• Title/Summary/Keyword: HEK293T cells

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Effects of Excretory/Secretory Products from Clonorchis sinensis and the Carcinogen Dimethylnitrosamine on the Proliferation and Cell Cycle Modulation of Human Epithelial HEK293T Cells

  • Kim, Eun-Min;Kim, June-Sung;Choi, Min-Ho;Hong, Sung-Tae;Bae, Young-Mee
    • Parasites, Hosts and Diseases
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    • v.46 no.3
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    • pp.127-132
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    • 2008
  • Clonorchis sinensis is one of the most prevalent parasitic helminths in Korea. Although cholangiocarcinoma can be induced by C. sinensis infection, the underlying mechanism is not clearly understood. To assess the role of C. sinensis infection in carcinogenesis, an in vitro system was established using the human epithelial cell line HEK293T. In cells exposed to the excretory/secretory products (ESP) of C. sinensis and the carcinogen dimethylnitrosamine (DMN), cellular proliferation and the proportion of cells in the G2/M phase increased. Moreover, the expression of the cell cycle proteins E2F1, p-pRb, and cyclin B was dramatically increased when ESP and DMN were added together. Similarly, the transcription factor E2F1 showed its highest level of activity when ESP and DMN were added simultaneously. These findings indicate that DMN and ESP synergistically affect the regulation of cell cycle-related proteins. Our results suggest that exposure to C. sinensis and a small amount of a carcinogen such as DMN can promote carcinogenesis in the bile duct epithelium via uncontrolled cellular proliferation and the upregulation of cell cycle-related proteins.

Knock-down of human MutY homolog (hMYH) decreases phosphorylation of checkpoint kinase 1 (Chk1) induced by hydroxyurea and UV treatment

  • Hahm, Soo-Hyun;Park, Jong-Hwa;Ko, Sung-Il;Lee, You-Ri;Chung, In-Sik;Chung, Ji-Hyung;Kang, Lin-Woo;Han, Ye-Sun
    • BMB Reports
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    • v.44 no.5
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    • pp.352-357
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    • 2011
  • The effect of human MutY homolog (hMYH) on the activation of checkpoint proteins in response to hydroxyurea (HU) and ultraviolet (UV) treatment was investigated in hMYH-disrupted HEK293 cells. hMYH-disrupted cells decreased the phosphorylation of Chk1 upon HU or UV treatment and increased the phosphorylation of Cdk2 and the amount of Cdc25A, but not Cdc25C. In siMYH-transfected cells, the increased rate of phosphorylated Chk1 upon HU or UV treatment was lower than that in siGFP-transfected cells, meaning that hMYH was involved in the activation mechanism of Chk1 upon DNA damage. The phosphorylation of ataxia telangiectasia and Rad3-related protein (ATR) upon HU or UV treatment was decreased in hMYH-disrupted HEK293 and HaCaT cells. Co-immunoprecipitation experiments showed that hMYH was immunoprecipitated by anti-ATR. These results suggest that hMYH may interact with ATR and function as a mediator of Chk1 phosphorylation in response to DNA damage.

Comparison of Immuno-Modulatory Regulatory Activities of Rubus coreanus Miquel by Ultra High Pressure Extracts Process (초고압 공정에 의한 복분자의 면역조절효능 비교)

  • Kwon, Min-Chul;Kim, Cheol-Hee;Na, Chun-Soo;Kwak, Hyeong-Geun;Kim, Jin-Chul;Lee, Hyeon-Yong
    • Korean Journal of Medicinal Crop Science
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    • v.15 no.6
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    • pp.398-404
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    • 2007
  • This study was performed to investigate the enhancement of anticancer activities and immuno modulatary activities from R. coreanus. by ultra high pressure extracts process. The cytotoxicity on human kidney cell (HEK293) was showed below 19.5% in adding 1.0 $mg/m{\ell}$ concentration. The anticancer activity was increased over 10% by high pressure processing in AGS and A549 cells. The immune cell growth using human immune B and T cells was improved by the high pressure extracts of Rubus coreanus in adding 1.0 $mg/m{\ell}$ concentration. The secretion of two kinds of cytokine, the IL-6 and $TNF-{\alpha}$ from human immune B and T cells were also enhanced in adding extracts by high pressure process of R. coreanus. The ultra high pressure extraction technique showed high efficiency in extracting of bioactive compound. The ultra high pressure technique could be used combined with other technique to improve the extracting rate and extracting efficiency.

Role of Tumor Necrosis Factor-Producing Mesenchymal Stem Cells on Apoptosis of Chronic B-lymphocytic Tumor Cells Resistant to Fludarabine-based Chemotherapy

  • Valizadeh, Armita;Ahmadzadeh, Ahmad;Saki, Ghasem;Khodadadi, Ali;Teimoori, Ali
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.18
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    • pp.8533-8539
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    • 2016
  • Background: B-cell chronic lymphocytic leukemia B (B-CLL), the most common type of leukemia, may be caused by apoptosis deficiency in the body. Adipose tissue-derived mesenchymal stem cells (AD-MSCs) as providers of pro-apoptotic molecules such as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), can be considered as an effective anti-cancer therapy candidate. Therefore, in this study we assessed the role of tumor necrosis factor-producing mesenchymal stem cells oin apoptosis of B-CLL cells resistant to fludarabine-based chemotherapy. Materials and Methods: In this study, after isolation and culture of AD-MSCs, a lentiviral LeGO-iG2-TRAIL-GFP vector containing a gene producing the ligand pro-apoptotic with plasmid PsPAX2 and PMDG2 virus were transfected into cell-lines to generate T293HEK. Then, T293HEK cell supernatant containing the virus produced after 48 and 72 hours was collected, and these viruses were transduced to reprogram AD-MSCs. Apoptosis rates were separately studied in four groups: group 1, AD-MSCs-TRAIL; group 2, AD-MSCs-GFP; group 3, AD-MSCs; and group 4, CLL. Results: Observed apoptosis rates were: group 1, $42{\pm}1.04%$; group 2, $21{\pm}0.57%$; group 3, $19{\pm}2.6%$; and group 4, % $0.01{\pm}0.01$. The highest rate of apoptosis thus occurred ingroup 1 (transduced TRAIL encoding vector). In this group, the average medium-soluble TRAIL was 72.7pg/m and flow cytometry analysis showed a pro-apoptosis rate of $63{\pm}1.6%$, which was again higher than in other groups. Conclusions: In this study we have shown that tumor necrosis factor (TNF) secreted by AD-MSCs may play an effective role in inducing B-CLL cell apoptosis.

Luteolin reduces fluid hypersecretion by inhibiting TMEM16A in interleukin-4 treated Calu-3 airway epithelial cells

  • Kim, Hyun Jong;Woo, JooHan;Nam, Yu-Ran;Seo, Yohan;Namkung, Wan;Nam, Joo Hyun;Kim, Woo Kyung
    • The Korean Journal of Physiology and Pharmacology
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    • v.24 no.4
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    • pp.329-338
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    • 2020
  • Rhinorrhea in allergic rhinitis (AR) is characterized by the secretion of electrolytes in the nasal discharge. The secretion of Cl- and HCO3- is mainly regulated by cystic fibrosis transmembrane conductance regulator (CFTR) or via the calcium-activated Cl- channel anoctamin-1 (ANO1) in nasal gland serous cells. Interleukin-4 (IL-4), which is crucial in the development of allergic inflammation, increases the expression and activity of ANO1 by stimulating histamine receptors. In this study, we investigated ANO1 as a potential therapeutic target for rhinorrhea in AR using an ANO1 inhibitor derived from a natural herb. Ethanolic extracts (30%) of Spirodela polyrhiza (SPEtOH) and its five major flavonoids constituents were prepared. To elucidate whether the activity of human ANO1 (hANO1) was modulated by SPEtOH and its chemical constituents, a patch clamp experiment was performed in hANO1-HEK293T cells. Luteolin, one of the major chemical constituents in SPEtOH, significantly inhibited hANO1 activity in hANO1-HEK293T cells. Further, SPEtOH and luteolin specifically inhibited the calcium-activated chloride current, but not CFTR current in human airway epithelial Calu-3 cells. Calu-3 cells were cultured to confluency on transwell inserts in the presence of IL-4 to measure the electrolyte transport by Ussing chamber. Luteolin also significantly inhibited the ATP-induced increase in electrolyte transport, which was increased in IL-4 sensitized Calu-3 cells. Our findings indicate that SPEtOH and luteolin may be suitable candidates for the prevention and treatment of allergic rhinitis. SPEtOH- and luteolin-mediated ANO1 regulation provides a basis for the development of novel approaches for the treatment of allergic rhinitis-induced rhinorrhea.

The Improvement of skin barrier function and anti-obesity effect of Codonopsis lanceolata by supercritical carbon dioxide extraction

  • Kim, Bora
    • Journal of the Korean Applied Science and Technology
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    • v.37 no.4
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    • pp.674-681
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    • 2020
  • The root of Codonopsis lanceolata has been used in traditional medicine. This study was conducted to confirm the comparative effect of ethanol solvent extraction (CLE) and supercritical carbon dioxide extraction (CLS) of C. lanceolata roots. CLS had higher antioxidant than CLE. For supercritical co-solvent modified carbon dioxide extraction (CLS), it were extracted at 250 bar 50℃ 150 min at a flow rate of ethyl alcohol 3 mL/min for 90min. In addition, CLS inhibited the adipocyte differentiation of 3T3-L1 cells. When treated with the extract at a concentration of 100 ㎍/mL, the Wnt/β-catenin pathway reporter luciferase activity of HEK 293-TOP cells increased approximately by 3-folds compared to that of the untreated control group. Also, the treatment by CLS (50 ㎍/mL) showed a significant increase of involucrin expression. These results indicate that supercritical carbon dioxide extract of C. lanceolatamay serve as a cosmeceutical agent for improving skin barrier function and the treatment of obesity.

The activation of PPAR-α and Wnt/β-catenin by Paeonia lactiflora root supercritical carbon dioxide extract

  • Kim, Bora
    • Journal of the Korean Applied Science and Technology
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    • v.36 no.4
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    • pp.1136-1142
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    • 2019
  • The root of Paeonia lactiflora has been used in Chinese medicine. We conducted to check the comparative qualities of ethanol solvent extraction (PLE) and supercritical carbon dioxide extraction (PLS) of P. lactiflora root. PLE had higher antioxidant and polyphenol contents than PLS. But, PLS were significantly increased peroxisome proliferator-activated receptor (PPAR)-α. In addition, PLS inhibited the adipocyte differentiation of 3T3-L1 cells. When treated with the extract at a concentration of 100 ㎍/mL, the Wnt/β-catenin pathway reporter luciferase activity of HEK 293-TOP cells increased approximately by 3-folds compared to that of the untreated control group. These results indicate that P. lactiflora supercritical carbon dioxide extract may serve as a cosmeceutical for improving skin barrier function and the treatment of obesity.

The Activation of PPAR-α and Wnt/β-catenin by Luffa cylindrica Supercritical Carbon Dioxide Extract

  • Kim, Bora
    • Natural Product Sciences
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    • v.25 no.4
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    • pp.341-347
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    • 2019
  • Luffa cylindrica (LC) is a very fast-growing climber and its fruit have been considered as agricultural wastes. We conducted to check the comparative qualities of ethanol solvent extraction (LCE) and supercritical carbon dioxide extraction (LCS) of L. cylindrica fruit and seed. LCS had higher antioxidant and polyphenol contents than LCE. LCS were significantly increased peroxisome proliferator-activated receptor (PPAR)-a and involucrin expression as epidermal differentiation marker in 3D skin equivalent model. LCS also showed antimicrobial activity against Staphylococcus aureus, a causative bacteria in atopic dermatitis. In addition, LCS inhibited the adipocyte differentiation of 3T3-L1 cells. When treated with the extract at a concentration of 100 ㎍/mL, the Wnt/β-catenin pathway reporter luciferase activity of HEK 293-TOP cells was increased approximately by 2-folds compared to that of the untreated control group. These results indicate that L. cylindrica supercritical carbon dioxide extract may serve as a cosmeceutical for improving skin barrier function and the treatment of obesity.

Analysis of the effects of δ-Aminolevulinic acid on the proliferation and apoptosis of mammalian cells (포유류 세포주에서 δ-Aminolevulinic acid (ALA)의 세포증식과 사멸에 미치는 영향분석)

  • Jun, Yong-Woo;Kim, Kun-Hyung;Jo, Su-Yeon;Lee, Jin-A;Jang, Deok-Jin
    • Analytical Science and Technology
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    • v.27 no.5
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    • pp.223-227
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    • 2014
  • ${\delta}$-Aminolevulinic acid (ALA) is a compound which is widely present in the biosphere and plays an important role in the living body as an intermediate of the tetrapyrrole compound biosynthesis pathway that leads to heme in mammals and chlorophyll in plants. ALA is of interest as a biodegradable mediator, a growth regulator, a precursor of heme proteins, and an effective agent used in therapy of cancer. It has been recently reported that ALA is commonly used in dermatology, due to good effects of skin therapy. Although for the last few decades a substantial amount of research has been focused on the elucidation of the mechanism of ALA and the improvement of its therapeutic activity, it's effect on the cell functions and growth was not cleared. Here, we identified that ALA treatment could attenuate cell proliferation of HEK293T and HaCaT cells. In addition, ALA treatement could induce apoptosis of HeLa cells. These results suggest that apoptosis induced by ALA treatment might be responsible for inhibition of cell proliferation. These results propose the possibility of the improved therapeutic strategy making ALA one of the effective drugs used in human cancers.

Extracellular Zinc Modulates Cloned T-type Calcium Channels

  • Lee, Jung-Ha;Park, Byong-Gon;Park, Jin-Yong;Lee, Joong-Woo;Jeong, Seong-Woo
    • Proceedings of the Korean Biophysical Society Conference
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    • 2002.06b
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    • pp.36-36
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    • 2002
  • In the present study, we investigated effects of extracellular zinc (Zn$\^$2+/) on T-type Ca$\^$2+/ channel isoforms (${\alpha}$lG, ${\alpha}$lH, and ${\alpha}$lI) stably expressed in HEK 293 cells. Ca$\^$2+/ currents were measured using 10 mM Ca$\^$2+/ as a charge carrier under whole cell-ruptured patch configuration. Zn$\^$2+/ blocked the ${\alpha}$lH currents with a 100- and 200-fold higher potency (IC$\sub$50/ = 2.5 ${\mu}$M) when compared with those for blockade of the ${\alpha}$1G and ${\alpha}$1I currents, respectively.(omitted)

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