• Environmental Analysis Health and Toxicology
• /
• v.18 no.2
• /
• pp.89-94
• /
• 2003

Tributyltin (TBT) used world-wide in antifouling paints for ships is a widespread environmental pollutant and cause reproductive organs atrophy in rodents. At low doses, antiproliferative modes of action have been shown to be involved, whereas at higher doses apoptosis seems to be the mechanism of toxicity in reproductive organs by TBT. In this study, we investigated that the mechanisms underlying DNA fragmentation induced by TBT in the rat leyding cell line, R2C. Effects of TBT on intracellular Ca$\^$2+/ level and reactive oxygen species (ROS) were investigated in R2C cells by fluorescence detector. TBT significantly induced intracellular Ca$\^$2+/ level in a time-dependent manner. The rise in intracellular Ca$\^$2+/ level was followed by a time-dependent generation of reactive oxygen species (ROS) at the cytosol level. Simultaneously, TBT induced the release of cytochrome c from the mitochondrial membrane into the cytosol. Furthermore, ROS production and the release of cytochrome c were reduced by BAPTA, an intracellular Ca$\^$2+/ chelator, indicating the important role of Ca$\^$2+/ in R2C during these early intracellular events. In addition, Z-DEVD FMK, a caspase-3 inhibitor, decreased apoptosis by TBT. Taken together, the present results indicated that the apoptotic pathway by TBT might start with an increase in intracellular Ca$\^$2+/ level, continues with release of ROS and cytochrome c from mitochondria, activation of caspases,and finally results in DNA fragmentation.

• The Korean Journal of Pharmacology
• /
• v.31 no.1 s.57
• /
• pp.63-74
• /
• 1995

It has been known that, during hypoxia, the adrenal medulla is activated to release catecholamines (CA) while hypoxia also inhibits high $K^+$ -induced CA secretion in the cultured bovine adrenal chromaffin cells. The present study was attempted to examine the effect of hypoxia on CA secretion evoked by chlinergic stimulation and membrane-depolarization from the isolated perfused rat adrenal glands and also to clarify its mechanism of action. For this purpose, using the isolated rat adrenal glands, the effects of hypoxia on CA release evoked by nicotinic ($N_1$) and muscarinic ($M_1$) receptor agonists, membrane-depolarizing agent, $Ca^{++}$-channel activator, intracellular $Ca^{++}$-releaser and ACh were determined. Experiments were carried out, perfusing Krebs solution pre-equilibrated with a gas mixture of 95% N_2$ and 5% $CO_2$. Hypoxia was maintained for $3{\sim}4$ hours through the experiments. Hypoxia gradually caused a time-dependent seduction in CA secretion evoked by DMPP ($100{\mu}M$), McN-A-343 ($100{\mu}M$), ACh (5.32 mM), Bay-K-8644 ($10{\mu}M$) and high $K^+$ (56 mM) respectively. How-ever, it did not affect CA secretion evoked by cyclopiazonic acid ($10{\mu}M$). Hypoxia itself also did fail to produce any influence on spontaneous secretory response of CA. These experimental results suggest that hypoxia depresses CA release evoked by both cholinergic stimulation and membrane-depolarization from the isolated rat adrenal medulla, and that this inhibitory activity may be due to the result of the direct inhibition of $Ca^{++}$ influx into the chromaffin cells without any effect on the calcium mobilization from the intracellular store.

• The Korean Journal of Pharmacology
• /
• v.29 no.2
• /
• pp.225-235
• /
• 1993

Pituitary LH release has been known to be regulated by the hypothalamic gonadotropin releasing hormone (GnRH) and the gonadal steroid hormones. In addition, neurotransmitters and neuropeptides are actively involved in the control of LH secretion. The alteration in LH release might reflect changes in biosynthesis and/or posttranslational processing of LH. However, little is known about the mechanism by which biosynthesis of LH subunits is regulated, especially at the level of transcription. In order to investigate if ovarian steroid hormones regulate the LH subunit gene expression, ${\alpha}\;and\;LH{\beta}$ steady state mRNA levels were determined in anterior pituitaries of ovariectomized rats. Serum LH concentrations and pituitary LH concentrations were increased markedly with time after ovariectomy. ${\alpha}\;and\;LH{\beta}$ subunit mRNA levels after ovariectomy were increased in a parallel manner with serum LH concentrations and pituitary LH contents, the rise in $LH{\beta}$ subunit mRNA levels being more prominent than the rise in ${\alpha}\;subunit$ mRNA. ${\alpha}\;and\;LH{\beta}$ subunit mRNA levels in ovariectomized rats were negatively regulated by the continuous treatment of ovarian steriod hormones for $1{\sim}4\;days$ and $LH{\beta}\;subunit$ mRNA seemed to be more sensitive to negative feedback of estradiol than progesterone. Treatment of estrogen antagonist, LY117018 or progesterone antagonist, RU486 significantly restroed LH subunit mRNA levels as well as LH release which were suppressed by estradiol or progesterone treatment. These results suggest that ovarian steroids negatively regulate the LH synthesis at the pretranslational level by modulating the steady state levels of ${\alpha}\;and\;LH{\beta}\;subunit$ mRNA and $LH{\beta}\;subunit$ mRNA seemed to be more sensitive to negative feedback action of estradiol than progesterone.

• Tuberculosis and Respiratory Diseases
• /
• v.42 no.5
• /
• pp.703-712
• /
• 1995

Background: Peripheral blood monocytes are important immune effector cells that play a fundamental role in cellular immunity. In addition to their antigen-presenting and phagocytic activities, monocytes/macrophage produce a vast array of regulatory and chemotactic cytokines. Interleukin-8(IL-8), a potent neutrophil-activating and chemotactic peptide, is produced in large quantities by mononuclear phagocytes and may be an important mediator of local and systemic inflammation. Overexpression by IL-8 of such inflammation may be an important step of tissue injury frequently seen in inflammatory reaction. So it could be hypothesized that the agents which block the production of IL-8 can decrease the inflammatory reaction and tissue injury. To evaluate this, we described the effect of Dexamethasone, $PGE_2$, Indomethacin and Interferon-$\gamma$(IFN-$\gamma$) on IL-8 mRNA and protein expression from LPS-stimulated human peripheral blood monocytes(PBMC). Method: PBMC was isolated from healthy volunteers. To evaluate the effect of Dexamethasone, $PGE_2$ & Indomethacin, these drug were treated for 1 hour before and after LPS stimulation and IFN-$\gamma$ was only treated I hour before the LPS stimulation. Northern blot analysis for IL-8 mRNA and ELISA for immunoreactive IL-8 protein in culture supernatant were performed. We repeated above experiment three times for Northern blot analysis and two times for ELISA and got the same result. Results: 1) Pre- and post-treatment of Dexamethasone suppressed both the LPS stimulated IL-8 mRNA expression and IL-8 protein release in PBMC. 2) IFN-$\gamma$ pre-treatment suppressed the IL-8 mRNA expression and IL-8 protein release in unstimulated cells. 3) In LPS stimulated cells, IFN-$\gamma$ suppressed the IL-8 mRNA expression but IL-8 protein release suppression was not observed. 4) $PGE_2$ and Indomethacin exert no effect on the LPS-stimulated IL-8 mRNA and protein expression in concentration used in this experiment ($PGE_2;10^{-6}M$, Indomethacin; $10{\mu}M$). Conclusion: One of the mechanism of antiinflammatory action of Dexamethasone can be explained by the suppressing effect of IL-8 production in some extent and by this antiinflammatory effect, dexamethasone can be used to suppress local and systemic inflammation mediated by IL-8.

• KOREAN JOURNAL OF CROP SCIENCE
• /
• v.25 no.2
• /
• pp.6-14
• /
• 1980

One of the physiological disease, sudden wiliting of Yushin variety suggested that low sunlight, excessive nitrogen application, and highly reduced soil condions either singly or combined, might be possible causes of the disorder. Some visual symptom of sudden wilting are discoloration of leaves, development of nodal roots above the soil surface, total root rot, and lodging. Those observations led to the hypothesis that suffocation of root tissues was a direct cause of the wilting. The oxygen transport characteristics of Yushin, IR262 and Tongil were examined by two methods. First, Soil-cultured plants of the three varieties were subjected to paraffin treatment to decrease the oxygen supply from the air to root tissues through the soil-water system, liquid paraffin was applied to the water surface in the pots at panicle formation stage. In this experiment, sudden wilting was observed of Yushin and IR262 at about a week after the treatment, but Tongil remained green and healthy. Wilting-resistant variety Tongil had higher oxygen release, whereas the susceptible Yushin and IR262 had lower oxygen release. Second, the number and size of the air spaces in each internode were investigated in the 5th internode from the top, all three varieties have a similar number of air spaces, although the air spaces of Thongil were larger. In the 4th internode, Tongil had plenty air spaces, Yushin and one of the Yushin's parents IR262 had scanty or none. The observations indicated that the ability of Yushin and IR262 for oxygen transport is very limited compared with that of Tongil. The limited oxygen supply due to poor development of air space in internode of rice plants may cause suffocation of root tissues, weaken metabolic activity of the tissues, and induce root rot, subsequently inducing sudden wilting and lodging under unfavorable weather, soil and cultural conditions.

• Tuberculosis and Respiratory Diseases
• /
• v.40 no.3
• /
• pp.213-222
• /
• 1993

Background : Bronchial asthma has been known as an inflmmatory disease. There have been many evidences that polymorphonuclear leukocytes (PMN) might play an important role in the pathogrnesis of asthma. Although many investigators suggested that pneumocyte injury by PMN-derived oxygen radicals may contribute to the pathogenesis of asthma, there has been few report for a direct evidence of oxygen radicals-mediated pneumocyte injury in bronchial asthma. Furthermore the exact mechanism of oxygen radicals-mediated pneumocyte injury is still controversy. This study was designed to establish a direct in vitro evidence and its clinical significance of pneumocyte injury by PMN-derived superoxide anion in bronchial asthma and to elucidate the main mechanism of superoxide anion-mediated pneumocyte injury. Methods : 12 stable asthmatics and 5 healthy volunteers were participated in this study. PMN was separated from peripheral venous blood samples by using dextran sedimentation and Ficoll-Hypaque density gradient separation method. Superoxide anion productions by PMN and plasma SOD activities were measured by spectrophotometric assay using the principle of SOD inhibitable cytochrome c reduction. PMN-mediated pneumocyte injuries were measured by $^{51}Cr$-release assay using A549 pneumocytes and were expressed as percent lysis and percent detachment. Results: 1) PMN from asthmatics produced more amount of superoxide anion compared to PMN from normal subjects ($6.65{\pm}0.58$ vs $2.81{\pm}0.95\;nmol/1{\times}10^6$ cells, p<0.05), and showed an inverse correlation with $FEV_1$(R=-0.63, p<0.05), but no correlation with $PC_{20}$ histamine in asthmatics. 2) Plasma SOD activities were decreased in asthmatics compared to normal subjects but not significant, and showed a positive correlation with $FEV_1$(R=0.63, p<0.05) but no correlation with $PC_{20}$ histamine in asthmatics. 3) There were a positive correlation between plasma SOD activity and superoxide anion production by PMN in normal subjects (R=0.88, p<0.05) but not in asthmatics. 4) PMN-mediated pneumocyte injury was predominantly expressed as cell detachment rather than cell lysis in both groups, and PMN from asthmatics showed more potent cytotoxic effect on A549 pneumocytes compated to PMN from normal subjects. PMN-mediated detachment rather than lysis of A549 pneumocytes was significantly inhibited by in vitro SOD but not by diluted serum. 5) PMN-mediated detachment rather than lysis of A549 pneumocytes showed a good correlation with superoxide anion production by PMN (R=0.90 in normal subjects, R=0.82 in asthmatics, p<0.05) but no correlation with plasma SOD activity. PMN-mediated pneumocyte injuries were not correlated with $FEV_1$ or $PC_{20}$ histamine in asthmatics. 6) There were no significant differences in PMN-mediated pneumocyte injuries between allergic and nonallergic asthmatics. Conclusion : Our results suggest that pneumocyte injury by PMN-derived superoxide anion may partially contribute to the pathogenesis of asthma and that cell detachment rather than cell lysis may be the mechanism of superoxide anion-mediated pneumocyte injury.

• The Korean Journal of Pharmacology
• /
• v.30 no.3
• /
• pp.299-311
• /
• 1994

The present study was attempted to investigate whether pentazocine, which is known to possess both opioid agonistic and antagonistic properties, produces catecholamines (CA) secretion from the isolated perfused rat adrenal gland, and to establish the mechanism of its action, and also to compare its action with that of some opioids. Pentazocine (30 to 300 ug) injected into an adrenal vein caused a dose-dependent secretory response of CA from the rat adrenal medulla. The pentazocine-evoked secretion of CA was remarkably diminished by the preloading with chlorisondamine $(10^{-6}\;M)$, naloxone $(1.22{\times}10^{-7}\;M)$, morphine $(1.7{\times}10^{-5}\;M)$, met-enkephalin $(9.68{\times}10^{-6}\;M)$, nicardipine $(10^{-6}\;M)$ and TMB-8 $(10^{-5}\;M)$ while was not influenced by the pretreatment of pirenzepine $(2{\times}10^{-6}\;M)$. The perfusion of $Ca^{++}$-free Krebs solution for 30 min into the gland also led to the marked reduction in CA secretion evoked by pentazocine. Furthermore, the CA release evoked by ACh and/or DMPP was greatly inhibited by the pretreatment with pentazocine $(1.75{\times}10^{-4}\;M)$ for 20 min. From these experimental results, it is thought that pentazocine causes markedly the increased secretion of CA from the isolated perfused rat adrenal medulla by a calcium-dependent exocytotic mechanism. The secretory effect of pentazocine appears to be mediated through activation of opioid receptors located on adrenal chromaffin cells, which may be also associated with stimulation of cholinergic nicotinic receptors.

• Biomolecules & Therapeutics
• /
• v.20 no.1
• /
• pp.43-49
• /
• 2012

Stimulation of mast cells through the high affinity IgE receptor (Fc${\varepsilon}$RI) induces degranulation, lipid mediator release, and cytokine secretion leading to allergic reactions. Although various signaling pathways have been characterized to be involved in the Fc${\varepsilon}$RI-mediated responses, little is known about the precious mechanism for the expression of tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) in mast cells. Here, we report that rapamycin, a specific inhibitor of mammalian target of rapamycin (mTOR), reduces the expression of TNF-${\alpha}$ in rat basophilic leukemia (RBL-2H3) cells. IgE or specific antigen stimulation of RBL-2H3 cells increases the expression of TNF-${\alpha}$ and activates various signaling molecules including S6K1, Akt and p38 MAPK. Rapamycin specifically inhibits antigeninduced TNF-${\alpha}$ mRNA level, while other kinase inhibitors have no effect on TNF-${\alpha}$ mRNA level. These data indicate that mTOR signaling pathway is the main regulation mechanism for antigen-induced TNF-${\alpha}$ expression. TNF-${\alpha}$ mRNA stability analysis using reporter construct containing TNF-${\alpha}$ adenylate/uridylate-rich elements (AREs) shows that rapamycin destabilizes TNF-${\alpha}$ mRNA via regulating the AU-rich element of TNF-${\alpha}$ mRNA. The antigen-induced activation of S6K1 is inhibited by specific kinase inhibitors including mTOR, PI3K, PKC and $Ca^{2+}$chelator inhibitor, while TNF-${\alpha}$ mRNA level is reduced only by rapamycin treatment. These data suggest that the effects of rapamycin on the expression of TNF-${\alpha}$ mRNA are not mediated by S6K1 but regulated by mTOR. Taken together, our results reveal that mTOR signaling pathway is a novel regulation mechanism for antigen-induced TNF-${\alpha}$ expression in RBL-2H3 cells.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.39 no.6
• /
• pp.813-819
• /
• 2010

The intracellular lipid droplets were stained with Oil Red O dye and quantified. Compared to the control, lipid accumulation was significantly decreased by 19.4% with the treatment of LCM at the concentration of $1000\;{\mu}g$/mL. Intracellular triglyceride (TG) level was also reduced by 21% at the concentration of $1000\;{\mu}g$/mL. To determine the mechanism for the reduction in TG content, levels of glucose uptake and glycerol release were measured. Incubation of the 3T3-L1 adipocytes with LCM did not affect the cellular uptake of glucose. However, the level of free glycerol released into the cultured medium drastically increased by 24.3% with the treatment of LCM. In subsequent measurements using quantitative real-time PCR, mRNA levels of hormone-sensitive lipase (HSL) and adipose triglyceride lipase (ATGL) except lipoprotein lipase (LPL) were significantly elevated at higher concentration. These results suggest that LCM partially stimulates the lipolysis through the induction of HSL and/or ATGL gene expression, resulting in the reduced lipid accumulation and increased glycerol release.

• The Korean Journal of Pharmacology
• /
• v.30 no.1
• /
• pp.87-100
• /
• 1994

It has been known for some time that dopamine-containing cells are existed in sympathetic ganglia, i.e., small, intensely fluorescent cells. However, its role and mechanism of action as a peripheral neurotransmitter are poorly understood so far. In the present study, an attempt was made to examine the effect of apomorphine, which is known to be a selective agonist of dopaminergic $D_2$. receptor on secretion of catecholamines (CA) from the isolated perfused rat adrenal gland. The perfusion of a low concentration of 10uM apomorphine into an adrenal vein for 20 min produced significant reduction in CA secretion induced by 5.32 mM ACh, 56 mM KCl, 100 uM DMPP and 100 uM McN-A-343. Increasing apomorphine concentration to 30 uM led to more markedly decreased CA secretion as compared to the case of 10 uM apomorphine and also did inhibit clearly CA release by $10^{-5}M$ Bay-K-8644. Furthermore, in adrenal glands preloaded with a higher dose of 100 uM apomorphine, CA releases evoked by ACh, excess $K^+$, DMPP and McN-A-343 were almost abolished by the drug. The perfusion of $3.3{\pm}10^{-5}M$ metoclopramide, which is well-known as a selective dopaminergic $D_2$ antagonist, produced significantly inhibitory effect of CA release by ACh, DMPP and McN-A-343 but did not affect that by excess $K^+$. However, preloading of 30uM apomorphine in the presence of metoclopramide did not modify the CA secretory effect of excess $K+$ and DMPP. These experimental results demonstrate that apomorphine causes dose-dependent inhibition of CA secretion by cholinergic receptor stimulation and also by membrane depolarization from the isolated perfused rat adrenal gland, suggesting that these effects appear to be exerted by inhibiting influx of extracellular calcium into the rat adrenal medullary chromaffin cells through activation of inhibitory dopaminergic receptors.