• Title/Summary/Keyword: protease treatment

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p53 signaling is involved in leptin-induced growth of hepatic and breast cancer cells

  • Shrestha, Mohan;Park, Pil-Hoon
    • The Korean Journal of Physiology and Pharmacology
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    • v.20 no.5
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    • pp.487-498
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    • 2016
  • Leptin, an adipokine predominantly produced from adipose tissue, is well known to induce tumor growth. However, underlying molecular mechanisms are not established yet. While p53 has long been well recognized as a potent tumor suppressor gene, accumulating evidence has also indicated its potential role in growth and survival of cancer cells depending on experimental environments. In the present study, we examined if p53 signaling is implicated in leptin-induced growth of cancer cells. Herein, we demonstrated that leptin treatment significantly increased p53 protein expression in both hepatic (HepG2) and breast (MCF-7) cancer cells without significant effect on mRNA expression. Enhanced p53 expression by leptin was mediated via modulation of ubiquitination, in particular ubiquitin specific protease 2 (USP2)-dependent manner. Furthermore, gene silencing of p53 by small interfering RNA (siRNA) suppressed leptin-induced growth of hepatic and breast cancer cells, indicating the role of p53 signaling in tumor growth by leptin. In addition, we also showed that knockdown of p53 restored suppression of caspase-3 activity by leptin through modulating Bax expression and prevented leptin-induced cell cycle progression, implying the involvement of p53 signaling in the regulation of both apoptosis and cell cycle progression in cancer cells treated with leptin. Taken together, the results in the present study demonstrated the potential role of p53 signaling in leptin-induced tumor growth.

Changes of Serum Levels of Alpha 1-Proteinase Inhibitor in Pre-and Post Open Heart Surgery (개심술 전후의 Alpha 1-Proteinase Inhibitor 의 추이)

  • Jeong, Jong-Hwa;Kim, Song-Myeong
    • Journal of Chest Surgery
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    • v.22 no.3
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    • pp.402-415
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    • 1989
  • Extracorporeal circulation leads to functional disorder and structural damage of organs, especially hematologic and pulmonary system, mainly by sequestration of neutrophils and deposition of macrophages at lung. Then, proteases are secreted, which insult vascular basement membrane of pulmonary capillary and alveolar septa of the lung. Among these, the most important protease at lung is elastase, because major component of lung is elastin. For prevention of lung injury, inactivators or antidotes to elastase should be necessary and Alpha 1-Proteinase Inhibitor is the elastase inactivator. Clinical experimental study was carried out to investigate the immediate postoperative change of serum Alpha 1-PI level following cardiopulmonary bypass for 20 heart cases [congenital 16 cases, acquired 4 cases] and 10 control [subtotal gastrectomy] cases. Also preliminary study was performed for 31 cases of open heart patients. The results were as follows: l. Immediate postoperative serum levels of Alpha 1-PI was significantly decreased at open heart surgery group [P< 0.005], but not decreased at control group. 2. There were no significant difference in change of serum Alpha 1-PI level between and membrane and bubble oxygenator group.Z 3. There were no significant difference in changes of serum Alpha 1-PI level between CHD and AHD. Alpha 1-PI is consumed at lung during cardiopulmonary bypass and increase after operation compensatedly and protect multiple organic damage especially lung. Therefore, Alpha 1-PI can be indicator for evaluation of prevention and treatment of pump-lung syndrome.

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Biochemical Characterization of a Putative Calcium Influx Factor as a Diffusible Messenger in Jurkat Cells, Xenopis Oocytes, and Yeast

  • Kim, Hak-Yong
    • Animal cells and systems
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    • v.7 no.1
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    • pp.75-79
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    • 2003
  • Highly purified high performance thin layer chromatography (HPTLC) fractions containing a putative calcium influx factor (CIF) were prepared from the Jurkat cells and Xenopus oocytes in which $Ca^{2+}$ stores were depleted by thapsigargin treatment and from the yeast in which intracellular $Ca^{2+}$ stores were also depleted by genetic means. Microinjection of the fractions has been shown to elicit $Ca^{2+}$ dependent currents in Xenopus oocytes. The nature of the membrane currents evoked by the putative CIF appeared to be carried by chloride ions since the current was blocked by the selective chloride channel blocker 1 mM niflumic acid and its reversal potential was about -24 mV. Injection of the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N, N, N',N'-tetraacetic acid (BAPTA) eradicated the current activities, suggesting the current responses are entirely $Ca^2$-dependent. Moreover, the currents were sensitive to the removal of extracellular calcium, indicating the dependence on calcium entry through the plasma membrane calcium entry channels. CIF activities were insensitive to protease, heat, and acid treatments and to Dische-reaction whereas the activities were sensitive to nucleotide pyrophosphatase and hydrazynolysis. The fraction might have a sugar because it was sensitive to Molisch test and Seliwaniff's resorcinol reaction. From the above results, CIF as a small and stable molecule seems to have pyrimidine, pyrophosphate, and a sugar moiety.oiety.

Isolation of a Lactococcus lactis Strain Producing Anti-staphylococcal Bacteriocin

  • Yang, Jung-Mo;Moon, Gi-Seong
    • Food Science of Animal Resources
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    • v.38 no.6
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    • pp.1315-1321
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    • 2018
  • Bacteriocin is ribosomally synthesized by bacteria and inhibits closely related species. In this study we aimed at isolating lactic acid bacteria producing bacteriocin presenting anti-staphylococcal activity. A Lactococcus lactis strain was isolated from kimchi for the purpose and identified by 16S rRNA gene sequencing. As preliminary tests, optimal culture conditions, stabilities against heat, solvents, and enzymes treatments, and type of action (bacteriostatic or bactericidal) of the bacteriocin were investigated. The optimal culture conditions for production of the bacteriocin were MRS broth medium and $25^{\circ}C$ and $30^{\circ}C$ culture temperatures. The bacteriocin was acidic and the activity was abolished by a protease treatment. Its stability was maintained at $100^{\circ}C$ for 15 min and under treatments of various organic solvents such as methanol, ethanol, acetone, acetonitrile, and chloroform. Finally, the bacteriocin showed bactericidal action against Staphylococcus aureus where 200 AU/mL of the bacteriocin decreased the viable cell count (CFU/mL) of S. aureus by 2.5 log scale, compared with a control (no bacteriocin added) after 4-h incubation.

Preparation of Yeast Extract from Waste Brewer's Yeast using Various Enzymes (각종 효소를 이용한 맥주 폐효모로부터 효모추출물 제조)

  • Lee, Ok-Hwan;Rhee, Seong-Kap;Son, Jong-Youn;Kim, Kyung-Im;Kim, Hyun-Duk;Lee, Boo-Yong
    • Korean Journal of Food Science and Technology
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    • v.34 no.5
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    • pp.867-872
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    • 2002
  • This study was performed to investigate the optimum process conditions for manufacturing yeast extract from waste brewer's yeast using various enzymes. Contents of IMP, GMP, free amino acids, and crude protein of yeast extracts were measured by enzymes treatment. Crude protein contents of yeast extracts subjected to cell wall digestion enzyme treatment were 21.1, 33.6, and 28.0% for the control grouup, glucanase (0.5%, 12 h), and tunicase (1%, 18 h), respectively. Crude protein contents of yeast extracts subjected to protease treatment were 22.0, 30.8, and 29.8% for control group, bromelin (1%, 3 h), and protamex (1%, 3 h), respectively. Crude protein content of yeast extract subjected to glucanase and protamex mixed treatment was 34.4%. The total contents of IMP and GMP of yeast extracts subjected to G+P+A (glucanase+phosphodiesterase+adenyldeminase) and G+Pro+P+A (glucanase+protamex+phosphodiesterase+adenyldeaminase) treatments were 1,066 and 1,047 mg/100 g, respectively. The content of free amino acids of yeast extract was the highest (2,302 mg/100 g) in G+Pro+P+A treatment. Optimum concentration and process condition of enzyme treatment to obtain yeast extract with high IMP, GMP, and free amino acid content were in the order of glucanase (0.5%, 12 h), protamex (1%, 3h), phosphodiesterase (0.1%, 3 h) and adenyldeaminase (1%, 1.5 h) treatments.

Effects of different inorganic: organic zinc ratios or combination of low crude protein diet and mixed feed additive in weaned piglet diets

  • Oh, Han Jin;Kim, Myung Hoo;Lee, Ji Hwan;Kim, Yong Ju;An, Jae Woo;Chang, Se Yeon;Go, Young Bin;Song, Dong Cheol;Cho, Hyun Ah;Jo, Min Seok;Kim, Dae Young;Kim, Min Ji;Cho, Sung Bo;Kim, Hyeun Bum;Cho, Jin Ho
    • Journal of Animal Science and Technology
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    • v.64 no.1
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    • pp.23-37
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    • 2022
  • Thirty-six weaned piglets with an initial body weight (BW) of 8.43 ± 0.40 kg (28 days of age, ([Landrace × Yorkshire] × Duroc) were randomly assigned to 6 treatments for a 2-week feeding trial to determine the effects of different inorganic zinc (IZ), organic zinc (OZ) or combination of low crude protein diet (LP) and Mixed feed additive (MFA) on diarrhea score, nutrient digestibility, zinc utilization, blood profiles, organ weight, and fecal microflora in weaned piglet diet. The pigs were individually placed in 45 × 55 × 45 cm stainless steel metabolism cages in an environmentally controlled room (30 ± 1℃). The dietary treatments included a negative control (NC), positive control (PC; zinc oxide, 1,000 mg/kg), T1 (IZ : OZ, 850 : 150), T2 (IZ : OZ 700 : 300), T3 (IZ : OZ, 500 : 500), and T4 (LP + MFA [0.1% Essential oils + 0.08% Protease + 0.02% Xylanase]). The daily feed allowance was adjusted to 2.7 times the maintenance requirement for digestible energy (2.7 × 110 kcal of DE/kg BW0.75). This allowance was divided into two equal parts, and the piglets were fed at 08 : 30 and 17 : 30 each day. Water was provided ad libitum through a drinking nipple. The diarrhea score was significantly increased (p < 0.05) in NC treatment compared with other treatments. The apparent total tract digestibility (ATTD) of dry matter (DM), nitrogen (N), and gross energy (GE) was significantly increased (p < 0.05) in the T2 treatment compared with the PC and NC treatments in week 1. In week 2, the ATTD of DM, N, and GE was significantly decreased (p < 0.05) in the NC treatment compared with other treatments. The T3 treatment had significantly higher (p < 0.05) ATTD and apparent ileal digestibility of zinc than the PC and T1 treatments. The Escherichia coli count in feces was significantly decreased in the T4 treatment compared with the NC and T2 treatments. The Lactobacillus count in feces was significantly increased in the T4 and T1 treatment compared with the T2 and T3 treatments. In conclusion, IZ : OZ 500 : 500 levels could improve nutrient digestibility and zinc utilization in weaned piglets, Moreover, MFA in LP diets could be used as a zinc alternative.

Comparison of Physicochemical and Sensory Properties of Freeze-concentrated Milk with Evaporated Milk during Storage

  • Hwang, J.H.;Lee, S.J.;Park, H.S.;Min, S.G.;Kwak, H.S.
    • Asian-Australasian Journal of Animal Sciences
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    • v.20 no.2
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    • pp.273-282
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    • 2007
  • This study was carried out to compare the changes of nutrients, sensory and chemical properties of freeze-concentrated and evaporated milks during storage. For pasteurization, the freeze-concentrated milk containing 27% of total solid was treated with 150 rpm ozone for 5 min, and 99% of microflora was eliminated. Also, the activities of protease and lipase decreased 93.31% and 96.15%, respectively, and phosphatase showed negative activity. Total bacteria count was maintained below$2.0{\times}10^4$CFU/ml. During storage, TBA absorbance was lower in freeze-concentrated milk than that in the evaporated milk. The production of short-chain free fatty acids and free amino acids increased proportionally to the storage period in both samples. While the short-chain free fatty acid production was lower in the freeze-concentrated milk compared with that in the evaporated milk, the production of individual free amino acid was similar in both samples. In sensory evaluation, cooked flavor and color were much lower in the freeze-concentrated milk than that in the evaporated milk. Overall acceptability score was higher in the freeze-concentrated than the evaporated milk. Based on above results, ozone treatment for the freeze-concentrated milk pasteurization was positive at the elimination of microflora and enzyme inactivation. During storage, the freeze-concentrated sample minimized the change of color and TBA absorbance, the production of short-chain free fatty acid and vitamins than the evaporated milk. Therefore, the freeze-concentrated milk process in the present study resulted in the positive effect in minimizing nutrient loss and keeping quality of milk during storage.

Screening of Bacterial Surface Display Anchoring Motif Using Tetrameric β-galactosidase in Bacillus subtilis Spore (Tetrameric β를 이용한 고초균 포자에서의 미생물 표면 발현 모체 선별)

  • Kim, June-Hyung;Pan, Jae-Gu;Kim, Byung-Gee
    • KSBB Journal
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    • v.26 no.3
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    • pp.199-205
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    • 2011
  • Using tetrameric ${\beta}$-galactosidase as a model protein, anchoring motives were screened in Bacillus subtilis spore display system. Eleven spore coat proteins were selected considering their expression levels and the location in the spore coat layer. After chromosomal single-copy homologous integration in the amyE site of Bacillus subtilis chromosome, cotE and cotG were chosen as possible spore surface anchoring motives with their higher whole cell ${\beta}$-galactosidase activity. PAGE and Wester blot of extracted fraction of outer layer of purified spore, which express CotE-LacZ or CotG-LacZ fusion verified the existence of exact size of fusion protein and its location in outer coat layer of purified spore. ${\beta}$-galactosidase activity of spore with CotE-LacZ or CotG-LacZ fusion reached its highest value around 16~20 h of culture time in terms of whole cell and purified spore. After intensive spore purification with lysozyme treatment and renografin treatment, spore of BJH135, which expresses CotE-LacZ, retained only 1~2% of its whole cell ${\beta}$-galactosidase activity. Whereas spore of BJH136, which has cotG-lacZ cassette in the chromosome, retained 10~15% of its whole cell ${\beta}$-galactosidase activity, proving minor perturbation of CotG-LacZ, when incorporated in the spore coat layer of Bacillus subtilis compared to CotE-LacZ. Usage of Bacillus subtilis WB700, of which 7 proteases are knocked-out and thereby resulting in 99.7% decrease in protease activity of the host, did not prevent the proteolytic degradation of spore surface expressed CotG-LacZ fusion protein.

Effects of Enzyme Treatments and Ultrasonification on Extraction Yields of Lipids and Protein from Soybean by Aqueous Process (Aqueous Process를 이용한 대두유와 대두단백의 추출중 효소와 초음파처리가 추출율에 미치는 영향)

  • Yoon, Suk-Hoo;Kim, In-Hwan;Kim, Seung-Ho;Kwon, Tai-Wan
    • Korean Journal of Food Science and Technology
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    • v.23 no.6
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    • pp.673-676
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    • 1991
  • Lipids and protein were extracted simultaneously from soybean flour by aqeous processing. Extraction yields of lipids and protein were 62 and 68%, respectively, when 120-150 mesh full-fat soybean flour was dispersed in six times of water (w/w) at $40^{\circ}C$ and pH 8. Supplementary treatment for the higher extraction yields such as proteolytic enzymes treatment improved extraction yields of lipids and protein up to 86 and 89%, respectively. Ultrasonification also improved extraction yields of lipids and protein up to 90%. Red and yellow colors of aqeous-extracted soybean oil were slightly darker than those of hexane-extracted oil, but were much lighter in colors than those of Folch-extracted oil.

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Multiple Signaling Pathways Contribute to the Thrombin-induced Secretory Phenotype in Vascular Smooth Muscle Cells

  • Jeong, Ji Young;Son, Younghae;Kim, Bo-Young;Eo, Seong-Kug;Rhim, Byung-Yong;Kim, Koanhoi
    • The Korean Journal of Physiology and Pharmacology
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    • v.19 no.6
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    • pp.549-555
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    • 2015
  • We attempted to investigate molecular mechanisms underlying phenotypic change of vascular smooth muscle cells (VSMCs) by determining signaling molecules involved in chemokine production. Treatment of human aortic smooth muscle cells (HAoSMCs) with thrombin resulted not only in elevated transcription of the (C-C motif) ligand 11 (CCL11) gene but also in enhanced secretion of CCL11 protein. Co-treatment of HAoSMCs with GF109230X, an inhibitor of protein kinase C, or GW5074, an inhibitor of Raf-1 kinase, caused inhibition of ERK1/2 phosphorylation and significantly attenuated expression of CCL11 at transcriptional and protein levels induced by thrombin. Both Akt phosphorylation and CCL11 expression induced by thrombin were attenuated in the presence of pertussis toxin (PTX), an inhibitor of Gi protein-coupled receptor, or LY294002, a PI3K inhibitor. In addition, thrombin-induced production of CCL11 was significantly attenuated by pharmacological inhibition of Akt or MEK which phosphorylates ERK1/2. These results indicate that thrombin is likely to promote expression of CCL11 via PKC/Raf-1/ERK1/2 and PTX-sensitive protease-activated receptors /PI3K/Akt pathways in HAoSMCs. We propose that multiple signaling pathways are involved in change of VSMCs to a secretory phenotype.