• Title/Summary/Keyword: intracellular pH

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Efficacy of Elaeagnus umbellata leaves on prevention of cadmium-induced toxicity in HepG2 cells

  • Jae-Yeul Lee;Seun-Ah Yang;Won-Bin Bae
    • Food Science and Preservation
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    • v.30 no.5
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    • pp.797-810
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    • 2023
  • Elaeagnus umbellata leaves have been reported to suppress inflammation, allergic responses, lung cancer proliferation and oral bacterial growth. Cadmium (Cd) is a heavy metal that has been found to cause many toxicities, including liver toxicity. The aim of this study was to evaluate the capacity of 70% ethanol extract of E. umbellata leaves (EUL) to protect human hepatocytes from Cd toxicity. After exposure of HepG2 cells to Cd at 10 𝜇M for 24 h, cell viability, expression levels of apoptosis- and antioxidant-related proteins, reactive oxygen species (ROS) accumulation and Cd uptake were assessed. EUL protected HepG2 cells from Cd-induced apoptosis as determined by MTT assay. A decrease in caspase-3 and p-p53 protein levels was observed in cells pretreated with EUL prior to Cd exposure. Furthermore, the Cd-induced increase in intracellular DCF fluorescence was attenuated by EUL, indicating that the Cd-induced apoptosis preventing effect was associated with the suppression of ROS accumulation. Moreover, EUL's effects on the inhibition of p38, JNK, and AKT phosphorylation also appear to be associated with protection against Cd toxicity. Moreover, EUL upregulated Cd-depressed expression of Nrf2, HO-1, catalase, and MT-1,2 proteins, suggesting that Cd uptake-induced apoptosis in HepG2 cells may be inhibited by EUL's antioxidative potential.

A study of Association of the H-FABP RFLP with Economic Traits of Pigs (돼지 H-FABP 유전자의 다형성 및 경제 형질과의 연관성 구명)

  • Choi, B.H.;Kim, T.H.;Lee, J.W.;Cho, Y.M.;Lee, H.Y.;Cho, B.W.;Cheong, I.C.
    • Journal of Animal Science and Technology
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    • v.45 no.5
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    • pp.703-710
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    • 2003
  • The purpose of this study was to detect association between genetic variation and economic trait in the porcine heart type fatty acid-binding protein gene as a candidate gene for the traits related with growth and meat quality in pigs. The H-FABP is a 15-kDa protein expressed in several tissues with high demand for fat metabolism such as cardiac and skeletal muscle and lactating mammary gland. H-FABP is small intracellular protein involved in fatty acid transport from the plasma membrane to the site of $\beta$-oxidation and/or triacylglycerol or phospholipid synthesis. In this study, H-FABP PCR-RFLP was performed in F$_2$ population composed of 214 individuals from an intercross between Korean Native Boars and Landrace sows. PCR products from two primer sets within H-FABP gene were amplified in 850bp and 700bp. Digestion of PCR products with the restriction digestion enzymes HaeⅢ and HinfⅠ, revealed fragment length polymorphisms(RFLPs). The genotype frequencies from H-FABP/HaeⅢ was .29 for genotype DD, .53 for genotype Dd, and .15 for genotype dd, respectively. The genotype frequencies of HH, Hh, and hh from H-FABP/HinfⅠ was .38, .41 and .20, respectively, in the population. Relationships between their genotypes and economic traits were estimated. In H-FABP/HaeⅢ locus, there were specific genotypes(Dd and dd) associated with economic traits such as body weights at 3, 5, 12, and 30 week of age (p〈.05 to .001). The ‘d’ allele was associated with gaining of body weight. In H-FABP/HinfⅠ locus, Genotypes of HH and Hh associated with growth traits such as body weights at 5, 12, and 30 week of age (p〈.05 or p〈.001) and back fat thickness, body fat including abdominal and trimmed fat (p〈.001) and intramuscular fat(p〈.05) The ‘H’ allele was positively associated with gaining of body weight and fatness deposition. In conclusion, a significant association of the H-FABP gene from its genetic variation was found on body weight, intramuscular fat and backfat thickness.

Pretreatment with Lycopene Attenuates Oxidative Stress-Induced Apoptosis in Human Mesenchymal Stem Cells

  • Kim, Ji Yong;Lee, Jai-Sung;Han, Yong-Seok;Lee, Jun Hee;Bae, Inhyu;Yoon, Yeo Min;Kwon, Sang Mo;Lee, Sang Hun
    • Biomolecules & Therapeutics
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    • v.23 no.6
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    • pp.517-524
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    • 2015
  • Human mesenchymal stem cells (MSCs) have been used in cell-based therapy to promote revascularization after peripheral or myocardial ischemia. High levels of reactive oxygen species (ROS) are involved in the senescence and apoptosis of MSCs, causing defective neovascularization. Here, we examined the effect of the natural antioxidant lycopene on oxidative stress-induced apoptosis in MSCs. Although $H_2O_2$ ($200{\mu}M$) increased intracellular ROS levels in human MSCs, lycopene ($10{\mu}M$) pretreatment suppressed $H_2O_2$-induced ROS generation and increased survival. $H_2O_2$-induced ROS increased the levels of phosphorylated p38 mitogen activated protein kinase (MAPK), Jun-N-terminal kinase (JNK), ataxia telangiectasia mutated (ATM), and p53, which were inhibited by lycopene pretreatment. Furthermore, lycopene pretreatment decreased the expression of cleaved poly (ADP ribose) polymerase-1 (PARP-1) and caspase-3 and increased the expression of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax), which were induced by $H_2O_2$ treatment. Moreover, lycopene significantly increased manganese superoxide dismutase (MnSOD) expression and decreased cellular ROS levels via the PI3K-Akt pathway. Our findings show that lycopene pretreatment prevents ischemic injury by suppressing apoptosis-associated signal pathway and enhancing anti-oxidant protein, suggesting that lycopene could be developed as a beneficial broad-spectrum agent for the successful MSC transplantation in ischemic diseases.

Production of Inulase Using Jerusalem Artichoke Tuber Extract (돼지감자 추출물을 이용한 이눌라아제 생산)

  • Choi, Weon-Sang;Choe, Yong-Kyung;Kim, Su-Il;Byun, Si-Myung
    • Applied Biological Chemistry
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    • v.27 no.4
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    • pp.238-244
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    • 1984
  • To produce inulase from Kluyveromyces fragilis No.351 using Jerusalem artichoke tuber extract, optimization of the condition was conducted. As results, the optimal concentration of artichoke tuber extract was 3.5 % and bactocasitone showed better production than yeast extract. The optimal temperature and pH were $30^{\circ}C$ and 5.5, respectively. The addition of $NH_4H_2PO_4$ increased the enzyme production. Inulase synthesis was growth-associated and the enzyme production increased as concentration of dissolved oxygen increased. A higher quantity of industrial grade inulase was prepared by the combination of ultrafiltration and ethanol precipitation with 72% recovery.

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Optimal Conditions for the Production of Intracellular Cytosine Deaminase from Chromobacterium violaceum YK 391. (Chromobacterium violaceum YK 391의 세포내 Cytosine Deaminase의 생성 최적조건)

  • Kim, Jung;Kim, Hyun-Soo;Yoo, Dae-Sik
    • Microbiology and Biotechnology Letters
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    • v.30 no.4
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    • pp.367-372
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    • 2002
  • Cytosine deaminase (cytosine aminohydrolase, EC 3.5.4.1) stoichiometrically catalyzes the hydrolytic deamination of cytosine and 5-fluorocytosine to uracil and 5-fluorouracil, respectively. Optimal medium compositions for production of cytosine deaminase from Chromobacterium violaceum YK 391 were 0.75% soluble starch, 1.5% peptone, 0.1% meat extract, 0.1% yeast extract, 0.01% NaCl, 0.01% $MgCl_2{\cdot}7H_2O$ and 0.05% $K_2HPO_4$. The optimal pH of medium and incubation temperature were 7.0 and $30^{\circ}C$, respectively. C. violaceum reached stationary phase after 30 hr, and produced a maximum cytosine deaminase (120 units/ml) after 72 h in batch culture.

Partial Characterization of Physicochemical and Kinetic Properties of $Ca^{++}-ATPase$ System in Luteal Membranes (황체막에서의 $Ca^{++}-ATPase$의 특성)

  • Choi, Gyu-Bog;Koo, Bon-Sook;Kim, In-Kyo
    • The Korean Journal of Physiology
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    • v.20 no.2
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    • pp.257-270
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    • 1986
  • It has been reported that the luteal function may be regulated by the intracellular calcium in luteal cells (Higuchi et al, 1976; Dorflinger et at, 1984; Gore and Behrman, 1984) which is adjusted partially by $Ca^{++}-ATPase$ activities in luteal cell membranes (Verma and Pennistion, 1981). However, the physicochemical and kinetic properties of $Ca^{++}-ATPase$ in luteal membranes were not fully characterized. This study was, therefore, undertaken to partially characterize the physicochemical and kinetic properties of $Ca^{++}-ATPase$ system in luteal membranes and microsomal fractions, known as an one of the major $Ca^{++}$ storge sites (Moore and Pastan, 1978), from the highly luteinized ovary Highly luteinized ovaries were obtained from PMSG-hCG injected immautre female rats. Light membrane and heavy membrane fractions and microsomal fractions were prepared by the differential and discontinuous sucrose density gradient centrifugation method desribed by Bramley and Ryan (1980). Light membrane and heavy membrane fractions and microsomal fractions from highly luteinized ovaries are composed of the two different kinds of $Ca^{++}-ATPase$ system. One is the high affinity $Ca^{++}-ATPase$ which is activated in low $Ca^{++}$ concentration (Km, 10-30 nM), the other is low affinity $Ca^{++}-ATPase$ activated in higher $Ca^{++}$ concentration $(K_{1/2},\;40\;{\mu}M)$. At certain $Ca^{++}$ concentrations, activities of high and low affinity $Ca^{++}-ATPase$ are the highest in light membrane fractions and are the lowest in microsomal fractions. It appeares that high affinity $Ca^{++}-ATPase$ system have 2 binding sites for ATP (Hill's coefficient; around 2 in all membrane fractions measured) and the positive cooperativity of ATP bindings obviously existed in each membrane fractions. The optimum pH for high affinity $Ca^{++}-ATPase$ activation is around S in all membrane fractions measured. The lipid phase transition temperature measured by Arrhenius plots of high affinity $Ca^{++}-ATPase$ activity is around $25^{\circ}C$. The activation energies of high affinity $Ca^{++}-ATPase$ below the transition temperature are similar in each membrane fractions, but at the above transition temperature, it is the hightest in heavy membrane fractions and the lowest in microsomal fractions. According to the above results, it is suggested that intracellular $Ca^{++}$ level, which may regulate the luteal function, may be adjusted primarily by the high affinity $Ca^{++}-ATPase$ system activated in intracellular $Ca^{++}$ concentration range $(below\;0.1\;{\mu}M)$.

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Body composition and hemodynamic changes in patients with special needs

  • Tsukamoto, Masanori;Hitosugi, Takashi;Esaki, Kanako;Yokoyama, Takeshi
    • Journal of Dental Anesthesia and Pain Medicine
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    • v.16 no.3
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    • pp.193-197
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    • 2016
  • Background: Some patients with special needs exhibit intellectual disability, including deficits in cognitive skills and decreased quality of life. The purpose of this study was to retrospectively compare changes in body composition and hemodynamics during general anesthesia in patients with and without special needs. Methods: The backgrounds of patients who underwent oral maxillofacial surgery under general anesthesia were recorded from medical records. Intracellular water (ICW), extracellular water (ECW), stroke volume variation (SVV), and heart rate (HR) were recorded for 3 h after the start of anesthesia. Categorical data were compared using an unpaired t-test, and a P-value of less than 0.05 was regarded as significant. Numerical data were compared using the Bonferroni correction, and a P-value of less than 0.0125 was regarded as significant. Results: A total of 21 patients were included in the study: 10 patients without special needs (non-S-group) and 11 patients with special needs (S-group). There were no significant differences in patients' backgrounds, except with regard to height (P = 0.03). In both groups, ICW and ECW were maintained, although they were lower in the S-group compared to the non-S-group. SVV was maintained in both groups, although it was higher in the S-group than the non-S-group. HR was significantly lower in the S-group 1 h after induction of anesthesia (P < 0.003). Conclusions: Changes in hemodynamics due to body fluid imbalance should be monitored during general anesthesia, especially for patients with special needs.

Nafamostat Mesilate Inhibits TNF-${\alpha}$-Induced Vascular Endothelial Cell Dysfunction by Inhibiting Reactive Oxygen Species Production

  • Kang, Min-Woong;Song, Hee-Jung;Kang, Shin Kwang;Kim, Yonghwan;Jung, Saet-Byel;Jee, Sungju;Moon, Jae Young;Suh, Kwang-Sun;Lee, Sang Do;Jeon, Byeong Hwa;Kim, Cuk-Seong
    • The Korean Journal of Physiology and Pharmacology
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    • v.19 no.3
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    • pp.229-234
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    • 2015
  • Nafamostat mesilate (NM) is a serine protease inhibitor with anticoagulant and anti-inflammatory effects. NM has been used in Asia for anticoagulation during extracorporeal circulation in patients undergoing continuous renal replacement therapy and extra corporeal membrane oxygenation. Oxidative stress is an independent risk factor for atherosclerotic vascular disease and is associated with vascular endothelial function. We investigated whether NM could inhibit endothelial dysfunction induced by tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$ ). Human umbilical vein endothelial cells (HUVECs) were treated with TNF-${\alpha}$ for 24 h. The effects of NM on monocyte adhesion, vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1) protein expression, p38 mitogenactivated protein kinase (MAPK) activation, and intracellular superoxide production were then examined. NM ($0.01{\sim}100{\mu}g/mL$) did not affect HUVEC viability; however, it inhibited the increases in reactive oxygen species (ROS) production and p66shc expression elicited by TNF-${\alpha}$ (3 ng/mL), and it dose dependently prevented the TNF-${\alpha}$ -induced upregulation of endothelial VCAM-1 and ICAM-1. In addition, it mitigated TNF-${\alpha}$ -induced p38 MAPK phosphorylation and the adhesion of U937 monocytes. These data suggest that NM mitigates TNF-${\alpha}$ -induced monocyte adhesion and the expression of endothelial cell adhesion molecules, and that the anti-adhesive effect of NM is mediated through the inhibition of p66shc, ROS production, and p38 MAPK activation.

Involvement of phospholipase $A_2$ in ATP-induced mucin release from cultured Hamster Tracheal Surface Epithelial cells

  • Jo, M.;Ko, K.H.;Kim, K.C.
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1996.04a
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    • pp.219-219
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    • 1996
  • Mucin release from hamster tracheal surface epithelial(HTSE) cells can be stimulated by extracellular ATP via activation of P$_2$ purinoceptors located on the cell surface which appears to be coupled to phospholipase C via G proteins. However, our preliminary data indicate that the ATP-induced mucin release involves, in part, activation of PKC, but not an increase in the intracellular Ca++ level, suggesting the presence of another pathway which is separate from the PLC-PKC pathway, In this study, we intended to confirm the previous observation and subsequently identify an additional mechanism. Confluent HTSE cells were metabolically labeled with either $^3$H-glucosamine or $^3$H-arachidonic acid(AA), and release of either $^3$H-mucin or $^3$H-AA was quantified following various treatments. $^3$H-mucin was assayed using the sepharose CL-4B gel-filtration method, whereas $^3$H-AA liberation was measured by counting $^3$H-radioactivity in the chase medium. We found that: (1)Desensitization of PKC by pretreatment with PMA completely abolished the mucin releasing effect of PMA but partially inhibited the ATP-induced mucin release; (2) ATP increases release of $^3$H-AA in a dose-dependent fashion; (3) mepacrine, an inhibitor of PLA$_2$, attenuates ATP-induced mucin release in a dose-dependent fashion. These results confirm our previous notion that the PLC-PKC pathway is responsible, in part, for ATP-induced mucin release. Furthermore, activation of PLA$_2$ appears to be an additional pathway which is involved in ATP-induced mucin release.

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Heterologous Expression of Human $\beta$-Defensin-1 in Bacteriocin-Producing Laetoeoeeus lactis

  • CHOI HAK JONG;SEO MYUNG JI;LEE JUNG CHOUL;CHEIGH CHAN ICK;PARK HOON;AHN CHEOL;PYUN YU RYANG
    • Journal of Microbiology and Biotechnology
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    • v.15 no.2
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    • pp.330-336
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    • 2005
  • Lactococcus lactis A164 is a nisin Z-producing strain isolated from kimchi. Its antimicrobial spectrum has been found to be active against most Gram-positive bacteria tested, yet inactive against Gram-negative bacteria [3]. Accordingly, to overcome this drawback, the current study attempted to express human $\beta$-defensin-l (hBD-l), which kills both Gram-positive and Gram-negative bacteria in L. lactis AI64. When the hBD-l cDNA was introduced using a nisin Z-controlled expression cassette, the L. lactis A164 transformants grew very poorly, due to the bactericidal effect of the expressed hBD-l against the transformants. Therefore, a gene fusion system was designed to reduce the toxicity of the expressed heterologous protein against the host cells. As such, the hBD-l gene was fused to the DsbC- Tag of pET -40b(+), then introduced to L. lactis A 164. The transformants expressed an intracellular 35.6-kDa DsbC-hBD-l fusion protein that exhibited slight activity against the host cells, yet not enough to strongly inhibit the cell growth. To obtain the recombinant hBD-l, the DsbC-hBD-l fusion protein was purified by nickel-affinity column chromatography, and the DsbC-Tag removed by cleaving with enterokinase. The cleaved mature hBD-l exhibited strong bactericidal activity against E. coli JM109, indicating that the recombinant L. lactis A 164 produced a biologically active hBD-I. In addition, the recombinant L. lactis A 164 was also found to produce the same level of nisin Z as the wild-type.