• 제목/요약/키워드: Up-regulated

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Establishment of a Pancreatic Cancer Stem Cell Model Using the SW1990 Human Pancreatic Cancer Cell Line in Nude Mice

  • Pan, Yan;Gao, Song;Hua, Yong-Qiang;Liu, Lu-Ming
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권2호
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    • pp.437-442
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    • 2015
  • Aim: To establish a pancreatic cancer stem cell model using human pancreatic cancer cells in nude mice to provide a platform for pancreatic cancer stem cell research. Materials and Methods: To establish pancreatic cancer xenografts using human pancreatic cancer cell line SW1990, nude mice were randomly divided into control and gemcitabine groups. When the tumor grew to a volume of $125mm^3$, they treated with gemcitabine at a dose of 50mg/kg by intraperitoneal injection of 0.2ml in the gemcitabine group, while the mice in control group were treated with the same volume of normal saline. Gemcitabine was given 2 times a week for 3 times. When the model was established, the proliferation of pancreatic cancer stem cells was observed by clone formation assay, and the protein and/or mRNA expression of pancreatic stem cell surface markers including CD24, CD44, CD133, ALDH, transcription factors containing Oct-4, Sox-2, Nanog and Gli, the key nuclear transcription factor in Sonic Hedgehog signaling pathway was detected by Western blot and/or RT-PCR to verify the reliability of this model. Results: This model is feasible and safe. During the establishment, no mice died and the weight of nude mice maintained above 16.5g. The clone forming ability in gemcitabine group was stronger than that of the control group (p<0.01). In gemcitabine group, the protein expression of pancreatic cancer stem cell surface markers including CD44, and ALDH was up-regulated, the protein and mRNA expression of nuclear transcription factor including Oct-4, Sox-2 and Nanog was also significantly increased (P<0.01). In addition, the protein expression of key nuclear transcription factor in Sonic Hedgehog signaling pathway, Gli-1, was significantly enhanced (p<0.01). Conclusions: The pancreatic cancer stem cell model was successfully established using human pancreatic cancer cell line SW1990 in nude mice. Gemcitabine could enrich pancreatic cancer stem cells, simultaneously accompanied by the activation of Sonic Hedgehog signaling pathway.

Hypoxia Induced High Expression of Thioredoxin Interacting Protein (TXNIP) in Non-small Cell Lung Cancer and its Prognostic Effect

  • Li, Yan;Miao, Li-Yun;Xiao, Yong-Long;Huang, Mei;Yu, Min;Meng, Kui;Cai, Hou-Rong
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권7호
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    • pp.2953-2958
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    • 2015
  • Although associations between thioredoxin interacting protein (TXNIP) and cancers have been recognized, the effects of TXNIP on non-small cell lung cancer (NSCLC) prognosis remained to be determined in detail. In addition, while hypoxia is a key characteristic of tumor cell growth microenvironment, the effect of hypoxia on TXNIP expression is controversial. In this study, formaldehyde fixed and paraffin embedded (FFPE) samples of 70 NSCLC patients who underwent resection between January 2010 and December 2011 were obtained. Evaluation of TXNIP and hypoxia inducible factor-$1{\alpha}$ ($HIF-1{\alpha}$) protein expression in FFPE samples was made by immunohistochemistry. By Kaplan-Meier method, patients with high TXNIP expression demonstrated a significantly shorter progression free survival (PFS) compared with those with low TXNIP expression (18.0 months, 95%CI: 11.7, 24.3 versus 23.0 months, 95%CI: 17.6, 28.4, P=0.02). High TXNIP expression level was also identified as an independent prognostic factor by Cox regression analysis (adjusted hazard ratio: 2.46; 95%CI: 1.08, 5.56; P=0.03). Furthermore, TXNIP expression was found to be significantly correlated with $HIF-1{\alpha}$ expression (Spearman correlation=0.67, P=0.000). To further confirm correlations, we established a tumor cell hypoxic culture model. Expression of TXNIP was up-regulated in all three NSCLC cell lines (A549, SPC-A1, and H1299) under hypoxic conditions. This study suggests that hypoxia induces increased TXNIP expression in NSCLC and high TXNIP expression could be a poor prognostic marker.

Solute Carrier SLC41A1 'A MINI REVIEW'

  • Basnet Hom Bahadur
    • 한국환경성돌연변이발암원학회지
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    • 제25권2호
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    • pp.60-65
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    • 2005
  • The human solute carrier, SLC41Al, is a $Mg^{2}+$ transporter that is regulated by extracellular magnesium. Although intracellular magnesium plays a fundamental role in cellular metabolism, little is known about how $Mg^{2}+$ is taken up and controlled by cells. Magnesium plays a fundamental role in cellular metabolism so that its control within the body is critical. Magnesium homeostasis is principally a balance between intestinal absorption of dietary magnesium and renal excretion of urinary magnesium. The kidney, mainly the distal convoluted tubule, controls magnesium reabsorption. Although renal reabsorption is under the influence of many hormones, selective regulation of magnesium transport is due to intrinsic control involving transcriptional processes and synthesis of transport proteins. Using microarray analysis, identification of the genetic elements involved with this transcriptional control has been begun. SLC41A1(GenBank Accession No. AJ514402), comprises 10 putative transmembrane domains, two of which are highly homologous to the integral membrane part of the prokaryote transports $Mg^{2}+$ and other divalent cations $Sr^2+,\;Zn^2+,\;Cu^2+,\;Fe^2+,\;Co^2+,\;Ba^2+,\;and\;Cd^2+,\;but\;not\;Ca^2+,\;Mn^2+,\;and\;Ni^2+.$ Transport of $Mg^{2}+$ by SLC41Al is rheogenic, voltage dependent, and not coupled to Na or Cl. Expressed SLC41Al transports a range of other divalent cations: $Mg^{2+},\;Sr^{2+},\;Zn^{2+},\;Cu^{2+},\;Fe^{2+},\;Co^{2+},\;Ba^{2+},\;and\;Cd^{2+}$. The divalent cations $Ca^{2+},\;Mn^{2+},\;and\;Ni^{2+}$and the trivalent ion $Gd^{3+}$ did not induce currents nor did they inhibit $Mg^{2+}$ transport. The nonselective cation $La^{3+}$ abolishes $Mg^{2+}$ uptake. Computer analysis of the SLC41Al protein structure reveals that it belongs to MgtE protein family & suggested that the human solute carrier, SLC41Al, might be a eukaryotic $Mg^{2+}$ transporter closely related $(60-70\%)$ protein encoded by SLC41A2 is a $Mg^{2}+$ transporter that might be involved in magnesium homeostasis in epithelial cells also transports a range of other divalent cations: $Ba^2,\;Ni^2,\;CO^2,\;Fe^2,\;or\;Mn^2,\;but\;not\;Ca^2,\;Zn^2,\;or\;Cu^{2+}$ that may have related functional properties.

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고 콜레스테롤 식이 투여 흰쥐에 있어서 녹차의 콜레스테롤 저하 효과 (Hypocholesterolemic Effects of Green Tea in Cholesterol-Fed Rats)

  • 진현화;양정례;정종화;김양하
    • 한국식품영양과학회지
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    • 제33권1호
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    • pp.47-51
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    • 2004
  • 본 연구는 녹차의 혈중 콜레스테롤 저하 효과 및 녹차 추출물인 catechin과 EGCG가 hypocholesterolemic 효과에 미치는 정도를 식이로 고콜레스테롤혈증을 유도한 흰쥐에서 평가하고자 수행하였다. 4주령의 수컷 SD계 흰쥐를 난괴법에 따라 각 군당 5마리씩 무녹차섭취군(대조군), 4% 녹차분말군(GTP), 1% 녹차 catechin군(Catechin),0.5% 녹차 EGCG군(EGCG)의 4군으로 7주간 사육을 하였다. 녹차 첨가군은 모두 0.5% EGCG를 함유하도록 녹차 첨가량을 결정하였으며 모든 식이는 고콜레스테롤혈증을 유도하기 위하여 1% cholesterol과 0.5% cholic acid를 첨가하였다. 식이섭취량은 각 군간에 유의적인 차이를 나타내지 않았다. 혈청 총 및 LDL-콜레스테롤 수준은 Catechin군이 189.2 mg/dL, 166 mg/dL, EGCG군에서는 161.4 mg/dL, 138 mg/dL로 나타나 대조군(245 mg/dL, 222 mg/dL)에 비하여 유의적으로 감소되었고 (p<0.05), EGCG군에서 그 효과는 더욱 크게 나타났다. 혈청의 중성지방농도는 녹차섭취군이 대조군보다 낮았으며 EGCG군에서 가장 낮게 나타났다. HDL-콜레스테롤 수준은 유의적이지는 않았지만 녹차섭취군이 대조군보다 높은 경향을 보였고, HDL/LDL-콜레스테롤 비율은 대조군에 비하여 녹차섭취군에서 증가하여 EGCG군에서 가장 높았다. 이와 같은 결과는 녹차의 섭취가 혈중 콜레스테롤 및 중성지방을 감소하고, HDL-콜레스테롤은 증가시킴으로써 동맥경화 위험을 줄일 수 있는 가능성을 반영하고 있다. 간에서의 CYP7Al활성과 유전자 발현정도는 Catechin군, EGCG군이 대조군에 비하여 높게 나타났으며, EGCG군에서 가장 많이 증가하여 혈청에서의 지질수준 감소 경향과 일치하였다. 이상의 본 연구 결과들은 녹차 섭취군에서의 혈중 콜레스테롤 저하작용 특히 EGCG군에서의 현저한 hypocholesterolemic 효과는 CYP7Al 유전자 발현 및 효소의 활성증가에 따른 콜레스테롤의 담즙산으로의 배설 증가에 기인하는 것으로 추측된다.

형질전환된 벼세포 배양에 있어서 삼투압 조절에 따른 hCTLA4-lg 생산성 변화 (Effect of Osmotic Pressure on hCTLA-lg Production in Transgenic Rice Cell Suspension Cultures)

  • 최성훈;이송재;홍석미;조지숙;김동일
    • KSBB Journal
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    • 제20권4호
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    • pp.278-284
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    • 2005
  • 식물세포의 느린 생장과 낮은 생산량의 이유로 지금까지 는 주로 미생물이나 동물세포에서 유전자 재조합 단백질을 생산하여 왔다. 그러나 저렴한 배지 가격, 동물 유래 바이러스 감염 위험성으로부터의 안정성, glycosylation 등의 post-translational modification이 가능하다는 장점들로 인하여 최근 들어 식물세포배양은 생물학적 활성을 가진 고부가가치의 단백질을 생산하는데 많이 이용되고 있다. 본 연구에서는 생장배지에 첨가했던 sucrose의 소비와 induction 배지로의 교환에서 오는 배지내의 삼투압을 조절하여 hCTLA4-Ig의 생산성을 높이고자 하였다. 다양한 삼투압 조절제 첨가 실험을 통해 sorbitol을 선별하고, 40 mM의 sorbitol 첨가에서 상대적으로 높은 생존도와 induction 후 7일째 대조구보다 1.7배 높은 생산성을 확인하였다. 또한, 저농도의 glucose 첨가를 통한 생산성 증대에 있어서는 8 mM glucose에서 induction 이후에도 높은 세포농도를 유지하면서 최대 37.3 mg/L까지 hCTLA4-Ig 생산량을 증가시켰다. 5-L bioreactor에서 회분식 배양과 induction시의 hCTLA4-Ig 생산량을 비교한 결과 induction시 배양 18일째 최고 45.3 mg/L까지 높일 수 있었으며, 회분식 배양에 비해 2.1배 증가됨을 확인하였다.

Testosterone 처리에 의한 넙치, Paralichthys olivaceus 난소에서 doublesex-and mab-3-related transcription factor-1 (DMRT-1) mRNA의 발현 유도 (Induced Expression of Doublesex-and mab-3-related Transcription Factor-1 (DMRT-1) mRNA by Testosterone in the Olive Flounder, Paralichthys olivaceus ovary)

  • 조필규;안광욱;김나나;최용기;조성환;민병화;임한규;최철영
    • 한국양식학회지
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    • 제20권3호
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    • pp.199-202
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    • 2007
  • 본 연구에서는 넙치(Paralichthys olivaceus) 정소로부터 DMRT-1 partial cDNA를 분리하였다. 넙치 DMRT-1은 317개의 염기로 구성되어 있으며, Atlantic halibut, 감성돔 및 무지개 송어와 각각 94%, 85%, 82%의 높은 상동성을 보였다. RT-PCR을 이용하여 DMRT-1 mRNA 발현은 난소보다 정소에서 높게 나타난 것을 관찰할 수 있었다. 또한, 암컷 넙치에 웅성호르몬인 testosterone 처리시 처리 농도에 따라 난소에서 DMRT-1 mRNA 발현이 증가함을 볼 수 있었다. 따라서, DMRT-1은 수컷에서 특이적으로 발현되는 유전자임을 알 수 있으며, 본 연구의 결과는 넙치의 성 전환 및 유도에 대한 기초자료를 제공할 것이다.

Molecular Cloning and Functional Analysis of the Gene Encoding 3-hydroxy-3-methylglutaryl Coenzyme A Reductase from Hazel (Corylus avellana L. Gasaway)

  • Wang, Yechun;Guo, Binhui;Zhang, Fei;Yao, Hongyan;Miao, Zhiqi;Tang, Kexuan
    • BMB Reports
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    • 제40권6호
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    • pp.861-869
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    • 2007
  • The enzyme 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR; EC1.1.1.34) catalyzes the first committed step of isoprenoids biosynthesis in MVA pathway. Here we report for the first time the cloning and characterization of a full-length cDNA encoding HMGR (designated as CgHMGR, GenBank accession number EF206343) from hazel (Corylus avellana L. Gasaway), a taxol-producing plant species. The full-length cDNA of CgHMGR was 2064 bp containing a 1704-bp ORF encoding 567 amino acids. Bioinformatic analyses revealed that the deduced CgHMGR had extensive homology with other plant HMGRs and contained two transmembrane domains and a catalytic domain. The predicted 3-D model of CgHMGR had a typical spatial structure of HMGRs. Southern blot analysis indicated that CgHMGR belonged to a small gene family. Expression analysis revealed that CgHMGR expressed high in roots, and low in leaves and stems, and the expression of CgHMGR could be up-regulated by methyl jasmonate (MeJA). The functional color assay in Escherichia coli showed that CgHMGR could accelerate the biosynthesis of $\beta$-carotene, indicating that CgHMGR encoded a functional protein. The cloning, characterization and functional analysis of CgHMGR gene will enable us to further understand the role of CgHMGR involved in taxol biosynthetic pathway in C. avellana at molecular level.

과발현 형질전환벼에서 CCCH type zinc-finger protein 유전자 OsZF2 기능 분석 (Functional characterization of a CCCH type zinc-finger protein gene OsZF2 by ectopic overexpression of the gene in rice)

  • 이정숙;윤인선;윤웅한;이강섭;변명옥;서석철
    • Journal of Plant Biotechnology
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    • 제36권1호
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    • pp.23-29
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    • 2009
  • 벼의 저온처리 cDNA 은행에서 분리된 CCCH 형태 zinc finger 단백질인 OsZF2의 기능을 분석하기 위하여 벼에서 CaMV 35S 프로모터 조절하에 OsZF2가 발현(35S:OsZF2)되는 형질전환벼 식물체를 개발하였다. 35S:OsZF2 형질전환벼에 대한 하이그로 마이신저항성 검정을 통해 동형접합체 계통을 선발하고 Northern 발현분석에 의해 OsZF2 유전자가 형질전환체에서 과발현되는 것을 확인하였다. 형질전환체와 대조구인 낙동벼를 100 mM NaCl 첨가 MS 배지에서 키운 후 잎과 뿌리의 길이를 측정하여 내염성 검정을 수행한 결과 대조구에 비해 형질전환체 생육이 다소 양호 한 것으로 나타났다. GMO 포장에서 생육상태를 관찰 한 결과 형질전환체는 생육지연으로 인한 왜화 현상을 나타내며 출수기 또한 열흘 정도 지연되나 등숙기에는 대조구와 같은 초장을 보였다. zinc finger 유전자는 식물체의 발달과 분화 단계 및 환경 스트레스 반응 등 중요한 역할을 하는 것으로 알려져 있으므로 유전체발현 분석으로 하위단계에서 조절되는 유전자 발현 양상을 분석하였다. 35S:OsZF2 전환체에서 낙동벼보다 4배 이상 발현이 증가된 유전자 중에서 게놈 주석에 기초한 기능을 유추하면 신호전달과 관련된 protein kinase, DNA 결합단백질과 대사에 관련된 효소 유전자, 스트레스 반응에 관여하는 일부 유전자 및 병 저항성과 관련된 유전자들의 발현이 증가되었다. 따라서 벼에서 분리된 OsZF2 CCCH type zinc finger 유전자는 벼 성장 발달과 스트레스에 반응하는 상위 조절자로서 기능을 할 것으로 추측된다.

Molecular Cloning and Function Analysis of an Anthocyanidin Synthase Gene from Ginkgo biloba, and Its Expression in Abiotic Stress Responses

  • Xu, Feng;Cheng, Hua;Cai, Rong;Li, Lin Ling;Chang, Jie;Zhu, Jun;Zhang, Feng Xia;Chen, Liu Ji;Wang, Yan;Cheng, Shu Han;Cheng, Shui Yuan
    • Molecules and Cells
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    • 제26권6호
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    • pp.536-547
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    • 2008
  • Anthocyanidin synthase (ANS, leucoanthocyanidin oxygenase), a 2-oxoglutarate iron-dependent oxygenase, catalyzed the penultimate step in the biosynthesis of the anthocyanin class of flavonoids, from the colorless leucoanthocyanidins to the colored anthocyanidins. The full-length cDNA and genomic DNA sequences of ANS gene (designated as GbANS) were isolated from Ginkgo biloba for the first time. The full-length cDNA of GbANS contained a 1062-bp open reading frame (ORF) encoding a 354-amino-acid protein. The genomic DNA analysis showed that GbANS gene had three exons and two introns. The deduced GbANS protein showed high identities to other plant ANSs. The conserved amino acids (H-X-D) ligating ferrous iron and residues (R-X-S) participating in 2-oxoglutarate binding were found in GbANS at the similar positions like other ANSs. Southern blot analysis indicated that GbANS belonged to a multi-gene family. The expression analysis by real-time PCR showed that GbANS expressed in a tissue-specific manner in G. biloba. GbANS was also found to be up-regulated by all of the six tested abiotic stresses, UV-B, abscisic acid, sucrose, salicylic acid, cold and ethylene, consistent with the promoter region analysis of GbANS. The recombinant protein was successfully expressed in E. coli strain with pET-28a vector. The in vitro enzyme activity assay by HPLC indicated that recombinant GbANS protein could catalyze the formation the cyanidin from leucocyanidin and conversion of dihydroquercetin to quercetin, suggesting GbANS is a bifunctional enzyme within the anthocyanidin and flavonol biosynthetic pathway.

Astragaloside IV Prevents Obesity-Associated Hypertension by Improving Pro-Inflammatory Reaction and Leptin Resistance

  • Jiang, Ping;Ma, Dufang;Wang, Xue;Wang, Yongcheng;Bi, Yuxin;Yang, Jinlong;Wang, Xuebing;Li, Xiao
    • Molecules and Cells
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    • 제41권3호
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    • pp.244-255
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    • 2018
  • Low-grade pro-inflammatory state and leptin resistance are important underlying mechanisms that contribute to obesity-associated hypertension. We tested the hypothesis that Astragaloside IV (As IV), known to counteract obesity and hypertension, could prevent obesity-associated hypertension by inhibiting pro-inflammatory reaction and leptin resistance. High-fat diet (HFD) induced obese rats were randomly assigned to three groups: the HFD control group (HF con group), As IV group, and the As IV + ${\alpha}$-bungaratoxin (${\alpha}-BGT$) group (As IV+${\alpha}-BGT$ group). As IV ($20mg{\cdot}Kg^{-1}{\cdot}d^{-1}$) was administrated to rats for 6 weeks via daily oral gavage. Body weight and blood pressure were continuously measured, and NE levels in the plasma and renal cortex was evaluated to reflect the sympathetic activity. The expressions of leptin receptor (LepRb) mRNA, phosphorylated signal transducer and activator of transcription-3 (p-STAT3), phosphorylated phosphatidylinositol 3-kinase (p-PI3K), suppressor of cytokine signaling 3 (SOCS3) mRNA, and protein-tyrosine phosphatase 1B (PTP1B) mRNA, pro-opiomelanocortin (POMC) mRNA and neuropeptide Y (NPY) mRNA were measured by Western blot or qRT-PCR to evaluate the hypothalamic leptin sensitivity. Additionally, we measured the protein or mRNA levels of ${\alpha}7nAChR$, inhibitor of nuclear factor ${\kappa}B$ kinase subunit ${\beta}/nuclear$ factor ${\kappa}B$ ($IKK{\beta}/NF-KB$) and pro-inflammatory cytokines ($IL-1{\beta}$ and $TNF-{\alpha}$) in hypothalamus and adipose tissue to reflect the anti-inflammatory effects of As IV through upregulating expression of ${\alpha}7nAChR$. We found that As IV prevented body weight gain and adipose accumulation, and also improved metabolic disorders in HFD rats. Furthermore, As IV decreased BP and HR, as well as NE levels in blood and renal tissue. In the hypothalamus, As IV alleviated leptin resistance as evidenced by the increased p-STAT3, LepRb mRNA and POMC mRNA, and decreased p-PI3K, SOCS3 mRNA, and PTP1B mRNA. The effects of As IV on leptin sensitivity were related in part to the up-regulated ${\alpha}7nAchR$ and suppressed $IKK{\beta}/NF-KB$ signaling and pro-inflammatory cytokines in the hypothalamus and adipose tissue, since co-administration of ${\alpha}7nAChR$ selective antagonist ${\alpha}-BGT$ could weaken the improved effect of As IV on central leptin resistance. Our study suggested that As IV could efficiently prevent obesityassociated hypertension through inhibiting inflammatory reaction and improving leptin resistance; furthermore, these effects of As IV was partly related to the increased ${\alpha}7nAchR$ expression.