• Title/Summary/Keyword: activity staining

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Apoptosis-related Genes Altered in Bovine Cystic Ovary (난소낭종 시 변화되는 세포사멸 관련 유전자)

  • Tak, Hyun-Min;Kim, Gyu-Tae;Kim, Eun-Jin;Mun, Yun-Ja;Choe, Chang-Yong;Son, Dong-Soo;Han, Jae-Hee;Kang, Da-Won
    • Journal of Embryo Transfer
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    • v.24 no.1
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    • pp.57-64
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    • 2009
  • This study was carried out to investigate expression of apoptosis-related differentially expressed gene (DEG) in ovaries of Korean cattle with follicular and luteal cysts and to identify the relationship between cyst and apoptosis using microarray, real-time PCR, TUNEL staining, and Western blot analysis. Microarray data showed that PIK3R2 and AKT1 were significantly up-regulated in follicular cyst, and TNF-RAF2, PRLR, FOXL2, STK4, and COL4A3 were up-regulated whereas INHA, CIDEB, BCL10, and FASLG were down-regulated in luteal cyst. Real-time PCR was performed to validate DEGs altered in luteal cyst. Of nine DEGs, four DEGs down-regulated in luteal cyst showed a positive corelation between microarray data and real-time PCR data. In this study, we focused on INHA, among many DEGs, which was highly down-regulated in both follicular and luteal cysts. Real-time PCR and micro array data showed that INHA was down-regulated by 12.3-fold and by 1.4-fold, respectively, in the bovine follicular cyst. TUNEL assay and Western blot analysis for ERK, JNK, p38, PI3K, and Akt, which were used to detect whether apoptosis is occurred, showed no significant changes in cystic ovaries (p>0.05). In the expression and activity of caspase-3, Bax, Bel-2, and Bel-xL, there was no significant changes between follicular cystic ovary and normal ovary. Rather, the expression levels of PI3K and p-Akt were decreased in follicular cystic ovary. These results suggest that deficiency of apoptosis in cystic ovary is associated with decreased expression of apoptotic effectors.

4-Hydroxynonenal Induces Endothelial Cell Apoptosis via ROS and Peroxynitrite Generation (4-Hydroxynonenal에 생성된 ROS와 peroxynitrite를 통한 내피세포의 세포사에 관한 연구)

  • Chung, Sang-Woon;Yee, Su-Bog;Lee, Ji-Young;Hossain, Mohammad Akbar;Kim, Dong-Hwan;Yoon, Jeong-Hyun;Chung, Hae-Young;Kim, Nam-Deuk;Kim, Nam-Deuk
    • Journal of Life Science
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    • v.21 no.7
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    • pp.961-968
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    • 2011
  • The formation of reactive lipid aldehydes, 4-hydroxynonenal (HNE) is shown to be derived from fatty acid hydroperoxides through the oxidative process. Among its known effects in cytotoxicity, HNE has been implicated in apoptotic cell death. To delineate its putative role as a potential mediator, we investigated the mechanism by which HNE induces apoptosis of endothelial cells (ECs). The anti-proliferative effects of HNE were tested through MTT assay after exposure to various concentrations ($5\sim15\;{\mu}M$) of HNE. We observed apoptotic bodies with propidium iodide staining, and measured the HNE induction of endothelial apoptosis by flow cytometry assay. We observed that cells exposed to HNE for 24 hr resulted in increased poly(ADP-ribose) polymerase cleavage and up-regulation of Bax. Data on the HNE action strongly indicated the involvement of reactive species, namely, intracellular ROS, nitrite, and peroxynitrite. To obtain evidence on the implication of ROS and peroxynitrite in HNE-induced apoptosis, a ROS scavenger, N-acetylcysteine (NAC), and a peroxynitrite scavenger, penicillamine, were tested. Results clearly indicate that the induction of apoptosis by HNE was effectively inhibited by NAC and penicillamine. Based on the present data, we conclude that the endothelial apoptosis induced by HNE involves both ROS generation and peroxynitrite activity. Our new data could lead to a redefinition of HNE action on apoptosis in ECs.

Apoptosis Induction of Human Breast Carcinoma Cells by Ethyl Alcohol Extract of Hizikia fusiforme (Apoptosis 유도에 의한 톳 ethyl alcohol 추출물의 인체 유방암세포 증식 억제)

  • Jung, Sun-Hwa;Hwang, Won-Deuk;Nam, Taek-Jeong;Choi, Yung-Hyun
    • Journal of Life Science
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    • v.19 no.11
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    • pp.1581-1590
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    • 2009
  • Hizikia fusiforme is a kind of brown edible seaweed that mainly grows in the temperate seaside areas of the northwest pacific, including Korea, Japan and China, and has been widely used as a health food for hundreds of years. Recently, H. fusiforme has been known to exert pharmacological activities including antioxidant, antimutagenic and anticoagulant activities. However, the molecular mechanisms of H. fusiforme in malignant cells have not been clearly elucidated yet. In this study, the effects of ethyl alcohol extract of H. fusiforme (EAHF) on the anti-proliferative effects of MDA-MB-231 and MCF-7 human breast cancer cells were investigated. EAHF treatment resulted in a concentration-dependent growth inhibition by including apoptosis in MDA-MB-231 cells and G1 phase arrest in MCF-7 cells, which could be proved by MTT assay, DAPI staining, agarose gel electrophoresis and flow cytometry analysis. In MDA-MB-231 cells, the increase in apoptosis induced by EAHF treatment correlated with up-regulation of pro-apoptotic Bax expression. EAHF treatment induced the proteolytic activation of caspase-3 and caspase-9, and a concomitant inhibition of poly (ADP-ribose) polymerase, $\beta$-catenin, phospholipase-${\gamma}1$ protein and DNA fragmentation factor 45/inhibitor of caspase-activated DNase. Taken together, these findings provide important new insights into the possible molecular mechanisms of the anti-cancer activity of H. fusiforme.

The Effect of IFN-$\gamma$ on the Phagocytosis of Mycobacterium tuberculosis and Activation of Human Pulmonary Alveolar Macrophage (Interferon-$\gamma$가 사람 폐포대식세포의 결핵균 탐식과 활성화에 미치는 영향)

  • Park, Jae-Seuk;Kim, Jae-Yeal;Lee, Gwi-Lae;Yoo, Chul-Gyu;Kim, Young-Whan;Han, Sung-Koo;Shim, Young-Soo
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.1
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    • pp.36-44
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    • 1998
  • Background: IFN-$\gamma$ is known to activate mononuclear phagocytes and to mediate host defense mechanism against some intracellular microorganisms, but little is known about anti-mycobacterial activity and mechanism of IFN-$\gamma$ in human. In this study, we investigated the role of IFN-$\gamma$ in the pathogenesis of tuberculosis by observing the effect of IFN-$\gamma$ on the phagocytosis of M.tuberculosis(MTB) and on the production of TNF-$\alpha$ by human pulmonary alveolar macrophage. Method: Pulmonary alveolar macrophage(PAM) were prepared with adhesion purification method from bronchoalveolar lavage fluid obtained from 8 persorn without active lung lesion and cultured($1{\times}10^6cells/ml$) with MTB($3{\times}10^7$ bacteria/ml) with or without IFN-$\gamma$(300U/ml), LPS(0.5ug/ml) and autologous serum(10%). After 2 hours, the percentage of PAM-phagocytosed MTB was counted after AFB staining(modified Kynion method). TNF-$\alpha$ production by PAM stimulated by IFN-$\gamma$(300U/ml), MTB($1{\times}10^6bacteria/ml$) and LPS(0.5ug/ml) for 24hours was measured in culture supernatant using ELISA method. The degree of phagocytosis of MTB by PAM stimulated with IFN-$\gamma$(300U/ml) and LPS(0.5ug/ml) for 24hours was also investigated. Results: IFN-$\gamma$ did not influence the phagocytosis of MTB by PAM(percentage of PAM-phagocytosed MTB: control: $22.1{\pm}4.9$, IFN-$\gamma$: $20.3{\pm}5.3$) and did not increase TNF-$\alpha$ production by PAM (control: $21{\pm}38pg/ml$, IFN-$\gamma$: $87{\pm}106pg/ml$), and the degree of phagocytosis of MTB by PAM pre-stimulated with IFN-$\gamma$ for 24 hours, was not increased (control: $24.5{\pm}9.5$, IFN-$\gamma$: $23.4{\pm}10.1$). Conclusion: IFN-$\gamma$ does not influence on the phagocytosis of MTB and TNF-$\alpha$ production by PAM.

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The Anti-diabetic Effects of Kocat-D1 on Streptozotocin-Induced Diabetic Rats (Kocat-D1의 streptozotocin으로 유도한 당뇨모델에 대한 항당뇨 활성)

  • Won, Hye-Jin;Lee, Hyun-Sun;Kim, Jong-Tak;Hong, Chung-Oui;Koo, Yun-Chang;Lee, Kwang-Won
    • Korean Journal of Food Science and Technology
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    • v.42 no.2
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    • pp.204-209
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    • 2010
  • This study was conducted to investigate the anti-diabetic activity of Kocat-D1, which is widely used in traditional medicine to treat diabetes in Shandong, China. Sprague Dawley rats (8 weeks of age) were separated into 4 groups: a normal control, streptozotocin (STZ)-induced diabetic rat group (DM control), Kocat-D1-1 (diabetic rat treated with 0.25 g/kg/day hot water extract), and Kocat-D1-2 (diabetic rat treated with 1 g/kg/day hot water extract). After eight weeks of treatment, the fasting blood glucose levels of the Kocat-D1-1 ($334.3{\pm}32.9\;mg/dL$) and Kocat-D1-2 group ($259.5{\pm}35.0\;mg/dL$) were significantly lower when compared to the DM control group ($451{\pm}42.6\;mg/dL$). Furthermore, the levels of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), albumin and high-density lipoprotein (HDL) cholesterol in the serum of the Kocat-D1-2 group were significantly normalized when compared to the DM control group. However, significant differences were not observed between the Kocat-D1-1 group and the DM control group. Histochemical staining of the liver of the Kocat-D1-2 group revealed no fat accumulation. The insulin level was significantly upregulated in the Kocat-D1-2 group ($0.13{\pm}0.02\;ng/mL$) when compared to the DM control group ($0.05{\pm}0.04\;ng/mL$). The relative volume of $\beta$-cells in the pancreas of the Kocat-D1-2 group ($49.4{\pm}4.2%$) also increased significantly when compared to the DM control group ($12.9{\pm}7.9%$). These results suggest that Kocat-D1 exerts an anti-hyperglycemic effect through the enhancement of insulin secretion.

Leaves of Cudrania tricuspidata on the Shoot Positional Sequence Show Different Inhibition of Adipogenesis Activity in 3T3-L1 Cells (꾸지뽕 신초 엽위별 잎 추출물의 항비만 효과)

  • Park, Ju Ha;Guo, Lu;Kang, He Mi;Son, Beung Gu;Kang, Jum Soon;Lee, Yong Jae;Park, Young Hoon;Je, Byoung Il;Choi, Young Whan
    • Journal of Life Science
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    • v.31 no.2
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    • pp.209-218
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    • 2021
  • This study aimed to evaluate the anti-obesity effects of Cudrania tricuspidata leaf extract in the order of leaf development on the shoot (L0, L1, L2, L3, L4, and L5). The leaves at the apex of a Cudrania tricuspidata shoot were classified as L0; the next leaves of the apex were classified as L1, L2, L3, and L4 from highest to lowest; and the lowest leaf was classified as L5. A series of 70% ethyl alcohol leaf extracts were screened for the inhibitory effects of adipogenesis in 3T3-L1 preadipocytes. We found that the apical leaf extract of Cudrania tricuspidata (CTL0) was the most effective. Next, a study was conducted on the inhibitory action mechanism of CTL0. Treatment with CTL0 significantly suppressed the differentiation of 3T3-L1 preadipocytes in a dose-dependent manner, as confirmed by the decrease in lipid droplet content observed with Oil Red O staining. Treatment with 12.5 ㎍/ml, 25 ㎍/ml, and 50 ㎍/ml of CTL0 significantly reduced the lipid droplet content. Glucose and cellular triglyceride concentrations were reduced in the 3T3-L1 cells on the CTL0-treated medium compared to the differentiation medium (DM control, DMEM + insulin + dexamethasone + rosiglitazone). Compared with DM, CTL0 significantly inhibited the expression of key pro-adipogenic transcription factors, including peroxisome proliferator-activated receptor γ (PPARγ), LPL, A-FABP, and Glut4. These findings show that CTL0 extract has potent anti-obesity effects.

Germinated Rhynchosia nulubilis Hydrolysate Ameliorates Dexamethasone-induced Muscle Atrophy by Downregulating MAFbx Expression in C2C12 Cells and C57BL/6 Mice (발아 서목태 가수분해물의 근위축 억제 효과)

  • Won Keong Lee;Eun Ji Kim;Sang Gon Kim;Young Min Goo;Young Sook Kil;Seung Mi Sin;Min Ju Ahn;Min Cheol Kang;Young-Sool Hah
    • Journal of Life Science
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    • v.33 no.3
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    • pp.277-286
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    • 2023
  • Sarcopenia is the age-related loss of muscle mass and function. It is a natural part of aging and can lead to decreased mobility and increased frailty. The ubiquitin-proteasome pathway, which is involved in muscle protein degradation, is closely linked to sarcopenia. Germinated Rhynchosia nulubilis hydrolysate (GRH) has been reported to have anti-inflammatory and antioxidant properties, but there have been no reports on its inhibitory effect on muscle reduction. However, no study has yet explored the relationship between GRH and muscle loss inhibition. In this study, we evaluated the effects of GRH on muscle atrophy inhibitory activity in dexamethasone (Dexa)-induced muscle atrophy C2C12 myotubes and mouse models. Moreover, we identified a molecular pathway underlying the effects of GRH on skeletal muscle. May Grunwald-Giemsa staining showed that the length and area of myotubes increased in the groups treated with GRH. In addition, the GRH-treated group significantly reduced the expression of muscle ring finger protein 1 and muscular atrophy F-box (MAFbx) in the Dexa-induced muscular atrophy C2C12 model. GRH also improved muscle strength in C57BL/6 mice with Dexa-induced muscle atrophy, resulting in prolonged running exhaustive time and increased grip strength. We found that muscle strengthening by GRH was correlated with a decreased expression of the MAFbx gene in mouse muscle tissue. In conclusion, GRH can attenuate Dexa-induced muscle atrophy by inhibiting the ubiquitin-proteasome pathway via downregulation of the MAFbx gene expression.

Effect of Ginseng on Visceral Nucleic Acid Content of Rats (고려인삼이 흰쥐의 장기조직 핵산 함유량에 미치는 영향)

  • Kim, Chul;Choi, Hyun;Kim, Chung-Chin;Kim, Jong-Kyu;Kim, Myung-Suk;Huh, Man-Kyung
    • The Korean Journal of Physiology
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    • v.5 no.1
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    • pp.23-42
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    • 1971
  • I. Chemical analysis A study was planned to see if administration of ginseng extract has any influence upon the adrenal, the hepatic, the splenic, and the pancreatic nucleic acid contents of rats, and to estimate the effect of ACTH administration as a substitute for stress reaction upon these nucleic acid contents of rats previously primed with ginseng. Ninety male rats$(body\;weight:\;150{\sim}200gm)$ were divided into the ginseng, the saline, and the normal control groups, which received for 5 days 0.5ml/100 gm body weight of ginseng extract solution (4 mg of ginseng alcohol extract in 1 ml of saline), same amount of saline, or no medication, respectively. On the 5th experimental day, each of the 3 groups was further divided into 2 subgroups yielding the ginseng, the ginseng-ACTIT, the saline, the saline-ACTH, the normal control, and the normal-ACTH subgroups. The ginseng, the saline, and the normal control subgroups were sacrificed 3 hours after the last medication, while the ginseng-ACTH, the saline·ACTH, and the normal-ACTH subgroups received ACTH(0.1 unit/subject) 1 hour after the last medication and were sacrificed after 1 more hour. The adrenal gland, the liver, the spleen and the pancreas of each rat were measured for RNA and DNA contents using the chemical method of Schmidt-Thannhauser-Schneider. Following results were obtained: 1. Adrenal RNA and DNA contents and RNA/DNA ratio were all significantly higher in the ginseng group compared with the values obtained from the normal control and the saline groups. Generally administration of ACTH reduced nucleic acid contents of the viscera examined. However, in the ginseng group the rate of decrease [(value of ginseng-ACTH subgroup-value of ginseng subgroup) x100/value of ginseng subgroup)] in adrenal RNA and DNA contents and in RNA/DNA ratio were more conspicuous than they were in the normal control and the saline groups. 2. Hepatic RNA and DNA contents and RNA/DNA ratio were all significantly less in the ginseng group than in the normal control and the saline groups. After ACTH, the rate of decrease in hepatic RNA, DNA, and RNA/DNA ratio of the ginseng· group was less conspicuous than those of the other 2 groups. 3. With regard to the splenic nucleic acid contents, the RNA and the RNA/DNA values of the ginseng group were higher than those of the normal control group but lower than those of the saline group, while the DNA value of the ginseng group was lower than that of the normal control group but higher than that of the saline group. Following administration of ACTH, the rate of decrease in RNA and DNA contents and in RNA/DNA ratio of the ginseng group was more conspicuous than that of the normal control group but less remarkable than that of the saline group. 4. Pancreatic RNA and DNA contents were notably lower in the ginseng group than in the normal control and the saline groups. However, the RNA/DNA ratio of the ginseng group was higher than that of the normal control and the saline groups.'After ACTH, the rate of decrease in pancreatic RNA and RNA/DNA ratio of the ginseng group was less than that of the normal. control group but more than that of the saline group, while the DNA content was actually increased in the ginseng group though it decreased in the normal control and the saline groups. Although the results are not clear enough for an accurate interpretation, they seem to indicate that ginseng exerts notable influence upon the RNA and DNA contents and the RNA/DNA ratio of the viscera stodied. On the whole the drug tends to increase the RNA and DNA contents and RNA/DNA ratio of the adrenal gland but seems to diminish the values of the other 3 viscera. In the early period following ACTH, ginseng facilitates the fall in RNA and DNA contents and RNA/DNA ratio of the adrenal gland, while it tends to reduce the fall in the values of the other viscera studied. II. Autoradiographic and histochemical analysis It was planned autoradiographically and histochemically to affirm and extend the results obtained in part I with regard to the chemically assessed change in the adrenal, the pancreatic, the hepatic and the splenic DNA and RNA contents under the influence of ginseng and ACTH. Fourty male mice (body weight: $18{\sim}20gm$) and 20 male rats were used. Each animal species was divided into the saline, the ginseng, the saline-ACTH, and the ginseng-ACTH groups according to the administered drugs. In the mice, the adrenal, the pancreatic, the splenic and the hepatic DNA-synthetic activity was assessed autoradiographically after administration of $^3H$-thymidine. In the rats, the RNA content of the above 4 organs was assessed histochemically after staining them with methylgreen pyronine. Following results were obtained: 1. Labeled cells were significantly more numerous in the adrenal cortex, the spleen and the liver of the ginseng group than in those of the saline group, although they were less numerous in the pancreas of the ginseng group than in the pancreas of the saline group. The adrenocortical, the pancreatic, the splenic and the hepatic tissues were stained with methylgreen pyronine more deeply in the ginseng group than in the saline group. 2. The adrenocortical, the pancreatic, the splenic and the hepatic tissues contained labeled cells less numerously in the saline-ACTH and the ginseng-ACTH group than in the saline and the ginseng groups. All these tissues were also stained with methylgreen pyronine less deeply in the saline-ACTH and the ginseng-ACTH groups than in the saline and the ginseng groups. 3. However, the adrenal cortex, the spleen, the pancreas, and the liver contained labeled cells more numerously in the ginseng-ACTH group than in the saline-ACTH group. the 4 tissues were stained with methylgreen pyronine more deeply in the ginseng-ACTH group than in the saline-ACTH group. It is inferred from the above results that though with exception, the ginseng mostly facilitates cellular synthesis of nucleic acids and mitigates reduction in nucleic acid content of tissues after administration of ACTH.

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