• 제목/요약/키워드: Western blotting analysis

검색결과 459건 처리시간 0.036초

Induction of Apoptotic Cell Death by Red Pericarp Rice (Jakwangchalbyeo) Extracts

  • Chi, Hee-Youn;Lee, Chang-Ho;Kim, Kwang-Ho;Kim, Sun-Lim;Chung, Ill-Min
    • Food Science and Biotechnology
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    • 제15권4호
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    • pp.534-542
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    • 2006
  • The effects of ethanol fractions of three different rice grain extracts, Jakwangchalbyeo, Hwasunchalbyeo, and Ilpumbyeo, on apoptotic cell death in the rat hepatoma H4IIE cell line were investigated using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] cell viability assay. One hundred mg/mL Jakwangchalbyeo extract significantly reduced cell viability to 69.5, 57.2, and 46.1% within 24, 48, and 72 hr, respectively. Fluorescence-activated cell sorting (FACS) analyses were also performed to characterize the cell death pattern caused by treatment with the rice grain extracts. Apoptotic cell death was clearly observed with time after treatment with the Jakwangchalbyeo extract. In Western blotting analysis, degradation of the 116 kDa poly-ADP-ribose polymerase (PARP) molecule was observed with concomitant formation of an 89 kDa product 24, 48, and 72 hr after treating cells with the Jakwangchalbyeo extract. This indicates that an apoptotic process caused cell death in these cells. In conclusion, red-pericarp Jakwangchalbyeo extract induced apoptotic cell death in H4IIE cells to a larger extent than the other rice extracts.

Apoptotic Signaling Cascade of 5-aminolaevulinic Acid-based Photodynamic Therapy in Human Promyelocytic Leukemia HL-60 Cells

  • Nagao, Tomokazu;Matsuzaki, Kazuki;Takahashi, Miho;Minamitani, Haruyuki
    • Journal of Photoscience
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    • 제9권2호
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    • pp.509-511
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    • 2002
  • In this study, we investigated apoptotic cell death induced by photodynamic therapy using 5-aminolaevulinic acid (ALA-PDT) in human promyelocytic leukemia cells (HL-60). ALA-PDT induced apoptosis in HL-60 cells as confirmed by DNA agarose gel electrophoresis and nuclear staining with Hoechst 33342. The apoptotic cell death was inhibited by addition of broad-spectrum caspase inhibitor Z-Asp-CH$_2$-DCB, indicating that the apoptotic cell death was induced in a caspase-dependent manner. Actually, western blotting analysis revealed that caspase-3 was processed as early as 1.5 h after ALA-PDT. Cytoplasmic cytochrome c released from mitochondria was detected by western blotting. However, inhibitor of caspase-9, a cysteine protease located in the downstream of cytochrome c release, was not able to reduce the apoptotic cell death. Therefore, the mitochondrial apoptotic pathway was not involved in the ALA-PDT-induced apoptosis. On the other hand, it was found that ALA-PDT-induced apoptosis was clearly inhibited by pretreatment of caspase-8 inhibitor. These data suggest that caspase-8-mediated apoptotic pathway is important in ALA-PDT-induced cell death.

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The Relationship between the Expression of Melanoma Differentiation-Associated Gene-7/Interleukin-24 (MDA-7/IL-24) and Clinicopathological Features in Colorectal Adenocarcinomas

  • Seo, Boram;Hong, Young Seob;Youngmin, Youngmin;Roh, Mee Sook
    • 대한의생명과학회지
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    • 제18권4호
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    • pp.413-419
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    • 2012
  • The melanoma differentiation-associated gene-7 (MDA-7) protein, also known as interleukin-24 (IL-24), is a novel candidate of tumor suppressor that can induce apoptosis experimentally in a variety of human malignant cells. However, there have been few studies about its role in colorectal cancer. We performed immunohistochemical detection of MDA-7/IL-24 in 399 tissue samples from primary colorectal adenocarcinoma patients using a tissue microarray. Western blotting was then done to confirm the immunohistochemical observations. MDA-7/IL-24 immunoreactivity was observed in 116 (29.1%) of the 399 colorectal adenocarcinoma cases. Analysis of the MDA-7/IL-24 expression by Western blotting confirmed the immunohistochemical results. The tumors with a negative MDA-7/IL-24 expression more frequently showed poor differentiation (P=0004), lymph node metastasis (P=0.001), deep invasion (P=0.008) and high stage (P=0.001). A subset of colorectal adenocarcinoma revealed a decreased expression of MDA-7/IL-24, and this was associated with progressive pathologic features. These findings suggest that loss of MDA-7/IL-24 expression may play a role in tumor growth and progression of colorectal adenocarcinomas.

기내배양과 재조합 단백질 생산을 위한 화분 Biofactory의 저장기술의 개발 (Storage of Pollen Biofactory for in vitro Growth and rProtein Synthesis)

  • 박희성;고재철
    • KSBB Journal
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    • 제19권3호
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    • pp.174-177
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    • 2004
  • 기내에서의 발아 및 형질전환 활성을 지닐 수 있는 백합 (Lilium longiflorum) 화분의 장기보존조건을 얻기 위하여 용매를 이용한 연구를 수행하였다. 화분의 용이한 수집을 위하여 용매를 사용했을 때, petroleum ether, n-heptane, benzene 등의 경우 화분의 기내발아에는 영향을 미치지는 않았으나 수집과 정 후 바로 화분건조 및 냉동 보관함으로써 발아활성을 유지 할 수 있었다. 이들의 활성유지는 또한 vacuum infiltration 및 Agrobacterium을 이용한 형질전환에 따른 외래유전자인 tissue plasminogen activator의 발현에 의하여 확인하였다.

Biofactory로서의 미나리 엽조직을 이용한 재조합단백질 발현 (Expression of recombinant protein from Oenanthe javanica DC. leaf tissues as a biofactory)

  • 신동일;박희성
    • KSBB Journal
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    • 제23권6호
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    • pp.554-556
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    • 2008
  • 미나리의 엽조직을 agroinfiltration 및 동시배양을 수행하기 전에 NaOH 용액에서 3 min 처리를 실시하였다. MTT를 이용한 세포사멸실험에서 0.7% NaOH 용액처리까지는 엽조직 세포활성에 안전한 것으로 판단되었다. GUS 효소활성의 형광분석 결과 0.5% NaOH를 처리한 미나리 엽조직에 대하여 Agrobacterium cells ($OD_{600}=0.5$에서 1.0)을 이용한 vacuum infiltration (20 min)을 실시할 경우 효율적 형질전환이 이루어짐을 알 수 있었다. 이러한 조건은 western blotting과 ELISA에 의한 HBsAg의 발현 검정에서 확인할 수 있었다.

점액표피양암종 세포주에서 Kochia scoparia 추출물의 세포자멸과 자가포식 유도 효과 (Apoptosis and Autophagy Induced by Methanol Extract of Kochia scoparia in Human Mucoepidermoid Carcinoma Cell Line)

  • 도미향;유미현;김욱규
    • 대한구강악안면병리학회지
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    • 제42권6호
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    • pp.167-174
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    • 2018
  • Natural products are vastly utilized as a source of chemotherapeutic agents for human cancers. Kochia scopraia is traditionally used for the cure of urological and dermatological diseases. Recently, methanol extract of Kochia scoparia (MEKS) has been shown to have anti-cancer activity to various human cancers. However, there is no report demonstrating the anti-cancer activity of MEKS in human mucoepidermoid carcinoma (MEC) cells. In this study, the authors studied the effects of MEKS on the cell proliferation and underlying mechanism in YD15 human MEC cells. MEKS decreased YD15 cell proliferation proven by trypan blue exclusion assay and induced apoptosis, evidenced by cell cycle analysis and western blotting. Autophagy induction by MEKS was verified by western blotting. In addition, MEKS regulated the expression of phosphorylated Akt, phosphorylated p38 and Nrf2 protein. This results can imply that MEKS might be a potential candidate for the treatment of human MEC cells.

Anti-Cancer Effects of the Pandanus tectorius Parkinson Extract: Reduction of YAP and TAZ Levels via Inhibition of the Hippo and Notch Signaling Pathways

  • Min Kyu Kang;Da Hyun Kim
    • 대한의생명과학회지
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    • 제30권3호
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    • pp.113-122
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    • 2024
  • In this study, we aimed to investigate the anti-cancer effects of the Pandanus tectorius extract on AGS cells. P. tectorius, commonly known as hala or screw pine, is a tropical plant traditionally used for its medicinal properties, including anti-inflammatory and antioxidant properties. Here, effects of the P. tectorius extract on cell proliferation, migration, and gene expression were evaluated using various assays, including the water-soluble tetrazolium salt (WST)-1, wound healing, migration, and western blotting assays. WST-1 assay revealed a significant dose- and time-dependent decrease in cell viability, with higher concentrations of the extract resulting in more pronounced viability inhibition. Wound healing and migration assays revealed that the P. tectorius extract effectively hindered cell migration, as the treated cells showed considerably slower wound closure and reduced migration than the control cells. Molecular analysis revealed that the extract significantly downregulated the expression levels of key oncogenic proteins, genes, and components of the Notch signaling pathway. Western blotting confirmed the substantial reduction in the marker protein levels in treated cells. These findings suggest that P. tectorius extract exerts its anti-cancer effects by inhibiting multiple signaling pathways crucial for cancer cell proliferation, migration, and survival. Overall, this study highlights the potential of P. tectorius extract as a therapeutic agent for gastric cancer treatment.

열 및 에탄을 shock이 Vibrio vulnificus의 막단백질에 미치는 영향 (Effects of heat and ethanol shock on the membrane proteins of Vibrio vulnificus)

  • 허문수;정초록
    • 한국어병학회지
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    • 제12권2호
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    • pp.89-99
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    • 1999
  • V. vulnificus ATCC 27562균주의 배양 온도를 $2^{\circ}C $, 20분간 및 6% 에탄올, 10분간으로 반응시켰을 때 SDS-PAGE상에서 새로운 16가지의 heat shock protein(hsps)과 10가지의 ethanol shock protein이 나타났다. Lethal temperature에 노출하기전에 미리 열 충격을 가한 경우 thermo tolerance가 유도되었다. 균체면역에 의해 생성된 항혈청과 열 충격 세포에서 분리된 막단백질과의 ELISA에서는 Outer Membrane Protein(OMP)에서 높은 면역반응을 나타냈으며 western blotting으로는 Inner Membrane Protein(IMP)에서는 62kDa, OMP에서는 69 kDa단백이 높은 면역원성을 나타냈다. ethanol 충격 반응에서는 IMP에서는 48 kDa, OMP에서는 오직 major밴드에서만 면역반응성이 확인되었다. anti-V, vulnificus혈청에 대한 균체 응집시험에서는 열 충격 반응 후의 균체가 정상 균체에 비해 응집반응성이 높았다.

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Analysis of the Apoptotic Mechanisms of Snake Venom Toxin on Inflammation-induced HaCaT Cell-line

  • Chun, Youl Woong;Song, Ho Sueb
    • Journal of Acupuncture Research
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    • 제34권1호
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    • pp.23-30
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    • 2017
  • Objectives : In this study, the roles of Interleukin (IL)-4 and Signal transducer and activator of transcription 6 (STAT6), which have been reported to play a role in the pathogenesis of inflammation and cancer, were evaluated in snake venom toxin (SVT)-induced apoptosis. Methods : Inflammation was induced in human HaCaT kerationocytes, by lipopolysaccharide (LPS; $1{\mu}g/mL$) or tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), followed by treatment with SVT (0, 1, or $2{\mu}g/mL$). Cell viability was assessed by MTT assays after 24 h, and the expression of levels of IL-4, STAT6, and the apoptosis-related proteins p53, Bax, and Bcl-2 were evaluated by western blotting. Electro mobility shift assays (EMSAs) were performed to evaluate the DNA binding capacity of STAT6. Results : MTT assays showed that inflammation-induced growth of HaCaT cells following LPS or TNF-${\alpha}$ stimulation was inhibited by SVT. Western blot analysis showed that p53 and Bax, which promote apoptosis, were increased, whereas that of Bcl-2, an anti-apoptotic protein, was decreased in a concentration-dependent manner in LPS- or TNF-${\alpha}$-induced HaCaT cells following treatment with SVT. Moreover, following treatment of HaCaT cells with LPS, IL-4 concentrations were increased, and treatment with SVT further increased IL-4 expression in a concentration-dependent manner. Western blotting and EMSAs showed that the phosphorylated form of STAT6 was increased in HaCaT cells in the context of LPS- or TNF-${\alpha}$-induced inflammation in a concentration-dependent manner, concomitant with an increase in the DNA binding activity of STAT6. Conclusion : SVT can effectively promote apoptosis in HaCaT cells in the presence of inflammation through a pathway involving IL-4 and STAT6.

삼세기(Shaggy sea raven, Hemitripterus villosus)의 carbonic anhydrase III에 관한 연구 (Presence of Carbonic Anhydrase III-like Protein in Shaggy Sea Raven, Hemitripterus villosus)

  • 권록은;고강희
    • 생명과학회지
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    • 제24권2호
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    • pp.186-190
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    • 2014
  • 본 논문에서는 삼세기(Shaggy sea raven, Hemitripterus villosus)를 실험 재료로 선택하여 Carbonic Anhydrase Isozymes (CAs) 중의 하나인 CA III에 대한 연구를 SDS-PAGE, Isoelectric Focusing (IEF), Western blot analysis의 방법을 통하여 진행하였다. SDS-PAGE와 Western blot 결과 삼세기 아가미, 혈액, 장, 간, 신당, 근육, 심장조직에서 CA III의 분자량인 30 kDa의 band가 확인되었다. 삼세기의 근육과 아가미에 대한 등전점 전기영동(IEF)과 Western blot analysis 결과 pI 7.0 부근에서 band가 형성되는 것을 확인할 수 있었다. 특히 SDS-PAGE와 IEF 실험결과 삼세기의 아가미 조직에서 CA III의 발현량이 다른 조직들에 비하여 우세하게 나타났다. 이는 아가미가 다른 조직들과 달리 어류의 생체기관 중 유일하게 외부와 직접 접촉이 가능한 조직으로서 활성산소에 대한 손상을 최소화하기 위한 것으로 사료된다.