• 제목/요약/키워드: Vector Line

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Overexpression of Cyclin L2 Inhibits Growth and Enhances Chemosensitivity in Human Gastric Cancer Cells

  • Li, Hong-Li;Huang, Ding-Zhi;Deng, Ting;Zhou, Li-Kun;Wang, Xia;Bai, Ming;Ba, Yi
    • Asian Pacific Journal of Cancer Prevention
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    • 제13권4호
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    • pp.1425-1430
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    • 2012
  • Cyclin L2 is a novel member of the cyclin family, recently implicated in the regulation of cell cycle progression and/or transcriptional regulation. The present study was undertaken to investigate the effects of overexpression on tumor cell growth and chemosensitivity in human gastric cells in vitro. Cyclin L2 was transfected into human gastric cancer cell line BCG823 and expressed with a mammalian expression vector pcDNA3.1. The effects and mechanisms of cyclin L2 on cell growth, cell cycling and apoptosis were studied. Compared to control vectors, overexpression of cyclin L2 inhibited the growth of BCG823 cells and enhance their chemosensitivity to fluorouracil, docetaxel and cisplatin. The anti-proliferative effects of cyclin L2 could be due to G0/G1 arrest and apoptosis. Cyclin L2 induced G0/G1 arrest and apoptosis involved upregulation of caspase-3 and down regulation Bcl-2 and survivin. The results indicated that overexpression of cyclin L2 protein may promote efficient growth inhibition and enhance chemosensitivity to chemotherapeutic agents in human gastric cancer cells by inducing G0/G1 cell cycle arrest and apoptosis.

5축 가공에 의한 SCM415 롤러 캠 개발과 표면조도 연구 (A Study on the Development and Surface Roughness of Roller Cam SCM415 by 5-Axis Machining)

  • 김진수;이동섭;강성기
    • 한국정밀공학회지
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    • 제30권4호
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    • pp.397-402
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    • 2013
  • In this study, we carried out the each lines of section, using GC (green silicon carbide) whetstone, the SCM415 material which separated by after and before heat treatments process, in 3+2 axis machining centers for integrated grinding after cutting end mill works, the spindle speed 8000 rpm and feed rate 150 mm/min. For the analysis of the centerline average roughness (Ra), we measured by 10 steps stages. Using Finite element analysis, we found the result of the load analysis effect of the assembly parts, when applied the 11 kg's load on both side of the ATC (Automatic tool change) arm. The result is as follows. For the centerline average roughness (Ra) in the non-heat treatment work pieces, are appeared the most favorable in the tenth section are $0.510{\mu}m$, that were shown in the near the straight line section which is the smallest deformation of curve. In addition, the bad surface roughness appears on the path is to long by changing angle, the more inclined depth of cut, because the chip discharging is not smoothly.

Expression of CYP2A6, CYP2D6 and CYP4A11 Polymorphisms in COS7 Mammalian Cell Line

  • Lee, Hye-Ja;Park, Mi-Kyung;Park, Young-Ran;Kim, Dong-Hak;Yun, Chul-Ho;Chun, Young-Jin;Shin, Hee-Jung;Na, Han-Sung;Chung, Myeon-Woo;Lee, Chang-Hoon
    • Toxicological Research
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    • 제27권1호
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    • pp.25-29
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    • 2011
  • The cytochrome P450 (P450, CYP) are the superfamily of heme-containing monooxygenase enzymes, found throughout all nature including mammals, plants, and microorganisms. Mammalian P450 enzymes are involved in oxidative metabolism of a wide range of endo- and exogenous chemicals. Especially P450s involved in drug metabolisms are important for drug efficacy and polymorphisms of P450s in individuals reflect differences of drug responses between people. To study the functional differences of CYP2A6, CYP2D6, and CYP4A11 variants, we cloned the four CYP2A6, three CYP2D6, and three CYP4A11 variants, which were found in Korean populations, in mammalian expression vector pcDNA by PCR and examined their expressions in COS-7 mammalian cells using immunoblots using P450 specific polyclonal antibodies. Three of four CYP2A6, two of three CYP4A11, and two of three CYP2D6 variants showed expressions in COS-7 cells but the relative levels of expressions are remarkably different in those of each variants. Our findings may help to study and explain the differences between functions of CYP variants and drug responses in Korean populations.

형질전환 토마토에서 Antisense Polygalacturonase 유전자의 발현 (Expression of Antisense Polygalacturonase Gene in Transgenic Tomato)

  • 김영미;김용환;이성갑;임명호;송경수
    • 식물조직배양학회지
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    • 제22권6호
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    • pp.351-355
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    • 1995
  • 국내 재배종 토마토 서광 품종으로부터 분리한 Polygalacturonase 유전자(PG2)의 3'측 1.1 kb cDNA 단편을 식물 형질전환용 운반체에 antisense 방향으로 삽입한 후 자엽을 이용하여 토마토내 도입하여 형질전환 토마토를 획득하였다. 형질전환 토마토(T$^{0}$ )를 도입시켜 그 종자를 1 mg/mL 농도의 kanamycin 함유 MS 배지에서 발아시켜 분리 집단 중에서 T$_1$9 식물체를 얻었다. T$_1$9의 Genomic Southern blot 분석 결과, antisense PG 유전자 1개가 염색체 내로 삽입되었음을 확인하였고 RNA gel blot 분석으로 endogenous PG mRNA보다 antisense PG RNA가 강하게 발현됨을 확인하였다. T$_1$9 계통 10개체의 성숙 토마토 과피조직내의 PG 효소 활성도 4~60%까지 저해되었다.

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초정밀 미세 패턴을 위한 전기 수력학 잉크젯 프린팅 시스템 (Electrohydrodynamic Inkjet Printing System for Ultrafine Patterning)

  • 노형래;고정국;권계시
    • 대한기계학회논문집B
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    • 제37권9호
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    • pp.873-877
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    • 2013
  • 잉크젯 기술은 가정용 프린터에서부터 제조 도구로 확대 되었다. 최근 인쇄전자 분야에서 고해상도 인쇄가 요구되고 있다. 기존의 잉크젯 인쇄 패터닝 방식을 향상 시키기 위해 전기수력학잉크젯 기술이 최근 주목을 받고 있는데 노즐 직경보다 작은 방울을 토출할 수 있고 넓은 점도 범위와 재료를 사용할 수 있기 때문이다. 본 논문에서는 미세 패터닝을 위한 EHD 프린팅 시스템이다. 요구 적출형 프린팅에 의해 다양한 패턴을 인쇄하고 벡터와 레스터 프린팅 알고리즘을 개발하였다. 내경이 $8{\mu}m$ 인 노즐을 이용하여 $7{\mu}m$ 이하의 미세 전도성 선폭을 EHD 방식을 통해 만들 수 있다.

RASAL1 Attenuates Gastric Carcinogenesis in Nude Mice by Blocking RAS/ERK Signaling

  • Chen, Hong;Zhao, Ji-Yi;Qian, Xu-Chen;Cheng, Zheng-Yuan;Liu, Yang;Wang, Zhi
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권3호
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    • pp.1077-1082
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    • 2015
  • Recent studies have suggested that the RAS protein activator like-1 (RASAL1) functions as a tumor suppressor in vitro and may play an important role in the development of gastric cancer. However, whether or not RASAL1 suppresses tumor growth in vivo remains to be determined. In the present study, we investigated the role of RASAL1 in gastric carcinogenesis using an in vivo xenograft model. A lentiviral RASAL1 expression vector was constructed and utilized to transfect the human poorly differentiated gastric adenocarcinoma cell line, BGC-823. RASAL1 expression levels were verified by quantitative real-time RT-PCR and Western blotting analysis. Then, we established the nude mice xenograft model using BGC-823 cells either over-expressing RASAL1 or normal. After three weeks, the results showed that the over-expression of RASAL1 led to a significant reduction in both tumor volume and weight compared with the other two control groups. Furthermore, in xenograft tissues the increased expression of RASAL1 in BGC-823 cells caused decreased expression of p-ERK1/2, a downstream moleculein the RAS/RAF/MEK/ERK signal pathway. These findings demonstrated that the over-expression of RASAL1 could inhibit the growth of gastric cancer by inactivation of the RAS/RAF/MEK/ERK pathway in vivo. This study indicates that RASAL1 may attenuate gastric carcinogenesis.

Adenovirus-mediated Double Suicide Gene Selectively Kills Gastric Cancer Cells

  • Luo, Xian-Run;Li, Jian-Sheng;Niu, Ying;Miao, Li
    • Asian Pacific Journal of Cancer Prevention
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    • 제13권3호
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    • pp.781-784
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    • 2012
  • The aim of this study was to evaluate the effect of the adenovirus-mediated double suicide gene (CD/TK) for selective killing of gastric cancer cells. Gastric cancer cells SCG7901 and normal gastric epithelial cell lines were infected by adenoviruses Ad-survivin/GFP and Ad-survivin/CD/TK. GFP expression and CD-TK were detected by fluorescence microscopy and reverse transcriptase polymerase chain reaction (RT-PCR), respectively. After treatment of the infected cells with the pro-drugs ganciclovir (GCV) and/or 5-FC, the cell growth status was evaluated by methyl thiazolyl tetrazolium assay. Cell cycle changes were detected using flow cytometry. In nude mice bearing human gastric cancer, the recombinant adenovirus vector was injected directly into the tumor followed by an intraperitoneal injection of GCV and/or 5-FC. The subsequent tumor growth was then observed. The GFP gene driven by survivin could be expressed within the gastric cancer line SCG7901, but not in normal gastric epithelial cells. RT-PCR demonstrated the presence of the CD/TK gene product in the infected SCG7901 cells, but not in the infected normal gastric epithelial cells. The infected gastric cancer SCG7901, but not the gastric cells, was highly sensitive to the pro-drugs. The CD/TK fusion gene system showed significantly greater efficiency than either of the single suicide genes in killing the target cells (P<0.01). Treatment of the infected cells with the pro-drugs resulted in increased cell percentage in G0-Gl phase and decreased percentage in S phase. In nude mice bearing SCG7901 cells, treatment with the double suicide gene system significantly inhibited tumor growth, showing much stronger effects than either of the single suicide genes (P<0.01). The adenovirus-mediated CD/TK double suicide gene driven by survivin promoter combined with GCV an 5-FC treatment could be an effective therapy against experimental gastric cancer with much greater efficacy than the single suicide gene CD/TK combined with GCV or 5-FC.

Overexpression of Hyoscyamine 6${\beta}$-Hydroxylase (h6h) Gene and Enhanced Production of Tropane Alkaloids in Scopolia parviflora Hairy Root Lines

  • KANG, YOUNG-MIN;LEE, OK-SUN;JUNG, HEE-YOUNG;KANG, SEUNG-MI;LEE, BYUNG-HYUN;CHANDRAKANT KARIGAR;THEERTHA PRASAD;BAHK, JUNG-DONG;CHOI, MYUNG-SUK
    • Journal of Microbiology and Biotechnology
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    • 제15권1호
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    • pp.91-98
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    • 2005
  • The hyoscyamine 6${\beta}$-hydroxylase (h6h) gene was introduced into the genome of Scopolia parviflora through the Agrobacterium rhizogenes binary vector system. The enzyme was expressed ally and tissue specific selectively in roots, resulting in five transgenic hairy root lines. The presence of the h6h gene in kanamycin-resistant hairy roots and its overexpression were confirmed by polymerase chain reaction (PCR), Northern blotting, and Western blotting, respectively. In the transgenic hairy root lines which constitutively expressed the H6H enzyme, hyoscyamine and scopolamine accumulated in high concentration. Among the transgenic hairy root lines that expressed the H6H enzyme, only two were more productive. The levels of tropane alkaloids in transgenic hairy root varied greatly: The best transgenic line (#5) contained 8.12 mg of scopolamine per g dry weight, which produced the compound three times more than wild-type root. These results suggest a possibility of improving the yield of tropane alkaloids in hairy root lines by genetic and metabolic engineering.

RASSF1A Suppresses Proliferation of Cervical Cancer Cells

  • Feng, Lei;Li, Jie;Yan, Ling-Di;Tang, Jian
    • Asian Pacific Journal of Cancer Prevention
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    • 제15권14호
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    • pp.5917-5920
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    • 2014
  • Background: This study aimed to explore the effects of ras association domain family 1 A (RASSF1A) on proliferation and apoptosis of human cervical cancer cell line Hela cells. Materials and Methods: RASSF1A was cloned into the pcDNA3.1(+) vector to generate pcDNA3.1(+)-RASSF1A plasmid for transfection into Hela cells. Changes in the proliferation and apoptosis of cultured Hela cells were examined by the 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium chloride assay and flow cytometry. A protein array was used to analyze the expression of apoptotic factors. Results: Plasmid pcDNA3.1(+)-RASSF1A was generated and transfected into Hela cells to stably express RASSF1A in Hela cells. RASSF1A transfection was effective in inhibiting the proliferation of Hela cells up to 52.4%, as compared to cells transfected with an empty plasmid. RASSF1A expression also successfully induced apoptosis in human cervical cells with an apoptosis rate of 20.5%. More importantly, protein array results showed that RASSF1 A transfection induced overexpression of p21 and caspase 8, while decreasing the expression of survivin in Hela cells. Conclusions: RASSF1A expression was effective in suppressing the proliferation and increasing apoptosis of Hela cells, and may be a potential therapy for cervical cancer in clinic.

Cloning and characterization of a novel gene with alternative splicing in murine mesenchymal stem cell line C3H/10T1/2 by gene trap screening

  • Wang, Mingke;Sun, Huiqin;Jiang, Fan;Han, Jing;Ye, Feng;Wang, Tao;Su, Yongping;Zou, Zhongmin
    • BMB Reports
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    • 제43권12호
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    • pp.789-794
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    • 2010
  • A novel gene, designated mgt-6, containing four splicing variants, was isolated from a gene trap clone library of C3H/10T1/2 cells transfected with retroviral promoterless gene-trap vector, ROSAFARY. The transcript variants were differentially expressed in murine tissues and cell lines and differentially responded to diverse stimuli including TGF-${\beta}1$ and mitogen-activated protein kinase (MAPK) inhibitors. The mgt-6 gene encoded a protein of 37 or 11 amino acid residuals with cytoplasmic distribution. However, when C3H/10T1/2 cells were treated with 5-azacytidine, the protein translocated into cell nucleus as indicated by fused LacZ or C-terminally tagged EGFP. Our preliminary results suggest that further study on the role of mgt-6 gene in cell transformation and differentiation may be of significance.