共查询到20条相似文献,搜索用时 15 毫秒
1.
Baby hamster kidney cells (BHK-21) were exposed to culture filtrates of 4 Fusarium moniliforme isolates containing varying
levels of fumonisin B1 (FMB1) and the effects upon RNA, DNA and protein synthesis were monitored. Cells were also grown on
medium amended with FMB1 only for comparison. After 24 h incubation FMB1 (100 μg/100 ml medium) reduced protein synthesis
by 4% and by 18% after 48 h. Culture filtrates containing the highest levels of FMB1 also caused the greatest inhibition in
protein synthesis after 24 h but after 48 h protein synthesis levels were the same as controls even though the FMB1 level
was 360 μg/100 ml. Only FMB1 reduced DNA synthesis, by 8% after 24 h but after 48 h DNA levels had increased by 40 % over
controls. The culture filtrates containing the highest levels of FMB1 (360 μg/100 ml) reduced DNA synthesis more than 50%
after 24 h and 48 h. Culture filtrates containing lesser amounts of FMB1 in some instances stimulated DNA synthesis and inhibited
it in others. There was also no correlation in the level of FMB1 with the inhibition of RNA synthesis by BHK cells. It appears
that metabolites other than fumonisin produced by F. moniliforme in culture can affect and both stimulate and inhibit RNA,
DNA and protein synthesis by BHK cells.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
2.
Thomas S. Vates Garry L. Brake S. K. Majumdar Gail L. Ferguson 《In vitro cellular & developmental biology. Plant》1986,22(6):305-310
Summary Cis-diamminedichloroplatinum II (cis-DDP), an antitumor drug and the inactive trans-isomer were studied to evaluate their
effects on cell multiplication, DNA synthesis, and surface morphology of the murine erythroleukemia cells (clone 6A11A). Short-term
treatment of cells (1h) with 5 and 10μg/ml of cis-DDP resulted in a significant inhibition of cell multiplication. Continuous
treatment with cis-DDP (up to 144 h) significantly arrested cell growth at 1,5, and 10μg/ml. The cells exposed to 10 μg/ml
trans-DDP exhibited a slight decrease in cell multiplication; however, the 25-μg/ml treatments showed a modest inhibition
of cell growth. Continuous treatment with cis-DDP resulted in a concentrationdependent decrease in DNA synthesis, although
low-dose treatment (0.05 and 0.1 μg/ml), with a few exceptions, had no relative inhibitory effect. Likewise, trans-DDP treatments
decreased tritiated thymidine incorporation; however, this inhibitory effect was not as drastic as with corresponding concentrations
of cis-DDP. Scanning electron microscope studies revealed the formation of many giant cells and blebs at all short-term treatment
concentrations of cis-DDP past the 48 h interval. Continuous treatment of cis-DDP at 1 μg/ml concentration produced giant
cells with minute holes, whereas the 5 and 10 μg/ml exposure resulted in the formation of blebs and large holes and reduction
of microvilli past the 48-h treatment period. At higher concentrations the continuous treatment of cis-DDP completely destroyed
the cells. The surface morphology of trans-isomer treated cells, in most instances, resembled the corresponding untreated
control cells. 相似文献
3.
In previous studies, tobacco protoplasts were transformed with the bacterial gene encoding neomycin phosphotransferase II (NPT II). Transformed calluses lost neomycin phosphotransferase II activity after several subcultures. Treatment of calluses with 5-azacytidine, a demethylating agent, restored enzyme activity, suggesting that methylation of npt II sequences might be responsible for loss of NPT II activity. Studies presented here were designed to test that hypothesis. Results indicated that the effect of 5-azacytidine could not be blocked by the DNA replication inhibitor, hydroxyurea, nor by the 5-azacytidine analogue, cytidine as would be expected with a DNA demethylation mechanism. The level of NPT II mRNA was not increased by 5-azacytidine. Treatment with cycloheximide, a protein synthesis inhibitor, had no effect on 5-azacytidine-increased NPT II activity. There was no increase of NPT II protein caused by 5-azacytidine, whereas 5-azacytidine increased activity of NPT II. In contrast, the auxin 2,4-D increased both the NPT II protein and activity. Assays for malate dehydrogenase demonstrated that the effect of 5-azacytidine and hydroxyurea on NPT II was not due to an overall effect on callus metabolism. In vitro studies involving standard bacterial NPT II enzyme and crude extracts from untreated and 5-azacytidine- or hydroxyurea-treated calluses showed that the activity of NPT II added to the untreated extracts was lower than the activity of NPT II added to the extracts from calluses treated with 5-azacytidine or hydroxyurea, indicating that there was an unknown factor (or factors) in callus extracts which affected the activity of NPT II and itself was affected by 5-azacytidine and hydroxyurea treatment. These results suggested that one effect of 5-azacytidine in increasing NPT II activity was posttranslational.Abbreviations ELISA
enzyme-linked immunosorbent assay
- NOS
nopalene synthase
-
nos
DNA segment encoding NOS
- NPT II
neomycin phosphotransferase
-
npt II
DNA segment encoding NPT II
- PAGE
polyacrylamide gel electrophoresis 相似文献
4.
Our previous study revealed that the exposure of the drug 5-Azacytidine and ethanol to the cultured cortical neurons of mice
causes demethylation of cytosine residues in the CpG island of the NMDA receptor NR2B gene (Marutha Ravindran and Ticku, Mol
Brain Res 121:19-27, 2004). In the present study, we further analyzed methylation in the CpG island with various concentration
frame and time frame of exposure of the cultured cortical neurons with 5-azacytidine to identify whether methylation in the
NR2B gene is site specific or region specific. Methylation was studied by digesting the genomic DNA with methylation sensitive
HpaII, MspI, AciI or HhaI enzyme following the exposure of cultured cortical neurons of mice with 5-azacytidine by performing PCR and Southern hybridization.
We observed demethylation of DNA at 1, 3 and 5 μM concentrations of 5-azacytidine in the regions (5982–6155), (6743–7466)
and at 3 and 5 μM concentrations of 5-azacytidine used in the region (6477–6763). Similarly in the time frame study with 5-azacytidine,
demethylation of DNA was observed at 24 h and 36 h of incubation with 5-azacytidine in the regions (5982–6155), (6743–7466)
and at 36 h of incubation with 5-azacytidine used in the region (6477–6763). Our experimental results demonstrate that the
methylation in the CpG islands of the NR2B gene may not be site specific or region specific in the cultured cortical neurons
of mice. 相似文献
5.
Natural killer (NK) cells are non-T, non-B cell lymphocytes that lyse a variety of tumor and virus-infected cells. In this
study, we demonstrated that phytohemagglutinin (PHA) rendered resistant autologous T cells extremely sensitive to natural-killer(NK)-cell-mediated
lysis. The sensitization was very rapid and concentration-dependent (0.01–1 μg/ml); 62% and 95% of autologous T cells were
lysed by interleukin-2-activated NK cells 5 min and 18 h respectively after treatment with PHA (1 μg/ml). The maximal decrease
in the level of MHC class I molecules observed on T cells was 22%. Induction of susceptibility to NK-mediated lysis was correlated
with the expression of activation markers on T cells treated for relatively long intervals (more than 18 h) with high concentrations
of PHA (more than 0.1 μg/ml). Sensitization of T cells required RNA and protein synthesis, although DNA synthesis was not
essential. We propose that this unique system is suitable for studying the mechanisms involved in recognition and killing
of target cells by NK cells.
Received: 11 February 1999 / Accepted: 28 July 1999 相似文献
6.
Sokolov AV Golenkina EA Kostevich VA Vasilyev VB Sud'ina GF 《Biochemistry. Biokhimii?a》2010,75(12):1464-1469
The interaction between ceruloplasmin (CP), the multicopper oxidase of human plasma, and 5-lipoxygenase (5-LO), the key enzyme
of leukotriene synthesis, is shown for the first time. By Western-blotting and mass spectrometry of tryptic fragments, it
is shown that 5-LO from protein extract of human leukocytes binds with immobilized CP. Dose-dependent influence of intact
CP on leukotrienes synthesis is found: CP reduced leukotrienes synthesis in leukocytes in a dose above 50 μg/ml (normal CP
concentration in plasma is about 300–400 μg/ml). Proteolyzed CP and apo-form of CP is unable to inhibit activity of 5-LO.
CP increased activity of 5-LO at low doses (5–10 μg/ml). On the whole, the influence of CP on phagocytosis index of leukocytes
coordinates with influence on activity of 5-LO: the index increased in the range of 2–10 μg/ml CP and decreased at doses of
CP above 40 μg/ml. The dual role of CP in regulation of cellular response of leukocytes is discussed. 相似文献
7.
Banfalvi G Ujvarosi K Trencsenyi G Somogyi C Nagy G Basnakian A 《Apoptosis : an international journal on programmed cell death》2007,12(7):1219-1228
Murine pre-B-cells grown in the presence of lower (1 μM) or higher (5 μM) concentration of cadmium chloride were separated
into 13 fractions by centrifugal elutriation. The rate of DNA synthesis after cadmium treatment determined in permeable cells
was dependent on cell culture density during cadmium treatment. Cell cycle analysis revealed a shift in the profile of DNA
synthesis from replicative to repair DNA synthesis upon cadmium treatment. The study of the relationship between cell culture
density and cell diameter at lower and higher cell densities in the presence of 1 μM cadmium chloride concentration showed
that a. at 5×105 cell/ml or lower densities cells were shrinking indicating apoptotic changes, b. at higher cell culture densities the average
cell size increased, c. the treatment of cells with low CdCl2 concentration (1 μM) at higher cell culture density (>5×105 cell/ml) did not change significantly the average cell diameter. At 5 μM cadmium concentration and higher cell culture densities
(>5×105 cell/ml) the average cell size decreased in each elutriated fraction. Most significant inhibition of cell growth took place
in early S phase (2.0–2.5 C value). Apoptotic chromatin changes in chromatin structure after cadmium treatment were seen as
large extensive disruptions, holes in the nuclear membrane and stickiness of incompletely folded chromosomes. 相似文献
8.
Purification and characterization of a DNA synthesis inhibitor protein from mouse embryo fibroblasts
A DNA synthesis inhibitor protein was purified from the conditioned medium of cycloheximide treated mouse embryo fibroblasts.
This protein has a molecular weight of 45,000 as determined by gel filtration and Polyacrylamide gel electrophoresis. The
levels of the [35S] methionine la belled 45 kDa protein in the medium and matrix were monitored across two cell cycles in synchronized cultures.
The 45 kDa protein was present in higher levels in the medium of non-S-phase cells depicting a peak between the two S-phases.
The DNA synthesis inhibitor protein was immunologically related to a chicken DNA-binding protein which showed similar cell
cycle specific variations at the intracellular level. The purified 45 kDa protein inhibited DNA synthesis in murine and human
cells. In mouse embryo fibroblasts, the DNA synthesis was inhibited to an extent of 86% by 0.25 μg/ml of the inhibitor, while
higher amounts of the inhibitor were required to arrest DNA synthesis in human skin fibroblasts: in these cells, 4 μg/ml of
the inhibitor inhibited DNA synthesis to an extent of 50%. The high levels of the 45 kDa protein in the medium of non-S phase
cells and its DNA synthesis inhibitory potential suggest that this protein may be involved in the regulation of DNA synthesis
during the cell cycle. 相似文献
9.
Robert A. Harper B. Allen Flaxman 《In vitro cellular & developmental biology. Plant》1981,17(5):393-396
Summary Primary cell cultures of normal rabbit epidermal cells (keratinocytes) were established without the use of enzymatic techniques.
Six experiments were carried out on cells from six different rabbits. When these cells were exposed to methotrexate (MTX)
for 24 h at 1 μg/ml, proliferation, as measured by cells entering mitosis, was significantly inhibited (P<0.05) in only one experiment. When the dose of MTX was elevated to 100 μg/ml, only two experiments showed significant inhibition
of mitosis. This minimal inhibition of mitosis by MTX was contrasted by the dramatic inhibitory effect of this antimetabolite
on DNA synthesis. At 1 μg/ml MTX for 24 h, DNA synthesis, as measured by [3H]deoxyuridine uptake, was inhibited >95%. We can conclude that under certain conditions, the rabbit keratinocyte may represent
a normal cell type that is inherently resistant to the antiproliferative effects of methotrexate.
The research was supported by National Cancer Institute Grant CA 11536. 相似文献
10.
Masayoshi Ono John W. Perry Takami Oka 《In vitro cellular & developmental biology. Plant》1981,17(2):121-128
Summary Cortisol was previously shown to elicit a concentration-dependent inhibition of α-lactalbumin accumulation in midpregnant
mouse mammary gland cultured in medium containing optimal concentrations of 5 μg/ml prolactin and insulin. In contrast, casein
accumulation under these conditions was progressively stimulated by addition of increasing amounts of cortisol (Ono, M.; Oka,
T. Cell 19: 473–480; 1980). In the present study we found that in the presence of a suboptimal concentration of 0.5 μg/ml
prolactin, 2.8×10−9
M to 2.8×10−7
M cortisol stimulated α-lactalbumin accumulation. Furthermore, higher concentrations of cortisol produced a smaller inhibition
of α-lactalbumin accumulation as compared to that obtained in cultures containing 5 μg/ml prolactin. The maximal increase
in α-lactalbumin accumulation attained in the presence of 1.4×10−8
M cortisol, 0.5 μg/ml prolactin, and insulin was comparable to that observed in culture containing 5 μg/ml prolactin and insulin.
Similar results were obtained in a cortisol concentration-response study of α-lactalbumin accumulation in cultures containing
a suboptimal concentration of 0.5 μg/ml human placental lactogen. Measurement of the rate of α-lactalbumin synthesis in cultured
tissue indicated that the opposing effects of low and high concentrations of cortisol on α-lactalbumin accumulation involved
an alteration in the rate of synthesis of the milk protein. In contrast to α-lactalbumin, the synthesis of casein was stimulated
in a concentration-dependent manner by addition of cortisol that acted synergistically with either 0.5 μg/ml or 5 μg/ml prolactin.
The maximal increases were obtained in the presence of 2.8×10−6
M cortisol. These results indicated that the action of cortisol on α-lactalbumin accumulation can be modulated by the concentration,
of prolactin and suggest that the interplay between cortisol and prolactin in regulation of α-lactalbumin synthesis may be
different from that involved in casein synthesis. 相似文献
11.
RNA synthesis required for DNA replication in Vicia seed embryos 总被引:1,自引:0,他引:1
Fukuei Keisuke; Sakamaki Tadashi; Takahashi Noribumi; Takaiwa Fumio; Tanifuji Shigeyuki 《Plant & cell physiology》1977,18(1):173-180
The synthesis of DNA and RNA during germination of Vicia seedswas examined. Incorporation of 3H-thymidine into DNA reacheda maximum at about 32 hr after the beginning of imbibition,and RNA synthesis was shown to precede DNA replication. Sedimentationanalyses of 3H-uridine-labeled RNAs indicated that the embryossynthesize all types of rRNA, heterodisperse RNA and 45SRNA before and also during the phase of DNA replication. Actinomycin-treatments at lower concentrations (50 or 100 µg/ml)resulted in the specific inhibition of rRNA synthesis. Suchinhibition did not lead to a large reduction in 3H-thymidineincorporation during the replication phase. However, DNA synthesiswas drastically inhibited by a higher level (200 µg/ml)of actinomycin D. The results strongly suggest the involvementof synthesis of heterodisperse RNA in DNA replication. (Received May 28, 1976; ) 相似文献
12.
The inhibitory activities of a novel antibiotic compound have been investigated. A synthetic version of the natural product
TAN-1057A was examined for its effects on translation and ribosomal subunit formation. The antibiotic at 6 μg/ml reduced the
growth rate of wild-type Staphylococcus aureus cells by 50%. The IC50 for inhibition of protein synthesis in these cells was 4.5 μg/ml. Pulse and chase labeling kinetics showed a strong inhibitory
effect on 50S ribosomal subunit formation as well. The IC50 for this process was 9 μg/ml, indicating an equivalent inhibitory effect of the antibiotic on translation and 50S synthesis.
The post-antibiotic effect of the drug was investigated. Protein synthesis resumed rapidly after removal of the drug from
cells, but full recovery of the normal 50S subunit complement in treated cells required 1.5 h. The dual inhibitory effects
of this compound are compared with other antimicrobial agents having similar effects on cell growth.
Received: 27 December 2000 / Accepted: 22 March 2001 相似文献
13.
Michiel B. Vandegehuchte Filip Lemière Lynn Vanhaecke Wim Vanden Berghe Colin R. Janssen 《Comparative biochemistry and physiology. Toxicology & pharmacology : CBP》2010,151(3):278-285
The purpose of this study is to investigate (1) the induction of epigenetic effects in the crustacean Daphnia magna using DNA methylation as an epigenetic mark and (2) the potential stable transfer of such an epigenetic effect to non-exposed subsequent generations. Daphnids were exposed to chemical substances known to affect DNA methylation in mammals: vinclozolin, 5-azacytidine, 2′-deoxy-5-azacytidine, genistein and biochanin A. Effects on overall DNA cytosine methylation, body length and reproduction were evaluated in 21 day experiments. Using a multi-generational experimental design these endpoints were also evaluated in the F1 and F2 generation of both exposed and non-exposed offspring from F0 daphnids exposed to 5-azacytidine, genistein or vinclozolin. A reduction in DNA methylation was consistently observed in daphnids exposed to vinclozolin and 5-azacytidine. Only in organisms exposed to 5-azacytidine was this effect transferred to the two subsequent non-exposed generations. A concurrent reduction in body length at day 7 was observed in these treatments. For the first time, exposure to environmental chemicals was shown to affect DNA methylation in the parental generation of D. magna. We also demonstrated a transgenerational alteration in an epigenetic system in D. magna, which indicates the possibility of transgenerational inheritance of environment-induced epigenetic changes in non-exposed subsequent generations. 相似文献
14.
Tonya D. Mitchell Ajmer S. Bhagsari Peggy Ozias-Akins Sarwan K. Dhir 《In vitro cellular & developmental biology. Plant》1998,34(4):319-324
Summary Transient expression of the β-glucuronidase (GUS) gene has been studied in leaf-derived embryogenic callus of sweetpotatoIpomoea batatas L. (Lam.) by electroporation. The influence of several factors including electric field strength, buffer composition, time
course of transientGUS gene expression, DNA concentration, enzyme, and polyethylene glycol (PEG) treatment was examined onGUS gene expression (number of blue spots). MaximumGUS gene expression (an average of 90 blue spots/fifty mg fresh weight callus tissue) was observed after 48 h when callus pieces
were preincubated with electroporation (EPR) buffer for 1 h, followed by electroporation with a single electric pulse of 500
V/cm discharged from a 960-μF capacitor in the presence of 20 μg DNA/ml and 8.3 μl NaCl (3M). Changing the electroporation buffer conductivity (by varying the buffer composition with low-high salt concentrations),
had only slight effect on the number of blue spots. Similarly, the time course study ofGUS gene expression revealed that GUS activity could be detected 12 h after electroporation with a maximum activity after 72
h (112 blue spots). Increasing the amount of DNA from 5 to 50 μg/ml in the EPR buffer had a slight effect on the expression
frequency (from 20–110 blue spots, and 112 blue spots with 20 μg/ml). The number of blue spots was increased by enzymatic
wounding of callus pieces for 10 min and by addition of 200 μl PEG 4000 (15%) before electroporation. These results suggest
that intact cell electroporation can be used for producing transgenic sweetpotato tissue. 相似文献
15.
Summary. The effect of α-difluoromethylornithine (DFMO) on the apoptosis of HC11 mouse mammary epithelial cells was investigated.
The involvement of reactive oxygen species (ROS) and Bcl-2 protein in the mechanism of apoptosis induced by ornithine decarboxylase
(ODC) inhibition was also assessed. DFMO (0.1, 1 and 5 mM) induced apoptosis of HC11 cells in dose- and time-dependent manner.
Apoptosis manifests itself with morphological features like: cell shrinkage, condensation of chromatin, pyknosis and fragmentation
of nucleus, followed by secondary necrosis (putrosis). The decrease in the nuclear DNA contents appearing as the hypodiploidal
peak sub-G1 in the DNA histogram was not dependent on the presence of prolactin (5 μg/ml) in DFMO-treated cultures. Apoptosis induced
by ODC inhibition was associated with a rapid increase in ROS concentration in HC11 cells observed within 1 h after DFMO treatment.
The down-regulation of Bcl-2 as a decrease in cell number expressing bcl-2 and a lowered Bcl-2 protein content in cells expressing this protooncogene was also noted. The administration of putrescine
(50 μM) lowered the number of early-apoptotic, late-apoptotic and necrotic cells. Moreover, it increased the number of cells
expressing bcl-2. In conclusion, the disturbance of cellular polyamine homeostasis by inhibition of their synthesis enhances mammary epithelial
cell susceptibility to apoptosis. It may occur in the mammary gland at the end of lactation, when the depletion of circulating
lactogenic hormones and activation of intra-mammary apoptogenic factors expression take place.
Received May 6, 1999; Accepted December 15, 1999 相似文献
16.
Modulation of the actin cytoskeleton in chondrocytes has been used to prevent or reverse dedifferentiation and to enhance
protein synthesis. We have hypothesized that an actin-modifying agent, staurosporine, could be used with fibrochondrocytes
to increase the gene expression and synthesis of critical fibrocartilage proteins. A range of concentrations (0.1–100 nM)
was applied to fibrochondrocytes in monolayer and evaluated after 24 h and after 4 days. High-dose staurosporine treatment
(10–100 nM) increased cartilage oligomeric matrix protein 60– to 500-fold and aggrecan gene expression two-fold. This effective
range of staurosporine was then applied to scaffoldless tissue-engineered fibrochondrocyte constructs for 4 weeks. Whereas
glycosaminoglycan synthesis was not affected, collagen content doubled, from 27.6 ± 8.8 μg in the untreated constructs to
55.2 ± 12.2 μg per construct with 100 nM treatment. When analyzed for specific collagens, the 10-nM group showed a significant
increase in collagen type I content, whereas collagen type II was unaffected. A concomitant dose-dependent reduction was noted
in construct contraction, reflecting the actin-disrupting action of staurosporine. Thus, staurosporine increases the gene
expression for important matrix proteins and can be used to enhance matrix production and reduce contraction in tissue-engineered
fibrocartilage constructs.
The authors gratefully acknowledge NIAMS R01 AR 47839–2 for funding this work, and the Hertz Foundation for their support
of G. Hoben. 相似文献
17.
Sung Gu Lee Hye Yeon Koh Hyuncheol Oh Se Jong Han Il-Chan Kim Hong Kum Lee Joung Han Yim 《Biotechnology letters》2010,32(4):471-475
Type I collagen is the major structural protein in dermis and its presence is used to monitor skin cell proliferation and
aging. Recently, novel usimine compounds have been found in the Antarctic lichen Ramalina terebrata. In the present study, usimine-C induced cell proliferation of human dermal fibroblast, CCD-986SK, up to 1.6-fold after treating
with 90 μg/ml for 48 h. Type I procollagen synthesis was significantly increased 1.3-fold, 3-fold, and 5-fold after treating
with 0.14, 0.72, and 3.6 μg usimine-C/ml for 24 h, respectively, whereas no significant increase in type I procollagen was
observed after treating with usimine-A or -B. Usimines are usnic acid derivatives. Considering that the difference among the
derivatives is a side chain, the proliferation activity may be related to this side chain, triggering an internal signal for
type I procollagen expression. Further studies still remain to clarify the signaling pathways for the type I procollagen induction,
which is activated by usimine-C. 相似文献
18.
DNA methylation levels in porcine fetal fibroblasts induced by an inhibitor of methylation, 5-azacytidine 总被引:1,自引:0,他引:1
Mohana Kumar B Jin HF Kim JG Song HJ Hong Y Balasubramanian S Choe SY Rho GJ 《Cell and tissue research》2006,325(3):445-454
Removal of the somatic DNA methylation pattern from donor cells and remodeling of embryonic status have been suggested as integral processes for successful nuclear transfer (NT) reprogramming. This study has investigated the effects of 5-azacytidine (5-azaC), a DNA methylation inhibitor, on global methylation changes in porcine fetal fibroblasts (PFF); this may improve NT attributable to the potential reprogramming of the methyl groups. PFF in 5th passage cultures were treated with 0, 0.5, 1.0, 2.0, and 3.0 μM 5-azaC for 96 h; 5-azaC inhibited the growth at all tested concentrations. At the higher concentrations of 5-azaC used, cells appeared to exhibit morphological changes and to become apoptotic as observed by TUNEL assay. Thus, cells were negatively affected by 5-azaC. Differences in cellular ploidy were also observed at higher concentrations. Analysis showed no considerable changes in the proportion of cells at the G1-phase of the cell cycle with 5-azaC concentrations. The fractional part of the methylated DNA of these cells was significantly reduced by 5-azaC treatment. Confocal microscopy confirmed the inhibition of methylation levels in PFF with increased concentrations of 5-azaC. Exposure to 5-azaC altered the expression of genes involved in imprinting (IGF2) or pro-apoptosis (BAX), whereas there was a reduction in the expression of the main enzyme responsible for replicating the DNA methylation pattern (DNMT1) and anti-apoptosis (BCL2L1). Therefore, 5-azaC induces a relative reduction in methylation in PFF, and cells treated with 0.5 μM 5-azaC may have enhanced potential for porcine NT.The financial support of BioGreen 21 (grant no. 100052004002000) and KOSEF (grant no. R05-2004-000-10702-0) in Korea is gratefully acknowledged. 相似文献
19.
Hans Hirsiger Urs Giger Urs A. Meyer 《In vitro cellular & developmental biology. Plant》1984,20(3):172-182
Summary Monolayer cultures were prepared from hepatocytes of 15 d chick embryos and maintained at high cell density in a chemically
defined medium. In the absence of growth stimulatory conditions DNA synthesis was observed only during the first 10 to 16
h of culture. Thus, after a 12 h exposure to [3H]thymidine ([3H]dThd, 4 to 16 h) 9.1±1% (
,n=4) of the hepatocyte nuclei were labeled. Labeled mitotic nuclei, up to late telophase, were regularly observed in these
cultures. Beyond 16 h less than 2% labeled nuclei were found (12 h of [3H]dThd), which indicates that the hepatocytes entered proliferative quiescence. DNA synthesis of “resting” hepatocytes was
stimulated by insulin and, only slightly, by hydrocortisone, glucagon, or fetal bovine serum. Triiodothyronine (T3), or the nucleoside inosine (i) did not stimulate. Combination of insulin (I) with hydrocortisone (H), T3 (T), or glucagon (G) resulted in a more than additive effect. Nearly maximal stimulation occurred with the combinations IHT
and ITG. Labeling increased at 10 ng/ml of each component and was maximal at 1 to 10 μg/ml. A lag period of 8 to 10 h after
hormone administration (IHiTG, 10 μg/ml) was observed before nuclear labeling increased. Within the subsequent 10 h a considerable
proportion of the hepatocytes (up to 30% or more) entered DNA synthesis. Mitotic activity (with nuclei in prophase up to late
telophase) also was stimulated. An increase of both total DNA and protein content was measured in several experiments. Hormonal
stimulation of hepatocyte DNA synthesis and mitotic activity was associated with decreased β-naphthoflavone-mediated induction
of cytochrome P450. A causal relationship between these two phenomena remains to be established. It is suggested that chick
embryo hepatocyte cultures are a useful tool for studies on hepatocyte proliferation and differentiation.
The present study is based on original observations by Dr. F. R. Althaus (presently at the Institute of Pharmacology and Biochemistry,
University of Zürich, Switzerland). This contribution of his and his incisive criticism are acknowledged.
The study was supported by Grant 3.893.81 from the Swiss National Research Foundation. 相似文献
20.
Mycelial yield and production of three trichothecenes, namely T-2 toxin, diacetoxyscirpenol (DAS) and neosolaniol (NEO) were
compared in control (CS) and carbendazim-resistant strains (RS) ofFusarium sporotrichioides. Each strain was exposed to graded concentrations of carbendazim (0, 1, 2, and 4 μg/ml media) for 2, 5 and 7 days under shake-culture
conditions at an incubation temperature of 25°C. Mycelial yield was significantly (P<0.001) affected by strain, carbendazim
concentration and incubation time. The strain differences in mycelial mass at 2 days (P<0.05) became more pronounced at 5
and 7 days of incubation (P<0.001). However, mycelial growth differences between the two strains were greatest following exposure
to carbendazim, with the effects becoming more divergent with time. Combined results for the three incubation times showed
dose related effects in carbendazim inhibition of T-2 toxin production by CS isolates. In contrast, RS cultures exposed to
the 2 μg/ml addition of carbendazim significantly increased T-2 toxin production (P<0.05 or better). At 1 and 4 μg/ml additions,
T-2 toxin inhibition occurred but the effect was less marked than in the CS series. RS yielded more DAS than CS at 5 days
(P<0.05) and at 7 days (P<0.01) of incubation. The major component of this strain difference arose from the effects of the
2 μg/ml addition of carbendazim (P<0.01). NEO production was also higher in RS than in CS, with the difference becoming progressively
more pronounced from day 5 (P<0.05) to day 7 (P<0.01) of incubation. However, these differences reflected enhanced NEO output
with carbendazim addition of 4 μg/ml (P<0.05) in day 5 extracts and of both 2 μg/ml (P<0.01) and 4 μg/ml additions (P<0.05)
in day 7 samples. Moreover, the ratio of NEO to T-2 toxin production was affected by an interaction involving incubation time,
strain and carbendazim dose (P<0.05 or better). On day 5, this ratio was greater in CS exposed to 2 μg/ml, but at 4 μg/ml,
the ratio was higher in RS. It is concluded that carbendazim resistance induced genuine differences in the synthesis of T-2
toxin and NEO. It is suggested that the strain difference may reside in the conversion of NEO to T-2 toxin which may be sensitive
to fungicide concentration. This would imply that carbendazim resistance induces changes in the terminal rather than initial
phases of trichothecene biosynthesis. 相似文献