• Title/Summary/Keyword: Growth Regulator

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Reconstructed Adeno-Associated Virus with the Extracellular Domain of Murine PD-1 Induces Antitumor Immunity

  • Elhag, Osama A.O.;Hu, Xiao-Jing;Wen-Ying, Zhang;Li, Xiong;Yuan, Yong-Ze;Deng, Ling-Feng;Liu, De-Li;Liu, Ying-Le;Hui, Geng
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.8
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    • pp.4031-4036
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    • 2012
  • Background: The negative signaling provided by interactions of the co-inhibitory molecule, programmed death-1 (PD-1), and its ligands, B7-H1 (PD-L1) and B7-DC (PD-L2), is a critical mechanism contributing to tumor evasion; blockade of this pathway has been proven to enhance cytotoxic activity and mediate antitumor therapy. Here we evaluated the anti-tumor efficacy of AAV-mediated delivery of the extracellular domain of murine PD-1 (sPD-1) to a tumor site. Material and Methods: An rAAV vector was constructed in which the expression of sPD-1, a known negative regulator of TCR signals, is driven by human cytomegalovirus immediate early promoter (CMV-P), using a triple plasmid transfection system. Tumor-bearing mice were then treated with the AAV/sPD1 construct and expression of sPD-1 in tumor tissues was determined by semi quantitative RT-PCR, and tumor weights and cytotoxic activity of splenocytes were measured. Results: Analysis of tumor homogenates revealed sPD-1 mRNA to be significantly overexpressed in rAAV/sPD-1 treated mice as compared with control levels. Its use for local gene therapy at the inoculation site of H22 hepatoma cells could inhibit tumor growth, also enhancing lysis of tumor cells by lymphocytes stimulated specifically with an antigen. In addition, PD-1 was also found expressed on the surfaces of activated CD8+ T cells. Conclusion: This study confirmed that expression of the soluble extracellular domain of PD-1 molecule could reduce tumor microenvironment inhibitory effects on T cells and enhance cytotoxicity. This suggests that it might be a potential target for development of therapies to augment T-cell responses in patients with malignancies.

Tumour Suppressive Effects of WEE1 Gene Silencing in Breast Cancer Cells

  • Ghiasi, Naghmeh;Habibagahi, Mojtaba;Rosli, Rozita;Ghaderi, Abbas;Yusoff, Khatijah;Hosseini, Ahmad;Abdullah, Syahrilnizam;Jaberipour, Mansooreh
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.11
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    • pp.6605-6611
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    • 2013
  • Background: WEE1 is a G2/M checkpoint regulator protein. Various studies have indicated that WEE1 could be a good target for cancer therapy. The main aim of this study was to asssess the tumor suppressive potential of WEE1 silencing in two different breast cancer cell lines, MCF7 which carries the wild-type p53 and MDA-MB468 which contains a mutant type. Materials and Methods: After WEE1 knockdown with specific shRNAs downstream effects on cell viability and cell cycle progression were determined using MTT and flow cytometry analyses, respectively. Real-time PCR and Western blotting were conducted to assess the effect of WEE1 inhibition on the expression of apoptotic (p53) and anti-apoptotic (Bcl2) factors and also a growth marker (VEGF). Results: The results showed that WEE1 inhibition could cause a significant decrease in the viability of both MCF7 and MDA-MB-468 breast cancer cell lines by more than 50%. Interestingly, DNA content assays showed a significant increase in apoptotic cells following WEE1 silencing. WEE1 inhibition also induced upregulation of the apoptotic marker, p53, in breast cancer cells. A significant decrease in the expression of VEGF and Bcl-2 was observed following WEE1 inhibition in both cell lines. Conclusions: In concordance with previous studies, our data showed that WEE1 inhibition could induce G2 arrest abrogation and consequent cell death in breast cancer cells. Moreover, in this study, the observed interactions between the pro- and anti-apoptotic proteins and decrease in the angiogenesis marker expression confirm the susceptibility to apoptosis and validate the tumor suppressive effect of WEE1 inhibition in breast cancer cells. Interestingly, the levels of the sensitivity to WEE1 silencing in breast cancer cells, MCF7 and MDA-MB468, seem to be in concordance with the level of p53 expression.

An Integrated Biological Control Using an Endoparasitoid Wasp (Cotesia plutellae) and a Microbial Insecticide (Bacillus thuringiensis) against the Diamondback Moth, Plutella xylostella (배추좀나방에 대한 프루텔고치벌과 미생물농약의 통합생물방제)

  • Kim, Kyusoon;Kim, Hyun;Park, Young-Uk;Kim, Gil-Hah;Kim, Yonggyun
    • Korean journal of applied entomology
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    • v.52 no.1
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    • pp.35-43
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    • 2013
  • All tested Korean populations of the diamondback moth, Plutella xylostella, are known to be resistant especially against pyrethroid insecticides by mutation in its molecular target, para-sodium channel. Moreover, P. xylostella is able to develop resistance against most commercial insecticides. This study was performed to develop an efficient control technique against P. xylostella by a combined treatment of an endoparasitoid wasp, Cotesia plutellae, and a microbial insecticide, Bacillus thuringiensis. To investigate any parasitism preference of C. plutellae against susceptible and resistant P. xylostella, five different populations of P. xylostella were compared in insecticide susceptibilities and parasitism by C. plutellae. These five P. xylostella populations showed a significant variation against three commercial insecticides including pyrethroid, organophosphate, neonicotinoid, and insect growth regulator. However, there were no significant differences among five P. xylostella populations in their parasitic rates by C. plutellae. Moreover, parasitized larvae of P. xylostella showed significantly higher susceptibility to B. thuringiensis. As an immunosuppressive agent, viral ankyrin genes (vankyrins) encoded in C. plutellae were transiently expressed in nonparasitized larvae. Expression of vankyrins significantly enhanced the efficacy of B. thuringiensis against the third instar larvae of P. xylostella. Thus an immunosuppression induced by C. plutellae enhanced the insecticidal efficacy of B. thuringiensis. These results suggest that a combined treatment of C. plutellae and B. thuringiensis may effectively control the insecticide-resistant populations of P. xylostella.

Production of Recombinant Human Hyperglycosylated Erythropoietin Using Cell Culture Technology by Improving Sialylation. (Sialic Acid 함량 증가 배양기술에 의한 재조합 인간 다당쇄 에리스로포이에틴의 생산)

  • 박세철;이승오;박만식;김승훈;김준환;송무영;이병규;고인영;강희일
    • Microbiology and Biotechnology Letters
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    • v.32 no.2
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    • pp.142-148
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    • 2004
  • Erythropoietin is a main regulator of human erythropoiesis. Recombinant human erythropoietin (rhEPO) is one of the glycoproteins produced in animal cells, and it has oligo saccharides chains which comprise about 40% of its molecular mass. Because the content of sialic acid can extend circulatory lifetime, the high degree of sialylation is often a desirable feature of therapeutic glycoproteins. In this study, the sialylation of rhEPO produced by chinese hamster ovary cell culture was maximized by supplementing the culture medium with N-acetylm-annosamine (ManNAc), a direct intracellular precursor for sialic acid synthesis and 2-deoxy-2,3-dehydro-N-acetylneuraminic acid (NeuAc2en), a sialidase inhibitor. Feeding of 20 mM ManNAc/0.5 mM NeuAc2en into culture medium increased the sialic acid content by nearly tenfold compared with unsupplemented medium. This effect was achieved without affecting the cell growth or product yield. Six erythropoietin fractions differing in sialic acid content, ranging from 11∼15% of EPO, were identified from chinese hamster ovary cell-derived rhEPO by mono Q column chromatography. It was found that, at 20 mM ManNAc/0.5 mM NeuAc2en feeding, productivity of hyper-glycosylated EPO increased up to 50%, compared with the unsupplemented medium.

In vitro shoot regeneration from leaf tissue of "Whangkeumbae" pear(Pyrus pyrifolia Nakai) (황금배(Pyrus pyrifolia Nakai) 잎 조직으로부터 기내 신초 재분화)

  • Chun, Jae An;Do, Kyung Ran;Kim, Se Hee;Cho, Kang-Hee;Kim, Hyun Ran;Hwang, Hae Sung;Shin, Il Sheob
    • Journal of Plant Biotechnology
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    • v.39 no.4
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    • pp.288-294
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    • 2012
  • In order to establish an efficient adventitious shoot regeneration conditions from leaf explants for Asian pear 'Whangkeumbae', the effect of concentration and kinds of plant growth regulator and carbon source was investigated. Leaf explants of cultures grown on Murashige and Skoog (MS) medium containing 8 g/L plant agar were used. When the medium contained 0.25 mg/L thidiazuron (TDZ) and 0.3 mg/L indolebutyric acid (IBA), the adventitious shoot regeneration rate (ASRR) was greater as 61.1% than others treated and higher TDZ concentrations (2.5 and 5 mg/L) treatment significantly reduced the ASRR. As the effect of IBA and indoleacetic acid (IAA) concentration on the ASRR, 0.5 mg/L TDZ plus different concentration of IAA exhibited relatively high ASRR and 0.5 mg/L TDZ plus 0.3 mg/L IAA showed the highest ASRR of 76.7%. Also the effect of sucrose and sorbitol as carbon source on regeneration was examined. The highest ASRR and the most shoots per explants averaged 94.4% and 3.49 by treatment of 30 mg/L sorbitol, respectably. Sorbitol is considered better carbon source than sucrose for shoot regeneration of 'Whangkeumbae' pear.

Characterization of cadC and cadR Mutants in Mediating the Expression of the Salmonella typhimurium cadBA Operon (Salmonella typhimurium cadBA 오페론의 발현에 관여하는 돌연변이체의 선별 및 그 특성)

  • 방성호;박용근
    • Korean Journal of Microbiology
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    • v.37 no.4
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    • pp.259-264
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    • 2001
  • It has been well known that the expression of S. typhimurium cadBA operon requires at least two extracellular signals: low pH and high concentration of lysine. To better understand the nature of pH-dependent and lysine dependent signal transduction, mutants were isolated in JF2238(cadA-lacZ) by Tn10 insertion, spontaneous mutagenesis, and EMS treatment. Two mutants were isolated from JF2238, expressed as a cadA-lacZ operon fusion in various growth conditions, and analyzed to have mutations in cadC, a gene encoding a function necessary for transcriptional activation of cadBA. One isolate (cadC6) conferred pH-independent and lysine-independent cadBA expression and the other(cadC4) showed pH-independent and lysine-dependent cadBA expression. cadR::Tn10 and cadR4 mutants were expressed in the absence of exogenously added lysine. They were also resistant to thiosine and complemented by lysP clone from E. coli. Thus, in the absence of exogenous lysine, cadR is a negative regulator of cadBA expression. Cadaverine, the product of lysine decarboxylation, was shown to inhibit expression of cadA-lacZ fusion in cad $C^+$ cell.

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Determination of Optimal Collecting Date and Exogenous Auxin Dipping Treatments in Cutting Transplants of 'Seolhyang' Strawberry (Fragaria × ananassa Duch.) ('설향' 딸기 삽목묘의 최적 삽수 채취시기와 오옥신 처리 구명)

  • Kim, Eun Ji;Uhm, Mi Jeong;Jung, Hyun Soo;Kim, Jong Yeob;Lee, Jun Gu
    • Journal of Bio-Environment Control
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    • v.29 no.3
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    • pp.252-258
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    • 2020
  • The aim of this study was to investigate the effects of timing of collecting date and concentration of IBA and NAA, in order to enhance initial activity and seedling quality of domestic strawberry. Strawberry cuttings were separately taken twice, in June 7 and in July 5, and IBA and NAA were treated with the concentrations of 0.025, 0.05 and 0.1% at cutting date, respectively. The seedlings were evaluated for the percentage of survival during 18 days at 6 times after tunnel cultivation. The NAA treatment was inappropriate for strawberry cutting due to the high rate of seedling mortality, regardless of the collecting date. The vitality of the seedlings was highest at IBA 0.1% in June collecting and at IBA 0.05% in July collecting. The seedlings from June collecting had a higher quantum yield at IBA 0.1% and the seedlings from July collecting at IBA 0.05%. Therefore, IBA could be more effectively applied than NAA to promote the vitality and quality with the appropriate concentration of 0.1% at June collecting and 0.05% at July collecting, respectively.

Shoot Primordium Culture for Multiplication of Carrot (당근의 다량증식을 위한 순원기 배양)

  • 서호범;이수성
    • Korean Journal of Plant Tissue Culture
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    • v.26 no.2
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    • pp.93-97
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    • 1999
  • Shoot tips with 2 leaf primordia were cultured to induce shoot primordia in MS liquid medium supplemented with several concentrations of BA and hIAA under the conditions of 10,000 lux illuminations for 24 h and of vertical shaking of 2 rpm in carrot. Two F$_1$ hybrids and two male sterility lines were used. Shoot primordia were only induced in the medium supplemented with 2.0 mg/L of BA and 0.2 mg/L of NAA. Genotypic specificity and seasonal effect of donor parents on shoot primordia induction were not observed and average 15-20% of the planted dornes developed to shoot primordia. The induced shoot primordia were successfully propagated by subculture in the same medium. However, they were grown into three different types during multiplication, that is, the type with multiple small shoots on the surface, the type of without any shoot, and the type of callus. Shoot primordia clusters with small shoots on the surface differentiated multiple shoots successfully in 1/2 MS solid medium supplemented with 0.2 to 1.0 mg/L of IAA and 0.2 to 1.0 mg/L of kinetin. New shoot primordia with small shoots were well formed when pieces bigger than 2 mm in diameter of the out layer of the shoot primordia cluster with small shoots were subcultured. No differences of multiplication and shooting ability and chromosomal variation of shoot primordia were observed until the 13th sub-culture.

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Gonadotropin Regulation of Regulator of G Protein Signaling 2 (RGS-2) Expression in the Rat Ovary (백서 난소에서 성선자극호르몬에 의한 RGS-2의 발현 조절)

  • Lee, Yu-Il;Lee, Eun-Suk;Kim, Sun-Ae;Kim, Mi-Young;Cho, Moon-Kyoung;Chun, Sang-Young
    • Clinical and Experimental Reproductive Medicine
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    • v.35 no.2
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    • pp.111-118
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    • 2008
  • Objective: The purpose of the present study was to examine the hormonal regulation of RGS-2 in the rat ovary. Methods: Immature rats were injected with 10 IU of PMSG to induce multiple growth of preovulatory follicles and 10 IU of hCG to induce ovulation. Northern blot analysis performed for gene expression and in situ hybridization performed for mRNA localization. Results: Northern blot analysis revealed that pregnant mare's serum gonadotropin (PMSG) treatment did not affect RGS-2 mRNA levels. In contrast, human chorionic gonadotropin (hCG) treatment of PMSG-primed rats resulted in an increase in RGS-2 expression within $1{\sim}3\;h$. The major cell-types expressing RGS-2 mRNA were oocytes regardless of follicle size. Interestingly, hCG treatment caused the stimulation of RGS-2 gene expression in granulosa cells of preovulatory and growing follicles. In contrast, cell types expressing RGS-2 protein were theca cells regardless of hCG treatment. Like in vivo, treatment of preovulatory granulosa cells with LH in vitro stimulated RGS-2 levels within 1 h. Interestingly, GnRH antagonist II enhanced the stimulatory action of LH. Conclusion: The present study demonstrates the LH/hCG induction of RGS-2 in preovulatory granulosa cells and suggests a role of RGS-2 in Gq protein signaling pathway during ovulation.

Erk AND RETINOIC ACID SIGNALING PARTICIPATE IN THE SEGREGATION AND PATTERNING OF FIRST ARCH DERIVED MAXILLA AND MANDIBLE (Erk와 retinoic acid의 제1인구둥 패터닝 조절)

  • Park, Eun-Ju;Tak, Hye-Jin;Park, Eun-Ha;Baik, Jeong-Mi;Zhengguo, Piao;Lee, Sang-Hwy
    • Maxillofacial Plastic and Reconstructive Surgery
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    • v.31 no.2
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    • pp.103-115
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    • 2009
  • In vertebrates, the face is mainly formed with neural crest derived neural crest cells by the inherent programs and the interactive environmental factors. Extracellular signaling-regulated kinase (Erk) is one of such programs to regulate the various cellular functions. And retinoic acid (RA) also plays an important role as a regulator in differentiation process at various stages of vertebrate embryogenesis. We wanted to know that the segregation as well as the patterning of maxillary and mandibular structure is greatly influenced by the maxillomandibular cleft (MMC) and the failure of this development may result in the maxillomandibular fusion (syngnathia) or other patterning related disorder. It has been well documented that the epithelium at this cleft region has significant expression of Fibroblast growth factor (Fgf) 8, and it is essential for the patterning of the first arch derived structures. By the morphological, skeletal, cell proliferation and apoptotic, and hybridization analysis, we checked the effects of Erk inhibition and/or RA activation onto MMC and could observe that Erk and RA signaling is individually and synergically involved in the facial patterning in terms of FGF signaling pathway via Barx-l. So RA and Erk signaling work together for the MMC patterning and the segregation of maxilla-mandible by controlling the Fgf-related signaling pathways. And the abnormality in MMC brought by aberrant Fgf signaling may result in the disturbances of maxillary-mandibular segregation.