong River basin to Malaysia, Pakistan, Bangladesh and Indonesia. A new cell line named TTCF was developed from the caudal fin of T. tor for the first time. The cell line was optimally maintained at 28°C in Leibovitz-15 (L-15) medium supplemented with 20% fetal bovine serum (FBS). The propagation of TTCF cells showed a high plating efficiency of 63.00%. The cytogenetic analysis revealed a diploid count of 100 chromosomes at passage 15, 30, 45 and 60 passages. The viability of the TTCF cell line was found to be 72% after 6 months of cryopreservation in liquid nitrogen (-196°C). The origin of the cell lines was confirmed by the amplification of 578- and 655-bp sequences of 16S rRNA and cytochrome oxidase subunit I (COI) genes of mitochondrial DNA (mtDNA) respectively. TTCF cells were successfully transfected with green fluorescent protein (GFP) reporter plasmids. Further, immunocytochemistry studies confirm its fibroblastic morphology of cells. Genotoxicity assessment of H₂O₂ in TTCF cell line revealed the utility of TTCF cell line as in vitro model for aquatic toxicological studies.
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NASA Astrophysics Data System (ADS)
Artanti, A. N.; Astirin, O. P.; Prayito, A.; Widiyaningsih, R. F.; Prihapsara, F.
2017-02-01
One of the compounds found effication as an anticancer agent on cervical cancer is acetogenin, a polyketide compound that is abundant in Annona muricata L. leaves. This study has been done to examine polyketide derivatives was isolated from Annona muricata L. which has potency to induce apoptosis by p53 expression on hela cell line. An approach recently develop to overcome side effect of chemoterapeutic agent is used of combined chemoterapeutic agent, i.e doxorubicin. The determination of cytotoxic combination activity from polyketide derivative and doxorubicin was evaluated using MTT assay to obtain the value of CI (combination index). The expression of p53 profile was evaluated by immunohistochemistry on hela cell line. Data analysis showed that combination of polyketide derivative from Annona muricata L. (38,5 µg/ml) and doxorubicin with all of concentration performed synergistic effect on hela cell line with CI value from 0,33 - 0,65. The analysis on immucytochemistry showed that polyketide derivative from Annona muricata L. leaves could enhance p53 pathway significantly on hela cell line.
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Lavdas, Alexandros A; Efrose, Rodica; Douris, Vassilis; Gaitanou, Maria; Papastefanaki, Florentia; Swevers, Luc; Thomaidou, Dimitra; Iatrou, Kostas; Matsas, Rebecca
2010-12-01
For biotechnological applications, insect cell lines are primarily known as hosts for the baculovirus expression system that is capable to direct synthesis of high levels of recombinant proteins through use of powerful viral promoters. Here, we demonstrate the implementation of two alternative approaches based on the baculovirus system for production of a mammalian recombinant glycoprotein, comprising the extracellular part of the cell adhesion molecule L1, with potential important therapeutic applications in nervous system repair. In the first approach, the extracellular part of L1 bearing a myc tag is produced in permanently transformed insect cell lines and purified by affinity chromatography. In the second approach, recombinant baculoviruses that express L1-Fc chimeric protein, derived from fusion of the extracellular part of L1 with the Fc part of human IgG1, under the control of a mammalian promoter are used to infect mammalian HEK293 and primary Schwann cells. Both the extracellular part of L1 bearing a myc tag accumulating in the supernatants of insect cultures as well as L1-Fc secreted by transduced HEK293 or Schwann cells are capable of increasing the motility of Schwann cells with similar efficiency in a gap bridging bioassay. In addition, baculovirus-transduced Schwann cells show enhanced motility when grafted on organotypic cultures of neonatal brain slices while they retain their ability to myelinate CNS axons. This proof-of-concept that the migratory properties of myelin-forming cells can be modulated by recombinant protein produced in insect culture as well as by means of baculovirus-mediated adhesion molecule expression in mammalian cells may have beneficial applications in the field of CNS therapies. ©2010 The Authors. Journal of Neurochemistry © 2010 International Society for Neurochemistry.
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Cold-induced retrotransposition of fish LINEs.
Chen, Shue; Yu, Mengchao; Chu, Xu; Li, Wenhao; Yin, Xiujuan; Chen, Liangbiao
2017-08-20
Classes of retrotransposons constitute a large portion of metazoan genome. There have been cases reported that genomic abundance of retrotransposons is correlated with the severity of low environmental temperatures. However, the molecular mechanisms underlying such correlation are unknown. We show here by cell transfection assays that retrotransposition (RTP) of a long interspersed nuclear element (LINE) from an Antarctic notothenioid fish Dissostichus mawsoni (dmL1) could be activated by low temperature exposure, causing increased dmL1 copies in the host cell genome. The cold-induced dmL1 propagation was demonstrated to be mediated by the mitogen-activated protein kinases (MAPK)/p38 signaling pathway, which is activated by accumulation of reactive oxygen species (ROS) in cold-stressed conditions. Surprisingly, dmL1 transfected cells showed an increase in the number of viable cells after prolonged cold exposures than non-transfected cells. Features of cold inducibility of dmL1 were recapitulated in LINEs of zebrafish origin both in cultured cell lines and tissues, suggesting existence of a common cold-induced LINE amplification in fishes. The findings reveal an important function of LINEs in temperature adaptation and provid insights into the MAPK/p38 stress responsive pathway that shapes LINE composition in fishes facing cold stresses. Copyright © 2017. Published by Elsevier Ltd.
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38 CFR 1.929 - Reduction of debt through performance of work-study services.
Code of Federal Regulations, 2011 CFR
2011-07-01
... for the performance of work-study services after the completion of each 50 hours of services (or in... performance of work-study services. 1.929 Section 1.929 Pensions, Bonuses, and Veterans' Relief DEPARTMENT OF... performance of work-study services. (a) Scope. (1) Subject to the provisions of this section VA may allow an...
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Development and characterization of two cell lines from gills of Atlantic salmon
Gjessing, Mona C.; Aamelfot, Maria; Batts, William N.; Benestad, Sylvie L.; Dale, Ole B.; Thoen, Even; Weli, Simon C.; Winton, James R.
2018-01-01
Gill disease in Atlantic salmon, Salmo salar L., causes big losses in the salmon farming industry. Until now, tools to cultivate microorganisms causing gill disease and models to study the gill responses have been lacking. Here we describe the establishment and characterization of two cell lines from the gills of Atlantic salmon. Atlantic salmon gill cell ASG-10 consisted of cells staining for cytokeratin and e-cadherin and with desmosomes as seen by transmission electron microscopy suggesting the cells to be of epithelial origin. These structures were not seen in ASG-13. The cell lines have been maintained for almost 30 passages and both cell lines are fully susceptible to infection by infectious hematopoietic necrosis virus (IHNV), viral hemorrhagic septicemia virus (VHSV), infectious pancreatic necrosis virus (IPNV), Atlantic salmon reovirus TS (TSRV) and Pacific salmon paramyxovirus (PSPV). While infectious salmon anemia virus (ISAV) did not cause visible CPE, immunofluorescent staining revealed a sub-fraction of cells in both the ASG-10 and ASG-13 lines may be permissive to infection. ASG-10 is able to proliferate and migrate to close scratches in the monolayer within seven days in vitro contrary to ASG-13, which does not appear to do have the same proliferative and migratory ability. These cell lines will be useful in studies of gill diseases in Atlantic salmon and may represent an important contribution for alternatives to experimental animals and studies of epithelial–mesenchymal cell biology.
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Kim, Dong-Hyun; Kim, Kyoung-Nam; Kim, Kwang-Mahn; Lee, Yong-Keun
2009-01-01
The delivery of hyperthermic thermoseeds to a specific target site with minimal side effects is an important challenge in targeted hyperthermia, which employs magnetic method and functional polymers. An external magnetic field is used to control the site-specific targeting of the magnetic nanoparticles. Polymer-coated magnetic nanoparticles can confer a higher affinity to the biological cell membranes. In this study, uncoated, chitosan-coated, and starch-coated magnetic nanoparticles were synthesized for use as a hyperthermic thermoseed. Each sample was examined with respect to their applications to hyperthermia using XRD, VSM, and FTIR. In addition, the temperature changes under an alternating magnetic field were observed. As in vitro tests, the magnetic responsiveness of chitosan- and starch-coated magnetite was determined by a simple blood vessel model under various intensities of magnetic field. L929 normal cells and KB carcinoma cells were used to examine the cytotoxicity and affinity of each sample using the MTT method. The chitosan-coated magnetic nanoparticles generated a higher DeltaT of 23 degrees C under an AC magnetic field than the starch-coated magnetite, and the capturing rate of the particles was 96% under an external magnetic field of 0.4 T. The highest viability of L929 cells was 93.7%. Comparing the rate of KB cells capture with the rate of L929 cells capture, the rate of KB cells capture relatively increased with 10.8% in chitosan-coated magnetic nanoparticles. Hence, chitosan-coated magnetic nanoparticles are biocompatible and have a selective affinity to KB cells. The targeting of magnetic nanoparticles in hyperthermia was improved using a controlled magnetic field and a chitosan-coating. Therefore, chitosan-coated magnetic nanoparticles are expected to be promising materials for use in magnetic targeted hyperthermia. 2008 Wiley Periodicals, Inc.
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Chen, Lindi; Humphreys, Angharad; Turnbull, Lisa; Bellini, Angela; Schleiermacher, Gudrun; Salwen, Helen; Cohn, Susan L; Bown, Nick; Tweddle, Deborah A
2016-12-27
Anaplastic Lymphoma Kinase (ALK) is a transmembrane receptor kinase that belongs to the insulin receptor superfamily and has previously been shown to play a role in cell proliferation, migration and invasion in neuroblastoma. Activating ALK mutations are reported in both hereditary and sporadic neuroblastoma tumours, and several ALK inhibitors are currently under clinical evaluation as novel treatments for neuroblastoma. Overall, mutations at codons F1174, R1275 and F1245 together account for ~85% of reported ALK mutations in neuroblastoma. NBLW and NBLW-R are paired cell lines originally derived from an infant with metastatic MYCN amplified Stage IVS (Evans Criteria) neuroblastoma, at diagnosis and relapse, respectively. Using both Sanger and targeted deep sequencing, this study describes the identification of distinct ALK mutations in these paired cell lines, including the rare R1275L mutation, which has not previously been reported in a neuroblastoma cell line. Analysis of the sensitivity of NBLW and NBLW-R cells to a panel of ALK inhibitors (TAE-684, Crizotinib, Alectinib and Lorlatinib) revealed differences between the paired cell lines, and overall NBLW-R cells with the F1174L mutation were more resistant to ALK inhibitor induced apoptosis compared with NBLW cells. This pair of cell lines represents a valuable pre-clinical model of clonal evolution of ALK mutations associated with neuroblastoma progression.
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7 CFR 929.51 - Recommendations for regulation.
Code of Federal Regulations, 2012 CFR
2012-01-01
... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... advisable to regulate the handling of cranberries in the manner provided in § 929.52, it shall so recommend... the factors affecting the supply of and demand for cranberries during the period when it is proposed...
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7 CFR 929.51 - Recommendations for regulation.
Code of Federal Regulations, 2014 CFR
2014-01-01
... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... advisable to regulate the handling of cranberries in the manner provided in § 929.52, it shall so recommend... the factors affecting the supply of and demand for cranberries during the period when it is proposed...
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7 CFR 929.51 - Recommendations for regulation.
Code of Federal Regulations, 2013 CFR
2013-01-01
... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE CRANBERRIES GROWN IN STATES OF... advisable to regulate the handling of cranberries in the manner provided in § 929.52, it shall so recommend... the factors affecting the supply of and demand for cranberries during the period when it is proposed...
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Fujii, Rika; Friedman, Eitan R; Richards, Jacob; Tsang, Kwong Y; Heery, Christopher R; Schlom, Jeffrey; Hodge, James W
2016-06-07
Chordoma, a rare bone tumor derived from the notochord, has been shown to be resistant to conventional therapies. Checkpoint inhibition has shown great promise in immune-mediated therapy of diverse cancers. The anti-PD-L1 mAb avelumab is unique among checkpoint inhibitors in that it is a fully human IgG1 capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC) of PD-L1-expressing tumor cells. Here, we investigated avelumab as a potential therapy for chordoma. We examined 4 chordoma cell lines, first for expression of PD-L1, and in vitro for ADCC killing using NK cells and avelumab. PD-L1 expression was markedly upregulated by IFN-γ in all 4 chordoma cell lines, which significantly increased sensitivity to ADCC. Brachyury is a transcription factor that is uniformly expressed in chordoma. Clinical trials are ongoing in which chordoma patients are treated with brachyury-specific vaccines. Co-incubating chordoma cells with brachyury-specific CD8+ T cells resulted in significant upregulation of PD-L1 on the tumor cells, mediated by the CD8+ T cells' IFN-γ production, and increased sensitivity of chordoma cells to avelumab-mediated ADCC. Residential cancer stem cell subpopulations of chordoma cells were also killed by avelumab-mediated ADCC to the same degree as non-cancer stem cell populations. These findings suggest that as a monotherapy for chordoma, avelumab may enable endogenous NK cells, while in combination with T-cell immunotherapy, such as a vaccine, avelumab may enhance NK-cell killing of chordoma cells via ADCC.
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Fujii, Rika; Friedman, Eitan R.; Richards, Jacob; Tsang, Kwong Y.; Heery, Christopher R.; Schlom, Jeffrey; Hodge, James W.
2016-01-01
Chordoma, a rare bone tumor derived from the notochord, has been shown to be resistant to conventional therapies. Checkpoint inhibition has shown great promise in immune-mediated therapy of diverse cancers. The anti-PD-L1 mAb avelumab is unique among checkpoint inhibitors in that it is a fully human IgG1 capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC) of PD-L1-expressing tumor cells. Here, we investigated avelumab as a potential therapy for chordoma. We examined 4 chordoma cell lines, first for expression of PD-L1, and in vitro for ADCC killing using NK cells and avelumab. PD-L1 expression was markedly upregulated by IFN-γ in all 4 chordoma cell lines, which significantly increased sensitivity to ADCC. Brachyury is a transcription factor that is uniformly expressed in chordoma. Clinical trials are ongoing in which chordoma patients are treated with brachyury-specific vaccines. Co-incubating chordoma cells with brachyury-specific CD8+ T cells resulted in significant upregulation of PD-L1 on the tumor cells, mediated by the CD8+ T cells' IFN-γ production, and increased sensitivity of chordoma cells to avelumab-mediated ADCC. Residential cancer stem cell subpopulations of chordoma cells were also killed by avelumab-mediated ADCC to the same degree as non-cancer stem cell populations. These findings suggest that as a monotherapy for chordoma, avelumab may enable endogenous NK cells, while in combination with T-cell immunotherapy, such as a vaccine, avelumab may enhance NK-cell killing of chordoma cells via ADCC. PMID:27172898