• Title/Summary/Keyword: ELISA assay

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Inhibitory Effect of Ophioglossum vulgatum on Free Radical and MMP Expression in UV-irradiated Human Dermal Fibroblasts (병이소초 추출물의 항산화 및 MMP 발현 저해 효과)

  • Kim, Jin-Hwa;Oh, Jung-Young;Lee, Geun-Soo;Zhang, Yong-He;Pyo, Hyeong-Bae
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.35 no.4
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    • pp.287-292
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    • 2009
  • Human skin is constantly exposed to environmental irritants such as smoke, chemicals and ultraviolet (UV). Free radicals and reactive oxygen species (ROS) caused by these environmental irritants play critical roles in cellular damage. In this study, to investigate the skin cell protective effect of Ophioglossum vulgatum extract, we investigated its effects on intercellular antioxidative activity and UVA-induced MMP expression in human dermal fibroblasts (HDFs). The dried O. vulgatum was extracted in a mixture of ethanol and water (1 : 1) for 24 h at room temperature. The extract was filtered and concentrated in vacuo and lyophilized. For testing intracellular ROS scavenging activity the cultured HDFs were analyzed by increase in DCF fluorescence upon exposure to UVB $20\;mJ/cm^2$. After treatment of O. vulgatum extracts, intracellular ROS levels were measured by luminescence spectrophotometer. Enzyme linked immuno sorbent assay (ELISA), and RT-PCR techniques were used for evaluating the effects of O. vulgatumon on MMP protein and mRNA expression in UVA irradiated HDFs. O. vulgatum extract was found to have ROS scavenging activity with the $IC_{50}$ values of $18.2\;{\mu}g/mL$ against superoxide radicals in the xanthine/xanthine oxidase system. After treatment of O. vulgatum extracts, the oxidation of CM-DCFDA was inhibited effectively and O. vulgatum extracts showed a potent free radical scavenging activity by 30.4 % at $100\;{\mu}g/mL$ in UVB-irradiated HDFs. UVA induced MMP protein expression was reduced 37.7 % by treatment with O. vulgatum extract, and MMP-1 mRNA expression was reduced in a dose-dependent manner. Taken together, these results suggest that O. vulgatum extract prevents the skin cell damage induced by UV irradiation, and implies that O. vulgatum extract may be useful as a new ingredient for anti-aging cosmetics.

The current status and control measures of BSE in the worldwide (국내, 외 광우병의 발생 현황과 대응 방안)

  • Yoo, Han-Sang
    • 한국환경농학회:학술대회논문집
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    • 2009.07a
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    • pp.273-282
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    • 2009
  • The transmissible spongiform encephalopathies (TSEs) disease group are fatal neurodegenerative disorders affecting a wide range of hosts. The group includes kuru and Creutzfeldt-Jakob disease (CJD) in humans, scrapie in sheep and goats and Bovine spongiform encephalopathy (BSE) in cattle. The exact nature of the infectious agent involved in the transmission of these diseases remains controversial. However, a central event in their pathogenesis is the accumulation in infected tissues of an abnormal form of a host-encoded protein, the prion protein (PrP). Whereas the normal cellular protein is fully sensitive to protease ($PrP^{sen}$), the disease-associated prion protein ($PrP^d$) is only partly degraded ($PrP^{res}$), its amino-terminal end being removed. BSE was first reported in the mid-80s in the UK. Ten years later, a new form of human prion disease, variant CJD (vCJD) developed in the wake of the BSE epidemic, and there is now strong scientific evidence that vCJD was initiated by the exposure of humans to BSE-infected tissues, thus indicating a zoonotic disease. However, the ban on the feeding of animal-derived proteins to ruminants, and the apparent lack of vertical transmission of BSE, have led to a decline in the incidence of the disease within cattle herd and therefore, an assumed decreased risk for human contacting vCJD. The origin of the original case(s) of BSE still remains an enigma even though three hypotheses have been raised. Hypotheses are i) sheep- or goat-derived scrapie-infected tissues included in meat and bone meal fed to cattle, ii) a previously undetected sporadic or genetic bovine TSE contaminating cattle feed or iii) originating from a human TSE through animal feed contaminated with human remains. A host cellular membrane protein ($PrP^C$), which is abundant in central nervous system tissue, appear to be conformationally altered in the diseased host into a prion protein ($PrP^{Sc}$). This $PrP^{Sc}$ is detergent insoluble and partially protease-resistant ($PrP^{res}$). The term $PrP^{res}$ is normally used to describe the protein detected after protease treatment, in techniques such as Western immunoblotting, and enzyme-linked immunosorbant assay using fresh/frozen tissue. Immunohistochemistry may performed with formalin-fixed tissues. Also, clinical signs of the BSE are one of the major diagnostic indicators. Recently, atypical forms (known as H- and L-type) of BSE have appeared in several European countries, Japan, Canada and the United States. An unusual case was also reported in a miniature zebu. The atypical BSE fall into two groups based on the relative molecular mass (Mm) of the unglycosylated $PrP^{res}$ band relative to that of classical BSE, one of the higher Mm (H-type) and the other lower (L-type). Both types have been detected worldwide as rare cases in older animals, at a low prevalence consistent with the possibility of sporadic forms of prion diseases in cattle. This raises the unwelcome possibility that vCJD could increase in the human population. Now, active surveillance program against BSE is going on in Korea. In regional veterinary service lab, ELISA is applied to screen the BSE in slaughter and confirmatory tests by Western immunoblotting and immunohistochemisty are carried out if there are positive or suspect in the screening test. Also, the ruminant feed ban is rigorously enforced. Removal of specified risk materials such as brain and spinal cord from cattle is mandatory process at slaughter to prevent the infected material from entering the human food chain.

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Serum Biomarkers for Early Detection of Hepatocellular Carcinoma Associated with HCV Infection in Egyptian Patients

  • Zekri, Abdel-Rahman;Youssef, Amira Salah El-Din;Bakr, Yasser Mabrouk;Gabr, Reham Mohamed;El-Rouby, Mahmoud Nour El-Din;Hammad, Ibtisam;Ahmed, Entsar Abd El-Monaem;Marzouk, Hanan Abd El-Haleem;Nabil, Mohammed Mahmoud;Hamed, Hanan Abd El-Hafez;Aly, Yasser Hamada Ahmed;Zachariah, Khaled S.;Esmat, Gamal
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.3
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    • pp.1281-1287
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    • 2015
  • Background: Early detection of hepatocellular carcinoma using serological markers with better sensitivity and specificity than alpha fetoprotein (AFP) is needed. Aims: The aim of this study was to evaluate the diagnostic value of serum sICAM-1, ${\beta}$-catenin, IL-8, proteasome and sTNFR-II in early detection of HCC. Materials and Methods: Serum levels of IL-8, sICAM-1, sTNFR-II, proteasome and ${\beta}$-catenin were measured by ELISA assay in 479 serum samples from 192 patients with HCC, 96 patients with liver cirrhosis (LC), 96 patients with chronic hepatitis C (CHC) and 95 healthy controls. Results: Serum levels of proteasome, sICAM-1, ${\beta}$-catenin and ${\alpha}FP$ were significantly elevated in HCC group compared to other groups (P-value<0.001), where serum level of IL-8 was significantly elevated in the LC and HCC groups compared to CHC and control groups (P-value <0.001), while no significant difference was noticed in patients with HCC and LC (P-value=0.09). Serum level of sTNFR-II was significantly elevated in patients with LC compared to HCC, CHC and control groups (P-value <0.001); also it was significantly higher in HCC compared to CHC and control groups (P-value <0.001). ROC curve analysis of the studied markers between HCC and other groups revealed that the serum level of proteasome had sensitivity of 75.9% and specificity of 73.4% at a cut-off value of $0.32{\mu}g/ml$ with AUC 0.803 sICAM-1 at cut off value of 778ng/ml, the sensitivity was 75.8% and the specificity was 71.8% with AUC 0.776. ${\beta}$-catenin had sensitivity and specificity of 70% and 68.6% respectively at a cut off value of 8.75ng/ml with an AUC of 0.729. sTNFR-II showed sensitivity of 86.3% and specificity of 51.8% at a cut off value of 6239.5pg/ml with an AUC of 0.722. IL-8 had sensitivity of 70.4% and specificity of 52.3% at a cut off value of 51.5pg/ml with AUC 0.631. Conclusions: Our data supported the role of proteasome, sICAM-1, sTNFR-II and ${\beta}$-catenin in early detection of HCC. Also, using this panel of serological markers in combination with ${\alpha}FP$ may offer improved diagnostic performance over ${\alpha}FP$ alone in the early detection of HCC.

Fatty acid analysis and regulatory effects of citron (Citrus junos Sieb. ex TANAKA) seed oil on nitric oxide production, lipid accumulation, and leptin secretion (유자씨유의 지방산분석 및 Nitric Oxide 생성, 지방축적능, 렙틴분비 조절효과)

  • Kim, Tae Woo;Kim, Kyoung Kon;Kang, Yun Hwan;Kim, Dae Jung;Choe, Myeon
    • Journal of Nutrition and Health
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    • v.47 no.4
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    • pp.221-228
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    • 2014
  • Purpose: Citron seed oil (CSO) has been reported to have high antioxidant activity. However, the composition and other biologically activities of CSO have not been reported. In this study, we confirmed the fatty acid composition of CSO, which may be beneficial to vascular disease and obesity. Methods: We investigated the oil composition of CSO using gas chromatography coupled with mass spectrometry (GC-MS) analysis, and cytotoxicity was confirmed by Cell Counting Kit-8 (CCK-8) assay. Nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs) was measured using Griess reagent, and lipid accumulation and leptin secretion in 3T3-L1 cells were measured by Oil-Red O staining and commercial ELISA kit, respectively. Results: GC-MS analysis indicated that CSO contains several components, including linoleic acid, oleic acid, palmitic acid, stearic acid, linolenic acid, palmitoleic acid, and arachidic acid. In physiological activity analysis, CSO did not induce cytotoxic effects in HUVECs and 3T3-L1 cells. Further, CSO significantly induced nitric oxide and leptin secretion as well as inhibited lipid accumulation. Conclusion: CSO increased NO release, inhibited lipid accumulation, and induced leptin secretion, suggesting it may be useful for the management of vessels and weight gain. Although further studies are required to investigate the safety and mechanism of action of CSO, our results show that the composition and physiological activity of CSO are sufficient for its use as functional edible oil.

IL-12 Regulates B7-H1 Expression in Ovarian Cancer-associated Macrophages by Effects on NF-κB Signalling

  • Xiong, Hai-Yu;Ma, Ting-Ting;Wu, Bi-Tao;Lin, Yan;Tu, Zhi-Guang
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.14
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    • pp.5767-5772
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    • 2014
  • Background and Aim: B7-H1, a co-inhibitory molecule of the B7 family, is found aberrantly expressed in ovarian cancer cells and infiltrating macrophage/dendritic-like cells, and plays a critical role in immune evasion by ovarian cancer. IL-12, an inducer of Th1 cell development, exerts immunomodulatory effects on ovarian cancer. However, whether IL-12 regulates B7-H1 expression in human ovarian cancer associated-macrophages has not been clarified. Therefore, we investigated the effects of IL-12 on the expression of B7-H1 in ovarian cancer-associated macrophages and possible mechanisms. Methods: PMA induced THP-1-derived macrophages or human monocyte-derived macrophages were treated with recombinant IL-12 (rIL-12) or infected with adenovirus carrying human IL-12 gene (Ad-IL-12-GFP) for 24 h, then cocultured with the SKOV3 ovarian cancer cell line for another 24 h. Macrophages were collected for real-time PCR and Western blot to detect the expression of B7-H1, and activation of the NF-${\kappa}B$ signaling pathway. Moreover, supernatants were collected to assay for IL-12, IFN-${\gamma}$ and IL-10 by ELISA. In addition, monocyte-derived macrophages treated with IFN-${\gamma}$ were cocultured with SKOV3 and determined for the expression of B7-H1. Furthermore, the expression of B7-H1 in monocyte-derived macrophages was also evaluated after blocking NF-${\kappa}B$ signaling. Results: The expression of B7-H1 was significantly upregulated in monocyte-derived macrophages treated with rIL-12 or Ad-IL-12-GFP compared with the control groups (p<0.05), accompanied by a remarkable upregulation of IFN-${\gamma}$ (p<0.05), a marked downregulation of IL-10 (p<0.05) and activation of NF-${\kappa}B$ signaling. However, the upregulation of B7-H1 was inhibited by blocking the NF-${\kappa}B$ signaling pathway (p<0.05). Expression of B7-H1 was also increased (p<0.05) in monocyte-derived macrophages treated with IFN-${\gamma}$ and cocultured with SKOV3. By contrast, the expression of B7-H1 in THP-1-derived macrophages was significantly decreased when treated in the same way as monocyte-derived macrophages (p<0.05), and IL-10 was also significantly decreased but IFN-${\gamma}$ was almost absent. Conclusions: IL-12 upregulates the expression of B7-H1 in monocyte-derived macrophages, which is possible though inducing the secretion of IFN-${\gamma}$ and further activating the NF-${\kappa}B$ signal pathway. However, IL-12 downregulates the expression of B7-H1 in THP-1-derived macrophages, associated with a lack of IFN-${\gamma}$ and inhibition of expression of IL-10.

Low Frequency Noise Induces Stress Responses in the Rat (흰쥐에서 저주파소음에 의한 스트레스 반응)

  • Choi, Woong-Ki;Lee, Kyu-Sop;Joung, Hye-Young;Lee, Young-Chang;Sohn, Jin-Hun;Lee, Bae-Hwan;Pyun, Kwang-Ho;Shim, In-Sop
    • Science of Emotion and Sensibility
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    • v.10 no.3
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    • pp.411-418
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    • 2007
  • Exposure to low frequency noise(LFN) can lead to vibroacoustic diseases(VADs), which include a systemic disease with lesions in a broad spectrum of organs and a psychiatric condition. It is known that VAD is an established risk factor for the development of many psychological conditions in humans and rodents, including major depression and anxiety disorder. The present study investigated the effects of LFN on neuronal stress responses in the rat brain. The neuronal expression of the proto-oncogene c-fos in the paraventricular nucleus(PVN) of the hypothalamus and tyrosine hydroxylase(TH) in the LC was observed. The immunocytochemical detection of the Fos protein and TH has been used as a marker of neuronal activation in response to stress. In addition, corticosterone concentration was evaluated by using an enzyme-linked immunosorbent assay(ELISA). The LFN groups were exposed to 32.5Hz and 125Hz of noise(4hr/day for 2days). The numbers of c-fos and TH-immunoreactive cells in the PVN and LC were significantly increased in the LFN groups(32.5Hz and 125Hz) compared to the normal group. Corticosterone concentration in plasma was also increased in LFN groups. The present results demonstrated that exposure with LFN produced a pronounced increase in expression of c-Fos and TH in stress-relevant brain areas. These results suggest that the neural characteristics involved in LFN are similar to those activated by typical processive stressors. These results also suggest that the central and peripheral activations by LFN may be related to LFN-related negative behavioral dysfunctions such as VADs.

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Regulatory Effect of Inflammatory Cytokines Secretion and Hypoxia-inducible $Factor-1{\alpha}$ Activation by Panax ginseng (인삼의 염증성 사이토카인 분비 및 저산소 유도인자-1${\alpha}$ 활성화 조절 효과)

  • Zo, Chul-Won;Lee, Seung-Hee;Kim, Dong-Woung;Lee, Seong-Kyun;Song, Bong-Keun
    • The Journal of Internal Korean Medicine
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    • v.27 no.4
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    • pp.864-878
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    • 2006
  • Purpose : Panax ginseng(PG) is considered to have salutary effects and stimulant actions on physical capacity. However, the effects of PG on the inflammatory cytokine secretion and hypoxia condition are still not understood. This study wasto elucidate the effect of PG on inflammatory cytokine secretion such as interleukin (IL)-1, IL-6, granulocyte macrophage colony stimulating factor (GM-CSF), and tumor necrosis factor $(TNF)-{\alpha}$. Also, the effects on the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1 (HIF-1) were measured. Methods : The water extract of PG was administrated to HMC-1 cells before phorbol myristate acetate (PMA)+A23187 treatment. $IL-1{\beta}$, IL-6, $TNF-{\alpha}$, GM-CSF, and VEGF secretion were measured by a modified enzyme-linked immunosorbent assay (ELISA). HIF-1 activation was measured by transcription factor enzyme-linked immunoassay (TF-EIA) Results : PG significantly decreased secretion of $IL-1{\beta}$, IL-6, $TNF-{\alpha}$, and GM-CSF in PMA+A23187-induced HMC-1 cells. VEGF secretion was not changed but HIF-1 activation was decreased by the treatment of PG. Conclusions : PG inhibited the secretion of inflammatory cytokines, which impliesPG might contribute to treatment of mast cell-mediated inflammatory disease. Also, PG inhibited PMA+A23187-induced $HIF -1{\alpha}activation}$ and DNA-binding activity for HIF-1. Therefore, these data demonstrate that PG modulates inflammatory cytokines through inhibition of $HIF-1{\alpha}activation}$ activation.

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Epstein-Barr Virus Antibodies in Korean Mothers and Their Neonates (산모와 신생아의 Epstein Barr Virus 항체가에 관한 연구)

  • Shin, Young Kyoo;Eun, Baik Lin;Park, Sang Hee;Lim, Chae Seung;Kim, Young Sik
    • Pediatric Infection and Vaccine
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    • v.5 no.1
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    • pp.121-127
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    • 1998
  • Purposes : This study was performed to evaluate the seropositivities and levels of term pregnant women and their neonates, and the transplacental transfer rate of maternal Epstein-Barr Virus-specific IgG(VCA IgG and EBNA IgG) from term pregnant women to their neonates. Subjects and Methods : During Jan. 1, 1997 to Mar. 31. 1997, we collected the 42 pairs of sera from pregnant women and umbilical cord of their neonates in Korea University Ansan Hospital. The serum levels of VCA IgG and EBNA IgG were measured by the ELISA method. Results : 1) The seropositivities of VCA IgG were 100% in mothers and neonates. There was no statistical difference of mean VCA IgG levels between mothers and neonates. There was significant correlation of VCA IgG levels between maternal sera and neonatal umbilical cord sera(correlation coefficient r=0.5214, P<0.001). 2) The seropositivities of EBNA IgG were 100% in mothers and neonates. There was no significant difference of the mean EBNA IgG levels between mothers and neonates. There was significant correlation of EBNA IgG levels between maternal sera and neonatal umbilical cord sera (correlation coefficient r=0.7244, P<0.001). 3) There was no correlation between VCA IgG and EBNA IgG levels of maternal sera. Conclusion : Seropositivities of EBV CA IgG and EBNA IgG of term-pregnant women and their neonates were 100% and no significant differences of antibody levels were found in two groups. It seems that EBV Antibody levels in Korean mothers and neonates were high enough to protect primary EBV infection during early infancy.

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Phellinus linteus Extract Regulates Macrophage Polarization in Human THP-1 Cells (상황버섯 추출물의 인간 유래 THP-1 단핵구 세포주의 분극화 조절)

  • Lee, Sang-Yull;Park, Sul-Gi;Yu, Sun-Nyoung;Kim, Ji-Won;Hwang, You-Lim;Kim, Dong-Seob;Ahn, Soon-Cheol
    • Journal of Life Science
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    • v.30 no.2
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    • pp.113-121
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    • 2020
  • Macrophages are initiators for regulating a host's defenses to eliminate pathogens and trigger tissue repair. Macrophages are classified into two types: classically (M1) activated macrophages and alternatively (M2) activated macrophages. M1-phenotype macrophages directly or indirectly kill infectious organisms and tumor cells via pro-inflammatory responses, whereas M2-phenotype macrophages remodel wounded tissue through anti-inflammatory responses. In this paper, we investigated how Phellinus linteus hot water extract passed from Diaion HP-20 resin (PLEP) regulates polarization of M1-like or M2-like macrophages in human THP-1 cells. PLEP did not have cytotoxicity at a high concentration of 300 ㎍/ml. We observed morphological alteration of the THP-1 cells, which are stimulated by PLEP, LPS/INF-γ (M1 stimulators) or IL-4/IL13 (M2 stimulators). PLEP exposure induced morphology contiguous with LPS/INF-γ. qPCR was also performed to determine whether PLEP influences M1 or M2 polarization-related genes. M1-phenotype macrophage-specific genes, such as TNF-α, IL-1β, IL-6, IL-8, CXCL10 and CCR7, were enhanced by PLEP in a dose-dependent manner similar to LPS/INF-γ. Conversely, M2-phenotype-specific genes, such as MRC-1, DC-SIGN, CCL17 and CCL22, were suppressed by PLEP. PLEP also significantly up-regulated secretory inflammation cytokines related to M1 polarization of macrophages, including TNFα, IL-1β and IL-6, which was similar to the gene expression. Further, MAPK and NF-κB signaling were increased by treatment with PLEP, resulting in enhancement of cytokine secretion. PLEP might therefore be used as a promising booster of pro-inflammatory responses through M1 polarization of human THP-1 cells.

Antimutagenic Effect and Cytotoxicity to Human Cancer Cell Lines of Colored Potato Extracts (유색감자 추출물의 항돌연변이 및 항암 활성)

  • Park, Young-Eun;Jeong, Jin-Cheol;Cho, Huyn-Mook;Hwang, Young-Sun;Lee, Hyeon-Jin;Choi, Su-San-Na;Lee, Su-Jin;Park, Eun-Sun;Ko, Eun-A;Kim, Nan-Sol;Lim, Jung-Dae;Choung, Myoung-Gun
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.53 no.1
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    • pp.75-84
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    • 2008
  • Colored potatoes are an excellent source of dietary polyphenols including anthocyanins. Generally, anthocyanins from fruits and vegetables exhibit anti-carcinogenesis and anti-cancer properties in vitro test. This experiment was conducted to know the effects of colored potato extracts contained anthocyanins on antimutagenic activity and anticancer activity to six human cancer cell lines containing LNCaP (androgen-dependent) prostate cancer cells. Extracts of three colored potatoes ('Hongyoung', 'Jayoung' and 'Jasim') and the white potato ('Superior') cultivars were used in this study. The extracts of three colored potatoes inhibited the mutagenicities induced by direct mutagen such as 4-nitro-quinoline-1-oxide (4-NQO) and another indirect mutagens of bezo(a)pyrene (BaP). Also, the extracts of 'Hoyoung' and 'Jayoung' showed higher antimutagenic activity than 'Jasim' and 'Superior' against to direct or indirect mutagen on both strains of TA98 and TA100. The activity of growth-inhibitory of extract of four potato cultivars were screened by SRB (sulphorhodamine B) method on diverse human cancer cells representing different types of cancers. Among the extract of four potato cultivars, the extract of 'Jasim' showed moderate inhibition on proliferation of LNCaP, ACHN and MOLT-4F cells and did not inhibit the proliferation of other cancer cells. On the other hand, extract of 'Superior' did not inhibit the proliferation of any tested cancer cell lines. However, the extracts of 'Hongyoung and Jayoung' inhibited the proliferation of cancer cells with $GI_{50}$ values ranging from 2.5 to $30\;{\mu}g/mL$. On the basis of the $GI_{50}$ values, it is clear that LNCaP cells were more sensitive to extracts of colored potato cultivars than other cancer cells. The extract of 'Jayoung' at $30\;{\mu}g/mL$ were more active and inhibited cell proliferation, and induced apoptosis in LNCaP cells. This result revealed that the extracts of colored potatoes are expected to be good candidate for development into source of antimutagenic and anticancer agent.