• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.123-130
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• 1997

Normal maturation of the mammalian oocytes is prerequisite for the fertilization and the early embryonic development. We have been tested the effects of purine and its de novo synthetic inhibitor, azaserine(Aza) on the maturation of germinal vesicle(GV) and germinal vesicle breakdown(GVBD) mouse oocytes. Denude-immature oocytes were cultivated in the media containing adenosine, guanosine, and/or azaserine, and checked the matruation stage by monitoring the prominent morphological changes. In GV stage oocytes, GV was arrested temporarily by the adenosine(1.0%) and protractedly by the guanosine(65.9%, P<0.001). The regression was increased significantly at the adenosine(90%, P<0.001) but decreased at the guanosine(1.6%, P<0.05). Inhibiting the de novo synthesis of purine, nuclear maturation rate was increase(90.4% : 96.7%), but GV arrest was significantly increased by cotreatment with guanosine(P<0.001). Polar body extraction significantly was increased at the Aza(P<0.05), but not in others. In GVBD oocytes, adenosine itself did not affect GVBD arrest. Guanosine, on the other hand, elevated GVBD arrest rate(P<0.001), but co-treated with Aza, decreased GVBD arrest(P<0.001). Aza increased GVBD arrest rate(20.2%, P<0.05) compared with control. From those results, we know that guanosine shows more prominent effect on the inhibition of nuclear maturation at the GV stage, and of the 1st polar body extrusion at the GVBD stage. Adenosine showed the cytoplasmic toxicity at GV stage oocyte. Our data speculate that cytoplasmic cAMP level is auto-regulated by endogenous adenylate cyclase while GVBD is inhibited by guanosine, since purine toxicity is not observed in the GVBD stage. And it is showed that purine metabolism is concerned with nuclear maturation, that the amounts of purine metabolism is not even during the oocyte maturation.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.111-116
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• 1997

We determined the effects of follicular fluid fractions in the maturation medium on bovine oocyte maturation, fertilization and subsequent development, as well as on number of cells in blastocysts following culture. Follicular fluid and oocytes from bovine follicles less than 5 mm in diameter were collected from the ovaries of slaughtered cows. Follicular fluid was separated into different molecular weight fractions by untrafiltration through a membrane using a centrifuge at 500$\times$g, for 2h. For the maturation medium, follicular fluid fractions (30%, v/v), whole fluid (30%) or PVP(3mg/ml) were added to TCM 199(0.1$\mu\textrm{g}$/ml estradiol-17$\beta$, 100IU hCG). After maturation for 24h, oocytes were fertilized in vitro with bull frozen-thawed spermatozoa and cultured on a monolayer of granulosa cells for 9 days after fertilization. There were no differences in maturation rates or fertilization rates among any maturation conditions. The rates of development to >2-cell stage of the oocytes were significantly decreased when fraction of follicular fluid below 10,000 MW were added into maturation medium, compared with control and fraction above 10,000 MW(26.0% vs 40.8% to 64.0%, respectveily. p<0.01). Likewise, the rates of development to blastocysts of fertilized oocytes were significantly decreased in maturation medium containing fraction of follicular fluid (<10,000 MW). The average cell number of blastocysts derived from oocytes that matured in the fraction(>10,000 MW) of follicular fluid was 154.7$\pm$13.7. These embryos contained more cells than those matured in whole follicular fluid, or the fraction(<10, 000 MW) of follicular fluid or control(107.0$\pm$8.4, 91.8$\pm$11.8 and 95.8$\pm$6.2, respectively). In conclusion, we found that fractions of follicular fluid contained factors stimulating or inhibiting oocyte cytoplasmic matruation. These suggest that a factor(s) inducing cytoplasmic maturation of oocytes may exist in >10,000 MW fraction of follicular fluid.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.5
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• pp.1243-1249
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• 2007

The aim of this study was to investigate the effect of ethanol extract of Fagopyrum esculentum on the melanogenesis. To determine whether ethanol extract of Fagopyrum esculentum suppress melanin synthesis in cellular level, B16F10 melanoma cells were cultured in the presence of different concentrations of Fagopyrum esculentum ethanol extract. In the present study, we examined the effects of Fagopyrum esculentum ethanol extract on cell proliferation, melanin contents, tyrosinase activity, expression of melanogenic enzyme proteins including tyrosinase, tyrosinase-related protein 1 (TRP-1) and tyrosinase-related protein 2 (TRP-2). Cell proliferation was slightly increased by treatment with ethanol extract of Fagopyrum esculentum $(25-200 {\mu}g/m{\ell}).$ The ethanol extract of Fagopyrum esculentum effectively suppressed melanin contents at a dose of $100 {\mu}g/m{\ell}).$ It was observed that the color of cell pellets was totally whitened compared with the control. The ethanol extract of Fagopyrum esculentum inhibited tyrosinase activity, regulate melanin biosynthesis as the key enzyme in melanogenesis. Using western blot analysis, the ethanol extract of Fagopyrum esculentum dose-dependently decreased tyrosinase and TRP-1 protein levels, and tyrosinase and TRP-1 were detected in similar manner. ${\alpha}-MSH$ leads to a stimulation of melanin synthesis through increase of tyrosinase activity, melanin contents and cytoplasmic dendricity. In this study, ethanol extract of Fagopyrum esculentum down-regulated the ${\alpha}-MSH$-induced tyrosinase activity, melanin contents and cytoplasmic dendricity. Regarding protein levels of the melanogenic enzymes, the amounts of tyrosinase and TRP-1 was increased after incubation with a-MSH. The treatment of ethanol extract of Fagopyrum esculentum decreased the ${\alpha}-MSH$-induced expression levels of tyrosinase and TRP-1. These results suggest that the ethanol extract of Fagopyrum esculentum exerts its depigmenting effects through the suppression of tyrosinase, TRP-1 and cytoplasmic dendricity. And it may be a potent depigmetation agent in hyperpigmentation condition.

• Applied Microscopy
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• v.24 no.4
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• pp.78-85
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• 1994

Sodium arsenite ($NaAsO_2$) was injected to the rat subcutaneously for the study of the acute toxicity of arsenite on hepatocytes, and the effects of pretreatment of arsenite and glutathione on the lethalty of the arsenite treated rats. Arsenite treated rat hepatocytes showed vacuolated cytosol and shrinked nuclear and expanded perinuclear space and cytoplasmic membrane whirl. Rats pretreated with BSO (L-Buthionine-SR-Sulfoximine), less survived than arsenite treated alone. It means that glutathione acts as a protecting agent against the arsenite. Subcutaneous sublethal dose (10mg/kg body weight) treatment was showed the protecting activity to lethality of lethal dose (15mg/kg body weight) treated rat. 10mg/kg body weight sublethal dose effects appeared in six hours intervals of between treatments.

• Journal of Microbiology and Biotechnology
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• v.17 no.11
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• pp.1797-1804
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• 2007

Lycopene, an acyclic carotenoid found in tomatoes (Lycopersicon esculentum) and a number off fruits, has shown various biological properties, but its antifungal effects remain poorly understood. The current study investigated the antifungal activity of lycopene and its mode of action. Lycopene showed potent antifungal effects toward pathogenic fungi, tested in an energy-independent manner, with low hemolytic effects against human erythrocytes. To confirm the antifungal effects of lycopene, its effects on the dimorphism of Candida albicans induced by fetal bovine serum (FBS), which plays a key role in the pathogenesis of a host invasion, were investigated. The results showed that lycopene exerted potent antifungal activity on the serum-induced mycelia of C. albicans. To understand the antifungal mode of action of lycopene, the action of lycopene against fungal cell membranes was examined by FACScan analysis and glucose and trehalose-release test. The results indicated that lycopene caused significant membrane damage and inhibited the normal budding process, resulting from the destruction of membrane integrity. The present study indicates that lycopene has considerable antifungal activity, deserving further investigation for clinical applications.

• Applied Microscopy
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• v.27 no.3
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• pp.225-234
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• 1997

Vincristine, a kind of anticancerous drugs, interferes with development of microtubles and synthesis of nucleic acid and proteins in cells, and destructs cytoplasmic membrane so that mitosis of cancer cells is inhibited. Unfortunately these anticancerous effects by vioneristime are not limited to specific cancer cells, so several side effects are produced. This study was performed to explore the effects of vincristine on the fine structure of cytoplasmic organelles and cartilagenous matrix in proximal epiphyseal plate of the tibia in rat. The results were as follows: 1. Cisternae of rough endoplasmic reticulum (RER) were fragmented and sacculated, and membrane-bound ribocomes of RER were detached at 3 and 6 hours after vincristine treatment. Severely dilated, fagmented and sacculated cisternae of RER were found at 12 hours after vincristine treatment, and at 24 hours after vincristine treatment a few cisternae were framented and sacculated. At 72 hours after vincristine treatment cisternae of RER were parallely well arraged. 2. Golgi complex was atrophied at 3, 6, and 12 hours after vincristine treatment, while at 72 hours after vincristine treatment the cisternae of Golgi complex were made of 5-6 layers. 3. Mitochondria with disorganized mitochondrial cristae and outer membrane-losed mitochondria were found at 3 hours after vincristine treatment. At 6 and 12 hours after vincristine treatment mitochondria had possessed disorganized cristae, and a few mitochondria with disorganized cristae were. observed at 24 hours after vincristine treatment. While at 72 hours after vincristine treatment mitochondria were shown distinct cristae and double membranes. 4. Phagosome were begun to observe at 3 hourse after vincristine treatment, and at 24 hourse after vincristine treatment many phagosomes were found, while at 72 hours after vincristine treatment a few phagosomes were observed. 5. In the cartilagenous matrix large-sized matrix granules were decreased and collagen fibrils were dispersed at 3, 6, and 12 hours after vincristine treatment, while at 72 hours after vincristine treatment many large-sized matrix granules and numerous matrix it is suggested that although vincristine may induce the degenerative changes of the chondrocyte, resulting in changes of components of the cartilagenous matirx, these toxic effects may be regressed with time.

• Toxicological Research
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• v.11 no.2
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• pp.199-203
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• 1995

The focus of this study was to investigate that cellular parameters and glucose uptake might be altered by extracellular calcium and starvation. Addition of 1 mM $Ca^{++}$ to hepatocytes (equalling to the free calcium concentration of blood) significantly increased intracellular $Na^+$ and decreased $Na^+$ & LDH leakage. This pertains to the hepatocytes of control rats as well as those of rats fasted for 24 and 48. hr. These effects might be come from the membrane-stabilizing effects of calcium. But calcium had no effects on cell volumes, superoxide-formation and glucose uptake. Actually hepatocytes of starved rats showed changes in several cellular parameters. Starvation increased LDH leakage, glucose uptake and the total concentration of $Na^+$ and $Na^+$ whereas it markedly decreased cell volumes. Since total tonicity remained unchanged, intracellular $Na^+$ and $Na^+$ could contribute to a higher share of total osmolarity in starvation. Starvation increased the cytoplasmic pH because $R-NH^{3+}$ions and their corresponding counterions disappeared. This increase may be related to suppress the protonization of amino groups in proteins. Starvation decreased hepatic glycogen, a major compound that affects cytosolic volume of hepatocytes. The data indicate that starvation increases the glucose transport activity. The possible molecular basis will be discussed.

• Biomedical Science Letters
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• v.8 no.1
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• pp.13-20
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• 2002

Asadisulphide were purified from Ferrula assafoetida by organic solvent extraction and chromatography. Since ethyl acetate extracts of F. assafoetida has the strongest inhibitory effects on adherence of HL-60 cells, it was reextracted with ethyl acetate, hexane, and ethyl ether and chromatographed three times to isolate asadisulphide. HL-60 cells were grouped into untreated control, TPA-treated, asadisulphide-teated and TPA＋asadisulphide-treated groups, and structural changes of these cells were observed using light microscope, scanning electron microscope and transmission electron microscope to examine the inhibitory effects of asadisulfide on the TPA-induced adherence of HL-60 cells. Light microscopic observations showed that asadisulphide has inhibitory effects on the cell aggregation, extention of cytoplasmic processes and inhibition of substrate adhesion of HL-60 cells. Using scanning and transmission electron microscope, it was observed that cell surfaces and several ultrastructures of TPA-treated HL-60 cell were different from control group, while there were no remarkable differences between asadisulphide-treated and TPA＋asadisulphide-treated group. These results could suggest that asadisulphide has the inhibitory effects on the TPA-induced structural changes of HL-60 cells.

• Journal of Korean Academy of Oral and Maxillofacial Radiology
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• v.25 no.1
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• pp.71-87
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• 1995

The purpose of this study was to investigate the early irradiation changes on the ultrastructure of the capillary endothelial cell in the rat submandibular glands. For the study, 110 Sprague-Dawley strain male rats were singly irradiated to their neck regions with the doses of 2Gy, 5Gy, and 10Gy by 6MV X -irradiation, and sacrificed on the 3 hours, 6 hours, 12 hours, 1 day, 3 days, 7 days, and 14 days after irradiation. The authors observed the histologic and ultrastructural changes of the capillary endothelial cell using the light and electron microscopes. The results were as follows: I. In the light microscopic examination, the capillary dilation was observed on the 6 hours group and the capillary density was slightly increased on the 12 hours group after 2Gy and 5Gy irradiation. And luminal size and capillary density were decreased on the 3 days and the 7 days groups after irradiation, after then, they were recovered. But capillary density was still decreased on the 14 days group after 10Gy irradiation. 2. In the transmission electron microscopic examination, the mild proliferation of cytoplasmic process of the endothelial cell and reduction in luminal size were observed on the 3 hours group after irradiation. After then, endothelial swelling, marked proliferation of cytoplasmic process, thickened basal lamina, and numerous pinocytotic vesicles were observed after the 1 day group after irradiation. Thickened basal lamina and numerous pinocytotic vesicles were still observed until the 7 days group after irradiation. These changes were recovered to normal on the 14 days group after 2Gy and 5Gy irradiation, but not after 10Gy irradiation. 3. In the scanning electron microscopic examination, the dilation of conduits and constriction, and meandering were observed on the 1 day group after 10Gy irradiation. These changes were observed with increased coarseness of the surface of the vascular resin casting on the 3 days group after irradiation. 4. From the above results, endothelial swelling, proliferation of cytoplasmic process, and thickening of the basal lamina appeared before the 6 hours group after irradiation. And these changes may also induce the increase of the capillary number and luminal size, after then, capillary permeability was increased via the increase of the number of pinocytotic vesicles. The changes were observed earlier and more apparent with the increase of the irradiation doses under the dose of 10Gy irradiation.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.4
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• pp.327-335
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• 2009

The aim of this study was to determine whether losartan, an angiotensin II (Ang II) type 1 ($AT_1$) receptor could influence the CA release from the isolated perfused model of the rat adrenal medulla. Losartan (5${\sim}$50 ${\mu}$M) perfused into an adrenal vein for 90 min produced dose- and time-dependent inhibition of the CA secretory responses evoked by ACh (5.32 mM), high $K^+$ (56 mM, a direct membrane depolarizer), DMPP (100 ${\mu}$M) and McN-A-343 (100 ${\mu}$M). Losartan failed to affect basal CA output. Furthermore, in adrenal glands loaded with losartan (15 ${\mu}$M) for 90 min, the CA secretory responses evoked by Bay-K-8644 (10 ${\mu}$M, an activator of L-type $Ca^{2+}$ channels), cyclopiazonic acid (10 ${\mu}$M, an inhibitor of cytoplasmic $Ca^{2+}$ -ATPase), veratridine (100 ${\mu}$M, an activator of $Na^+$ channels), and Ang II (100 nM) were markedly inhibited. However, at high concentrations (150${\sim}$300 ${\mu}$M), losartan rather enhanced the CA secretion evoked by ACh. Collectively, these experimental results suggest that losartan at low concentrations inhibits the CA secretion evoked by cholinergic stimulation (both nicotininc and muscarinic receptors) as well as by membrane depolarization from the rat adrenal medulla, but at high concentration it rather inhibits ACh-evoked CA secretion. It seems that losartan has a dual action, acting as both agonist and antagonist to nicotinic receptors of the rat adrenal medulla, which might be dependent on the concentration. It is also thought that this inhibitory effect of losartan may be mediated by blocking the influx of both $Na^+$ and $Ca^{2+}$ into the rat adrenomedullary chromaffin cells as well as by inhibiting the $Ca^{2+}$ release from the cytoplasmic calcium store, which is thought to be relevant to the $AT_1$ receptor blockade, in addition to its enhancement of the CA release.