• Title/Summary/Keyword: MTT reduction assay

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Oxidative Stress-Induced Apoptosis in Chronic Myelogenous Leukemia K562 Cells by an Active Compound from the Dithio-Carbamate Family

  • Khoshtabiat, Laya;Mahdavi, Majid;Dehghan, Gholamreza;Rashidi, Mohammad Reza
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.9
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    • pp.4267-4273
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    • 2016
  • Previous studies suggested that dithio-carbamates are potent apoptosis and anti-apoptosis inducing agents in various cancer cells. Here, the anti-proliferative and apoptosis inducing effects of a new derivative (2-NDC) from the dithio-carbamate family was examined in human leukemia K562 cells. We use thiazolyl blue tetrazolium bromide (MTT) to measure viability and cell growth inhibition. The 2-NDC showed effects on viability in a dose and time-dependent manner, inhibiting proliferation at concentrations of $10-30{\mu}M$ after 24-48 hours of treatment and increasing values after 72 hours at $40-120{\mu}M$. The cytotoxic effect of the compound was calculated with an $IC_{50}$ of $30{\mu}M$ after 24-hour. Apoptosis induction was confirmed by acridine orange-ethidium bromide (AO/EtBr) staining, DNA fragmentation assay, flow cytometric assessment and also caspase-3 activation assay. Furthermore, enzymes level such as superoxide dismutase (SOD) and catalase (CAT) involved in oxidative stress were evaluated. The results of this study demonstrated insignificant increase of intracellular ROS levels for 24 hours and reduction after 48-72 hours. In addition to reduction of intracellular thiol, caspase-3 like activity was also decreased in a time-dependent manner in cells treated with 2-NDC. Thus 2-NDC can be considered as a good candidate for further pharmaceutical evaluations.

The Anti-Cancer Effect of β-Gluconsan Calcium on Oral Cancer Cell (β-Gluconsan Calcium이 구강암 세포의 성장에 미치는 영향)

  • Kim, Ji-Hye;Jung, Yun-Sook;Kim, Hye-Young;Lee, Young-kyun;Kim, Jae-Young;Choi, Youn-Hee;Song, Keun-Bae
    • Journal of dental hygiene science
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    • v.15 no.6
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    • pp.794-799
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    • 2015
  • In recent years, there has been a global trend toward the importance of natural extracts for the prevention and treatment of human diseases. ${\beta}$-glucan is known to have anti-inflammatory activity, anti-cancer, and improvement of immune system. Polycan is purified ${\beta}$-glucan from Aureobasidum pullulans SM-2001. The anti-cancer effects of ${\beta}$-gluconsan calcium, polycan and calcium gluconate complex, were evaluated in human oral cancer YD-10B cells. YD-10B cells were cultured in the presence of 0, 0.5, 0.75, 1 mg/ml ${\beta}$-gluconsan calcium for 48 hours. MTT assay, cell counting, and observation of cell morphology were conducted. The number of cells decreased and cell morphology changed in the 0.5 mg/ml of ${\beta}$-gluconsan calcium. Almost all cells were dead in the 0.75 and 1 mg/ml. MTT assay showed a dose-dependent reduction in cell proliferation (p<0.05). These results indicate that ${\beta}$-gluconsan calcium exhibiting anti-cancer effects in YD-10B cells through changes in cell morphology and cell death.

Cytotoxic Effect of Extracts from Styela clava against Human Cancer Cell Lines (암세포주에 대한 미더덕 추출물의 세포독성 효과)

  • Jung, Eun-Sil;Kim, Ju-Young;Park, Eun-Ju;Park, Hae-Ryong;Lee, Seung-Cheol
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.35 no.7
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    • pp.823-827
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    • 2006
  • The present study describes the preliminary evaluation of the cytotoxic effect from Styela clava (Korean name : miduduk) extracts. S. clava was treated with methanol, ethanol, acetone, and water, then cytotoxic effect of the extracts were evaluated by the MTT reduction assay. The ethanol extracts from S. clava showed the cytotoxic activity on the HT-29 human colon cancer cells. The ethanol extracts was further fractionated with n-hexane, diethyl ether, ethyl acetate, and water according to the degree of polarity. The diethyl ether fraction showed high cytotoxic activity of HT-29 cells. however, the other fractions showed low cytotoxicity. The diethyl ether layer also showed the cytotoxic activity against SW620, HeLa, and MCF-7 cells. These studies support that extracts from S. clava may be a potential candidate as a possible chemotherapeutic agent against human cancer cells.

Whitening Effect and Antioxidant Activity of Horseradish Subcritical Water Extracts (겨자무 아임계수 추출물의 항산화활성 및 미백효과)

  • In-Jae Kim;Su-Bhin Eun;Won-Hee Kim;Seon-Bhin Park;Hee-Bin Ku;Gyo-Nam Kim;Seung-Cheol Lee;Youngim Choi;Sanggeun Park;Hae-Ryong Park
    • Journal of Life Science
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    • v.34 no.4
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    • pp.236-244
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    • 2024
  • Melanin is a natural pigment found in most plants and animals, and it is involved in determining the color of the skin and hair. Melanogenesis is a reactive occurrence in melanocytes aiming to protect the skin from external stimuli, such as ultraviolet rays. Tyrosine is involved in the biosynthesis of the substrate tyrosine into melanin. However, melanin overproduction can lead to skin diseases, such as melasma, blotching, hyperpigmentation, and skin cancer. Although many studies have been conducted on whitening substances, such as kojic acid and arbutin, some countries have banned or refrained from using them due to their side effects. Therefore, this study assessed the potential of horseradish (HR) as a new whitening agent in cosmetic products. For efficient extraction, subcritical water extraction was conducted. The results showed that the horseradish subcritical water 200℃ (SW 200) extract showed high DPPH radical scavenging ability, total phenolic contents (TPC), inhibiting tyrosinase activity and inhibiting melanin production of B16-F10 melanoma cell lines. To investigate its cytotoxicity to the B16-F10 melanoma cell lines, MTT reduction assay and morphological changes were observed. No cytotoxicity was found in horseradish methanol extract and SW 200. In conclusion, this research suggests the possibility of horseradish subcritical water may be useful as a natural whitening ingredient to be used in cosmetic products.

Brine shrimp lethality and cytotoxicity assay of Araucaria bidwillii Hook in human carcinoma cell lines

  • Ahamed, KFH Nazeer;Kumar, V;Manikandan, L;Wahile, Atul M;Mukherjee, Kakali;Saha, BP;Mukherjee, Pulok K
    • Advances in Traditional Medicine
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    • v.5 no.1
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    • pp.21-28
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    • 2005
  • The leaf extracts of Araucaria bidwillii Hook. (Araucariaceae) were evaluated for their cytotoxic effect in various human cancer cell lines. Preliminary investigation by brine shrimp lethality assay indicated that $LC_{50}$ value of various successive extracts were found to be less than $1000\;{\mu}g/ml$, where the ethyl acetate extract showed maximum activity of less than $100\;{\mu}g/ml$. Further cytotoxic evaluation of various leaf extracts of Araucaria bidwilli Hook was carried out in four different human cancer cell lines-acute myeloblastic leukemia (HL-60), chronic myelogenic leukemia (K-562), breast adenocarcinoma (MCF-7) and cervical epithelial carcinoma (HeLa). Cytotoxicity was assessed by trypan blue dye exclusion method and 3-(4,5-dimethyl thiazole-2yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction assay. From the present investigation it was found that the ethyl acetate and methanol extract of Araucaria bidwilli Hook was found to be more effective in leukemic cell lines and was less effective in MCF-7 and HeLa. The $IC_{50}$ value of the ethyl acetate extract in leukemic cell lines was found to be $28.18\;and\;34.64\;{\mu}g/ml$ and methanol extract was found to be $33.11\;&\;39.81\;{\mu}g/ml$. It can be concluded that various extract from the leaves of Araucaria bidwillii Hook. posses cytotoxic activity tested in brine shrimps and various human carcinoma cell lines.

Antioxidant Effects of Gamma-oryzanol on Human Prostate Cancer Cells

  • Klongpityapong, Papavadee;Supabphol, Roongtawan;Supabphol, Athikom
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.9
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    • pp.5421-5425
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    • 2013
  • Background: To assess the antioxidant effects of gamma-oryzanol on human prostate cancer cells. Materials and Methods: Cytotoxic activity of gamma-oryzanol on human DU145 and PC3 prostate cancer cells was determined by proliferation assay using 3-(4, 5-dimethylthiazol, 2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) reagent. mRNA levels of genes involved in the intracellular antioxidant system, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and glutathione reductase (GSR) were determined by reverse transcription-polymerase chain reaction (RT-PCR). Cancer cell lysates were used to measure lipid peroxidation using thiobarbituric acid reactive substance (TBARS). Glutathione contents of the cell lysates were estimated by the reaction between sulfhydryl group of 5, 5'-dithio (bis) nitrobenzoic acid (DTNB) to produce a yellow-color of 5-thio-2-nitrobenzoic acid using colorimetric assay. Catalase activity was also analysed by examining peroxidative function. Protein concentration was estimated by Bradford's assay. Results: All concentrations of gamma-oryzanol, 0.1-2.0mg/ml, significantly inhibited cell growth in a dose- and time-dependent fashion in both prostate cancer cell lines, DU145 and PC3. Gene expression of catalase in DU145 and PC3 exposed to gamma-orizanol at 0.5mg/ml for 14 days was down regulated, while mRNA of GPX was also down regulated in PC3. The MDA and glutathione levels including catalase activity in the cell lysates of DU145 and PC3 treated with gamma-oryzanol 0.1 and 0.5mg/ml were generally decreased. Conclusions: This study highlighted effects of gamma-oryzanol via the down-regulation of antioxidant genes, catalase and GPX, not cytotoxic roles. This might be interesting for adjuvant chemotherapy to make prostate cancer cells more sensitive to free radicals. It might be useful for the reduction of cytotoxic agents and cancer chemoprevention.

Effects of Puerariae Radix extract on Cisplatin-Induced Apoptosis of Rat Mesangial Cells (갈근(葛根) 추출물이 cisplatin에 의해 유도된 rat mesangial cell의 apoptosis에 미치는 영향)

  • Hong, Jae-Eui;Shin, Jo-Young;Ju, Sung-Min;Jeon, Byung-Hun;Lee, Si-Hyeong
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.24 no.2
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    • pp.220-227
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    • 2010
  • Puerariae radix (PR) is a popular natural herb and a traditional food in Asia, which has antithrombotic and anti-allergic properties and stimulates estrogenic activity. One of the major side effects of cisplatin is nephrotoxicity, leading to acute renal failure. Recent study has suggested a role of ROS and p53 in renal cell injury by cisplatin. We studied that protective effects of PR on cisplatin-induced apoptosis in rat mesangial cell. Rat mesangial cell was preincubated with PR (50, 100, 150 and 200 ${\mu}g/m{\ell}$) for 12 hr and then treated with 30 ${\mu}M$ cisplatin for 24 hr. Protective effect of PR on cisplatin-induced apoptosis in ECV304 cells was determined using MTT assay, FDA-PI staining, flow cytometric analysis, caspase-3 activity assay, ROS assay and western blot. Our results showed that PR inhibited in cisplatin-induced apoptosis and ROS production in ECV304 cells. Moreover, PR reduced ERK, p38 and JNK activation that increased in cisplatin-treated rat mesangial cell. Furthermore, activation of p53 by cisplatin in rat mesangial cell was inhibited by PR treatment. These results suggest that protective effect of PR on cisplatin-induced apoptosis in rat mesangial cell may be associated with reduction of ERK, p38, JNK, p53 activation.

Rosuvastatin Induces ROS-mediated Apoptosis in Human Prostate Cancer PC-3 Cells (Rosuvastatin이 유도하는 ROS가 전립선암 PC-3 세포주의 세포사멸 유도에 미치는 영향)

  • Choi, Hyeun Deok;Baik, Jong Jin;Kim, Sang Hun;Yu, Sun Nyoung;Chun, Sung Hak;Kim, Young Wook;Nam, Hyo Won;Kim, Kwang Youn;Ahn, Soon Cheol
    • Journal of Life Science
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    • v.26 no.4
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    • pp.398-405
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    • 2016
  • Statins, the inhibitors of 3-hydroxy 3-methylglutaryl coenzyme A (HMG-CoA) reductase, are widely used in treatments of hypercholesterolemia and newly known as anti-cancer effect of various cancer cells. Recently, several studies suggested that reactive oxygen species (ROS) play a critical role on cell death signaling. However, mechanism of ROS by rosuvastatin is currently unclear. This study aimed to explore the molecular mechanism of apoptosis by rosuvastatin in human prostate cancer PC-3 cells. Cell viability and apoptosis-related protein expression were measured by MTT assay and western blotting, respectively. In addition, the levels of apoptosis and ROS were analyzed. The results showed that rosuvastatin dramatically reduced cell viability in a dose- and time-dependent manner. We confirmed that rosuvastatin induced apoptosis through reduction of procaspase-3 and cleavage of poly (ADP-ribose) polymerase (PARP) in PC-3 cells. In addition, rosuvastatin stimulated ROS production in a dose-dependent manner and pre-treatment with N-acetylcysteine (NAC), a ROS scavenger, significantly recovered rosuvastatin-induced ROS and apoptosis. Thus, we concluded that rosuvastain induces apoptosis through generation of ROS in human prostate cancer PC-3 cells and provides a promising approach to improve the efficacy of cancer therapy.

Effects of Ixeris dentata Extract on Radical Oxygen Species and Bcl-2 Family in Human Breast Cancer Cells (씀바귀 추출물이 인체유방암세포의 활성 산소 및 Bcl-2 Family에 미치는 영향)

  • Kim, Hee-Jung;Kang, Keum-Jee
    • Journal of the East Asian Society of Dietary Life
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    • v.24 no.6
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    • pp.739-747
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    • 2014
  • The aim of the study was to determine the effects of Ixeris dentata extract (IDE) on anticancer activity in human breast cancer MDA-MB-231 cells at both cellular and molecular levels. The cells were cultured in the presence of 0, 20, 30 and $40{\mu}g/mL$ Ixeris dentata extract for 24 hours, respectively. At the end of culture, cytochemical analyses for MTT activity, trypan blue dye exclusion, Annexin V-FITC Apoptosis, and radical oxygen species (ROS) were conducted. RT-PCR was also performed to determine whether or not alterations in cell viability affect the Bax/Bcl-2 ratio. MTT assay showed that relative cell viability decreased in a dose-dependent manner (p<0.05). Reduction of cell viability matched well with increased cell membrane permeability as determined by trypan blue dye exclusion test (p<0.05). The rates of intracellular ROS also increased in a similar manner to those of TB-stained cells. There was an associated shift of apoptotic cells from early to late apoptosis between the 30 and $40{\mu}g/mL$. Bax/Bcl-2 ratio significantly increased along with significant decreases in Bcl-2 expression between 30 and $40{\mu}g/mL$ groups (p<0.05). In conclusion, anticancer activity of Ixeris dentata extract is modulated by a reduction in cell viability along with increased membrane permeability, leading to ROS accumulation within cells, and subsequently cell death through an apoptotic pathway that involves Bax and Bcl-2 in human breast cancer MDA-MB-231 cells.

Protective Effect of Perilla frutescens Extract against Oxidative Stress-Induced Cell Death in a Staurosporine-Differentiated Retinal Ganglion Cell Line (Staurosporine에 의해 분화된 망막신경절세포에서 산화 스트레스 유도 세포사멸에 대한 차조기 추출물의 보호 효능)

  • Lee, Bo Kyung;Choe, Lira;Lee, Ji In;Lee, Doo Yi;Chang, Sun-Young;Kim, So Hee;Jung, Yi-Sook
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.46 no.2
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    • pp.161-168
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    • 2017
  • In this study, we examined the effect of Perilla frutescens extract (PFE) on oxidative stress-induced cell death in RGC-5 cell lines. Staurosporine-differentiated RGC-5 (ssdRGC-5) cells obtained by treating RGC-5 cells with $1{\mu}M$ staurosporine were incubated with PFE for 30 min and then exposed to buthionine sulfoximine plus glutamate (B/G) for 20 h. Cell death was detected using lactate dehydrogenase release assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. To investigate the mechanism underlying cell death, we determined caspase-3 activity, level of reactive oxygen species (ROS) formation, and expression levels of cytoplasmic cytochrome c and mitochondrial Bax. Treatment of ssdRGC-5 cells with B/G increased intracellular ROS and induced apoptosis with increasing caspase-3 activity. PFE rescued ssdRGC-5 cells from oxidative stress-induced cell death by inhibiting intracellular ROS production and caspase-3 activation and regulating apoptosis-related proteins such as cytochrome c and Bax. These findings suggest that PFE may have a beneficial neuroprotective effect against oxidative stress-induced apoptotic death in ssdRGC-5 cells.