• Journal of Microbiology and Biotechnology
• /
• v.15 no.4
• /
• pp.756-766
• /
• 2005

The signaling mechanism underlying aburatubolactam C-induced FasL upregulation was investigated in human Jurkat T cells. After treatment with aburatubolactam C, the src-family PTKs $p56^{lck}\;and\;p59^{fyn}$, and MAP kinases ERK2 and JNK1, were activated prior to FasL upregulation; Both $p56^{lck}\;and\;p59^{fyn}$ were directly activated 2.4- and 2.2-fold, respectively, in vitro by aburatubolactam C. The aburatubolactam C-induced cellular changes, including the activation of ERK2 and INK1, and FasL upregulation, were completely prevented by the PTK inhibitor genistein. The activation of protein kinase C (PKC$\delta,\;\epsilon\;and\;\mu$ was also induced following aburatubolactam C treatment. Although the activation of $p56^{lck}$ and tyrosine phosphorylation of the cellular proteins were not blocked by the PKC inhibitor GFl09203X, the activation of ERK2 was completely abrogated, along with a detectably enhanced JNK1 activation; FasL upregulation, and apoptosis. However, the FasL upregulation and apoptosis were significantly inhibited by the PKC activator PMA, with a remarkable increase in the ERK2 activation. The cytotoxic effect of aburatubolactam C was reduced in the presence of the anti-Fas neutralizing antibody ZB-4. Although ectopic expression of Bcl-2 failed to completely block the cytotoxicity of aburatubolactam C, it was clearly suppressed. The c-Fos mRNA expression was upregulated in a biphasic manner, where the second phasic expression overlapped with the FasL upregulation. Accordingly, these results demonstrate that aburatubolactam C-induced apoptosis is exerted, at least in part, by FasL upregulation dictated by activation of the PTK ($p56^{lck}\;and\;p59^{fyn}$) /JNKI pathway, which is negatively affected by the concurrent activation of the PKC/ERK2 pathway proximal to PTK activation.

• The Korean Journal of Physiology
• /
• v.17 no.2
• /
• pp.125-133
• /
• 1983

Red cell glycolytic intermediates, metabolites and metabolic ratios were studied. Glycolytic intermediates were measured in neutralized perchloric acid extracts of red cell suspensions after 3 hr incubation at $37^{\circ}C$ in the presence and absence of saponin. Adenosine triphosphate(ATP), adenosine diphosphate(ADP), pyruvate and lactate were measured by enzymatic procedures involving stoichiometric oxidation or reduction of a pyridine nucleotide. Glucose was determined using glucose oxidase after zinc hydroxide extraction. The redox state was calculated from the lactate dehydrogenase equilibrium. Adenosine triphosphatase activity(ATPase) was measured by determining the amount of phosphate released from ATP by washed erythrocyte membranes(ghost) during 20 min. incubation. Both total hydrolysis and the amount of hydrolysis that occured in the presence of ouabain were measured. The second measurement yields Mg-ATPase and represents nonspecific ATPase activity of the membranes. The difference between total and Mg-ATPase activity can be attributed to Na-K-ATPase. For the measurement of sodium fluxes, human erythrocytes were preincubated in $^{22}Na$ for 3 hr at $37^{\circ}C$, washed and suspended in a tracer-free medium. The amount of $^{22}Na$ transported out of cells at any time was determined by analysis of supernatant samples taken at various time after addition of the labeled cells to isotope-free medium. The cells and medium were separated and the radioactivity appearing in the medium was measured. From the total radioactivity in the suspension and the radioactivity appearing in the medium at known time, the rate constant for sodium release was computed. The results are summarized as follows: 1) ATP and ATP/ADP were found to increase at every concentration of saponin tested whereas ADP declined at every cone. of saponin. The increase in pyruvate and lactate were observed at every cone, of saponin and thus $NAD^+/NADH$ computed from pyruvate/lactate also increased. Glucose utilization was stimulated by saponin. 2) $Na^+-K^+-ATPase$ activities showed a biphasic response to saponin, first increasing in lower concentration and then decreasing in higher concentration of saponin. 3) The efflux of sodium was significantly increased by saponin in the range of 5 to 10 mg%. The stimulatory effect of saponin on the rate constants for active(ouabain-sensitive) sodium efflux was inhibited by addition of ouabain.

• Asian-Australasian Journal of Animal Sciences
• /
• v.20 no.5
• /
• pp.693-700
• /
• 2007

The present study was conducted to examine the effect of different levels of essential and nonessential amino acid in NCSU-23 medium on the in vitro-produced porcine embryo as it develops from the zygote to the blastocyst stage. Four experiments were performed, each with a completely randomized design involving 5 to 8 replications of treatments. In order to know the effect of nonessential amino acids in NCSU-23 medium, 0, 5, 10 and $20{\mu}/ml$ MEM were supplemented there to, (Exp. 1) and the medium was supplemented with same level of essential amino acids (Exp. 2). The combined effect of nonessential (0, 5, 10 and $20{\mu}/ml$ MEM) and essential amino acids (0, 5, 10 and $10{\mu}/ml$ MEM) in NCSU-23 medium (Exp. 3), first 72 h with non-essential amino acids (at 0, 5, 10 and $20{\mu}/ml$ MEM), and last 4 d with essential amino acids with the same level as NEAA (Exp. 4) were examined. The embryo development was monitored and the quality of blastocysts was evaluated by counting the number of total cells and determining the ratio of inner cell mass (ICM) to trophoectoderm (TE) cells. When Eagle's nonessential amino acids (MEM) added to NCSU-23 medium, it significantly increased the likelihood of development to the 2- to 4-cell stage and subsequent blastocyst development. Supplementation of different levels of essential amino acids in the NCSU-23 medium decreased cleavage rate, rate of morula and blastocyst development and the number of ICMs. In the case of the combined effect of essential and nonessential amino acids, better and significant results were found for blastocysts, hatching blastocysts and for ICM numbers which were also dose dependent. With respect to the biphasic effect of nonessential and essential amino acids, nonessential amino acids increased cleavage whereas essential amino acids increased the total cell number. Neither the nonessential nor the essential group of amino acids, on their own, affected blastocyst cell number or the differentiation of cells in the blastocyst. In conclusion, this study determined the role of nonessential and essential amino acids in the culture of the porcine embryo and showed that the embryo requires different levels of amino acids as it develops from the zygote to the blastocyst stage.

• Journal of Microbiology and Biotechnology
• /
• v.26 no.4
• /
• pp.790-798
• /
• 2016

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.

• The Korean Journal of Physiology
• /
• v.24 no.1
• /
• pp.91-102
• /
• 1990

The effects of extracellular $Ca^{2+}$ and various $Ca^{2+}$ antagonists on endothelium-dependent relaxation to acetylcholine were studied in the isolated rabbit thoracic aorta in order to elucidate the control mechanism of endothelium derived relaxing factor (EDRF) release. Endothelium was removed from aortic strips by gentle rubbing with cotton ball. The effect of hemoglobin on basal tension was also observed with hemolysate. The results obtained were as follows: 1) Endothelium-dependent relaxation (EDR) to acetylcholine (ACh) showed biphasic pattern; the initial rapid relaxation phase and the late slow relaxation phase. 2) With the depletion of the extracellular $Ca^{2+}$, EDR was gradually suppressed, especially the late slow relaxation. 3) Verapamil, nifedipine, $Mn^{2+}$ and $Cd^{2+}$ had not any effect on EDR, while $La^{3+}$ and $Co^{2+}$ suppressed EDR completely. 4) The resting tension of the strips with rubbed endothelium was not altered by the addition of hemoglobin. That of the strips with intact endothelium, however, was enhanced and EDR to ACh was completely blocked From these results, we suggest that extracellular $Ca^{2+}$ is necessary for ACh-induced slow relaxation while $Ca^{2+}$ antagonists have not any effect on EDR.

• YAKHAK HOEJI
• /
• v.41 no.3
• /
• pp.335-344
• /
• 1997

The absorption, distribution and excretion of $^{14}C$ labeled YH1885 {5,6-Dimethyl-2(4-fluorophenylamino)-4-(1-methyl-1,2,3,4-tetrahydroisoquinolin-2-yl)pyrimidine hydroc hloride), a new proton pumpinhibitor, were investigated in rats after a single administration of $^{14}C$-YH1885. 1. After intravenous administration of 5mg/kg, the blood level of radioactivity declined in a biphasic fashion with the mean terminal elimination half-life of 12.4hr. 2. After oral administration of 20mg/kg, the maximum blood level of radioactirity was reached at 4.0hr in female rats. The blood level of radioactivity-time profiles in male and female rats were similar, and the absorptionof $^{14}C$-YH1885 was not affected by food. 3. Appproximately 89% and 1% of radioactivity of the total dose were excreted in feces and urine, respectively. 4. Biliary excretion of radioactivity was 47.9% of the dose. Enterohepatic circulation of radioactivity was 49.6%. 5. Radioactivity was excreted maily into feces via bile. 6. The concentration of radioactivity in most tissues reached the peak level at 4.0hr after dosing, and then declined. Autoradiograms of male rats showed that the radioactivity levlels in the fat, harder's gland, liver and G-Itract were higher than those in the other tissues and the elimination of radioactivity from fat and liver was slow. 7. Autoradiograms of a pregnant rat showed that radioactivity was transferred to mammary gland, placenta and fetus. The radioactivity level in the mammary gland was higher than that in the blood.

• Journal of the korean academy of Pediatric Dentistry
• /
• v.25 no.4
• /
• pp.788-796
• /
• 1998

The purpose of this study was to investigate the effect of various electroacupuncture duration induced by acupuncture point-Zusanli ($S_{36}$) electrical stimulation on inhibition of amplitude of digastric electromyogram (dEMG) evoked by noxious electrical stimuli around the mental foramen. intraperitoneal sodium pentobarbital in an initial dose of 50mg/kg and maintenance doses of 4.5mg/kg/h were given through a cannula in the femoral vein using a constant infusion pump. A pair of stimulating electrodes were inserted for noxious stimuli around the mental foramen. An irritant electronic stimuli pulse (0.2 Hz, 0.1 ms duration) was produced with an intensity of about $1.5{\times}2$ times threshold for evoking the dEMG. The anterior belly of the digastric muscle was exposed and a pair of 0.1mm wire electrodes were inserted for dEMG recording. Acupuncture point stimulation on Zusanli (2 Hz, 250 ${\mu}s$, biphasic pulse, 2 V) was delivered by Dental Electronic Anesthesia (3M, U.S.A). For periods of electronic stimulation of 10, 20, and 30min, the amplitudes of dEMG were measured on the oscilloscope and on the monitor connected to the amplifier. The following results were obtained: The dEMG was decreased to 73.4% of that in the control set after 10 min electroacupunture stimulation (Group I); The dEMG was decreased to 77.1% (10min), 54.0.% (20min) of that in the control set after 20minutes of electroacupunture stimulation (Group II). The dEMG was decreased to 73.3% (10min), 61.9% (20min), 76.2% (30min) of that in the control set after 30 min of electroacupunture stimulation (Group III). From these results, it may be that in the electroacupuncture stimulation on the Zusnali resulted in a reduction of amplitude of dEMG and that the most effective electroacupuncture stimulation period was 20min.

• Biomolecules & Therapeutics
• /
• v.25 no.4
• /
• pp.452-459
• /
• 2017

In this study, the effect of particle size of genistein-loaded solid lipid particulate systems on drug dissolution behavior and oral bioavailability was investigated. Genistein-loaded solid lipid microparticles and nanoparticles were prepared with glyceryl palmitostearate. Except for the particle size, other properties of genistein-loaded solid lipid microparticles and nanoparticles such as particle composition and drug loading efficiency and amount were similarly controlled to mainly evaluate the effect of different particle sizes of the solid lipid particulate systems on drug dissolution behavior and oral bioavailability. The results showed that genistein-loaded solid lipid microparticles and nanoparticles exhibited a considerably increased drug dissolution rate compared to that of genistein bulk powder and suspension. The microparticles gradually released genistein as a function of time while the nanoparticles exhibited a biphasic drug release pattern, showing an initial burst drug release, followed by a sustained release. The oral bioavailability of genistein loaded in solid lipid microparticles and nanoparticles in rats was also significantly enhanced compared to that in bulk powders and the suspension. However, the bioavailability from the microparticles increased more than that from the nanoparticles mainly because the rapid drug dissolution rate and rapid absorption of genistein because of the large surface area of the genistein-solid lipid nanoparticles cleared the drug to a greater extent than the genistein-solid lipid microparticles did. Therefore, the findings of this study suggest that controlling the particle size of solid-lipid particulate systems at a micro-scale would be a promising strategy to increase the oral bioavailability of genistein.

• Biomedical Science Letters
• /
• v.6 no.4
• /
• pp.229-235
• /
• 2000

In situ brain-pancreas perfusion was performed on male adult Sprague-Dawley rats, of which the central nervous systems (CNS) were intact during the perfusion procedure. The modified Krebs-Ringer buffer with 100 mg/dL of glucose and 20 mM of arginine was perfused for 30 min. In the experimental groups, a cephalic glucopenia was induced at 0 min (GLP1 group) or at 16 min (GLP2 group). The glucopenia was not induced in the control (CONT group). Insulin and glucagon concentrations in the effluent samples from the pancreas were measured using a RIA method. In all three groups, the first and second phases in the dynamics of the insulin and glucagon secretion were observed, which was a typical biphasic secretory pattern. The amount of insulin secretion tended to decrease in the GLP1 and GLP2 groups, but there was no statistically significant difference among the groups. However, the amount of glucagon secretion during 0~15 min of the perfusion period in the GLP1 group was greater as compared to the CONT group (p<0.05). The amount of glucagon secretion during 16~30 min of the perfusion period in the GLP2 group tended to be greater as compared to the CONT group, however there was no statistical significance. These data indicate that the cephalic glucopenia stimulates the direct secretion of glucagon from the pancreas during the early period of perfusion in the CNS-intact pancreatic perfused rats.

• The Korean Journal of Physiology
• /
• v.18 no.2
• /
• pp.139-150
• /
• 1984

Vanadate is a potent inhibitor of Na-K-ATPase. Ouabain, the another specific inhibitor of Na-K-ATPase, induces the contraction in cardiac muscle and smooth muscle. But, some investigators observed the discrepancies between vanadate and ouabain-induced contraction in cardiac muscle. The difference of vanadate and ouabain-induced contraction was investigated in the cat ileal smooth muscle. The following results were obtained. 1) Ouabain-induced contraction was biphasic, but vanadate-induced contraction had one peak. 2) Atropine inhibited ouabain·induced contraction, but did not inhibit vanadate-induced contraction. 3) Changes in external $Ca^{++}$concentration or $Ca^{++}$ antagonists had a greater influence on the contraction induced hy vanadate than by ouabain. 4) Removal of $Na^+$ from incubation medium and high $K^+$ abolished ouabain-induced contraction, but had no effect on vanadate-induced contraction. 5) Vanadate-induced contraction was potentiated in the presence of ouabain. 6) After 3 hrs incubation with vanadate, there was no change in intracellular $Na^+$ concentrations in contrast with ouabain. These results suggest that vanadate contracts ileal smooth muscle through the mechanism different from ouabain, and this is independent of the inhibition of Na-K-ATPase activity.