Editorial: NK cell-based cancer immunotherapy

En este tema de investigación, hemos recopilado varios artículos que ponen de relieve el gran potencial que las células NK exhiben como una herramienta eficaz en la inmunoterapia de cáncer.In this research topic, we have collected several articles that highlight the exciting potential that NK cells exhibit as an effective tool in cancer immunotherapy.• BIOEF-EiTB Maratoia Pediatric Cancer. Beca BIO13/CI/011, para Francisco Borrego Rabasco • BIOEF-EiTB Maratoia Transplantation. Beca BIO14/TP/001, para Susana Larrucea Bilbao • Ministerio de Economía y Competitividad. Beca SAF2013-46161-R, para Raquel Tarazona Rabasco • Ministerio de Salud. Beca PI13/02691, para Rafael Solana LarapeerReviewe

Role of transcription factor Sp1 and CpG methylation on the regulation of the human podocalyxin gene promoter

BACKGROUND: Podocalyxin (podxl) is a heavily glycosylated transmembrane protein mainly found on the apical membrane of rat podocytes and also in endothelial, hematopoietic, and tumor cells. Despite of its interest no much is known about the transcriptional regulation of podxl in different cells. Thus, we aimed at studying the functional features of the 5'-regulatory region of the human Podxl gene. RESULTS: The promoter region of the human Podxl gene has been cloned and its structure and function were analyzed. The primary DNA sequence is rich in G+C and is devoid of TATA or CAAT boxes. The sequence contains recognition sites for several putative transcription factors; however, the basic promoter activity seems to rely entirely on Sp1 transcription factor since supershift analysis was positive only for this factor. The region encompassed by 66 to -111 nts conferred the minimal transcriptional activity that increases as the number of Sp1 sites augmented with the length of the promoter fragment. In Sp1-lacking insect cells the Podxl promoter constructs showed activity only if cotransfected with an Sp1 expression plasmid. Finally, mutation of the Sp1 sites reduced the promoter activity. We analyzed whether methylation of the CpG dinucleotides present in the first ~600 nts of the promoter region of Podxl could explain the variable rates of expression in different types of cells. Inactivation of methyltransferases by 5'-aza-2'deoxicitidine showed a dose-dependent increase in the podxl content. Moreover, in vitro methylation of the promoter constructs -111,-181 and -210 led to an almost complete reduction of the promoter activity. A correlation was found between the degree of methylation of the CpG promoter dinucleotides and the rate of podxl expression in different cell lines. CONCLUSION: Our results indicate that transcriptional regulation of Podxl is supported primarily by Sp1 site(s) and that DNA-methylation of the CpG promoter islands contributes to control the tissue specific expression of podxl

Estudio del mecanismo de adhesión celular mediado por el factor J

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 8-7-199

NK Cell-Based Cancer Immunotherapy

Natural killer (NK) cells are innate lymphoid cells that have a significant role in regulating the defenses against cancer development and certain viral infections. They are equipped with an array of activating and inhibitory receptors that stimulate or diminish NK cell activity, respectively. Inhibitory receptors include, among others, the MHC class I ligands killer cell immunoglobulin-like receptors (KIR) in humans, and members of the Ly49 family of receptors in mice, and CD94/NKG2A. Activating receptors include cytokine and chemokine receptors, and those that interact with ligands expressed on target cells, such as the natural cytotoxicity receptors or NCRs (NKp30, NKp44 and NKp46), NKG2D, CD244 and DNAM-1. In addition, NK cells express Fc?RIIIA or CD16, the receptor that exerts antibody-dependent cell mediated cytotoxicity (ADCC). NK cells also express the death ligands FasL and TRAIL. The killing or sparing of target cells depends on the integration of distinct signals that originate from NK cell receptors. NK cells spare healthy cells that express normal levels of MHC class I molecules and low amounts of stress-induced self-molecules, whereas they kill target cells that down-regulate MHC class I molecules and/or up-regulate stress-induced self-molecules. The latter are common signatures of virus-infected cells and tumors. All the accumulated knowledge on NK cell biology, along with many clinical observations, is driving multiple efforts to improve the arsenal of NK cell-based therapeutic tools in the fight against malignant diseases. Indeed, NK cell-based immunotherapy is becoming a promising approach for the treatment of many cancers. It is well known that NK cells have a significant role in the anti-tumor effect of therapeutic antibodies that use ADCC as a mechanism of action. In addition to this, administration of autologous and allogeneic NK cells after activation and expansion ex vivo is used in the treatment of cancer. Moreover, adoptive transfer of NK cell lines has been tested in humans, and genetically modified NK cells expressing chimeric antigen receptors are being studied in preclinical models for potential use in the clinic

Competition between normal [674C] and mutant [674R] subunits: role of the molecular chaperone BiP in the processing of GPIIb-IIIa complexes

8 páginas, 8 figuras -- PAGS nros. 2640-2647This work aimed at investigating the function of the [C674R] mutation in GPIIb that disrupts the intramolecular 674 to 687 disulfide bridge. Individuals heterozygous for this mutation show a platelet GPIIb-IIIa content approximately 30% of normal controls, which is less than expected from one normal functioning allele. Coexpression of normal [674C]GPIIb and mutant [674R]GPIIb with normal GPIIIa produced a [674R]GPIIb concentration-dependent inhibition of surface exposure of GPIIb-IIIa complexes in Chinese hamster ovary (CHO) cells, suggesting that [674R]GPIIb interferes with the association and/or intracellular trafficking of normal subunits. Mutation of either 674C or 687C had similar effects in reducing the surface exposure of GPIIb-IIIa. However, substitution of 674C for A produced a much lesser inhibition than R, suggesting that a positive-charged residue at that position renders a less efficient subunit conformation. The mutant [674R]GPIIb but not normal GPIIb was found associated with the endoplasmic reticulum chaperone BiP in transiently transfected CHO cells. BiP was also found associated with [674R]GPIIb-IIIa heterodimers, but not with normal GPIIIa or normal heterodimers. Overexpression of BiP did not increase the surface exposure of [674R]GPIIb-IIIa complexes, indicating that its availability was not a limiting step. Platelets from the thrombasthenic patient expressing [674R]GPIIb-IIIa were found to bind soluble fibrinogen in response to physiologic agonists or dithiothreitol treatment. Thus, the [674R]GPIIb mutation leads to a retardation of the secretory pathway, most likely related to its binding to the molecular chaperone BiP, with the result of a defective number of functional GPIIb-IIIa receptors in the cell surfaceSupported in part by grants from the Dirección General de Investigación Cientı́fica y Técnica (DGICYT PB97-1240 and DGICYT PM97-0016), Fondo de Investigaciones Sanitarias (96/2014), and Comunidad Autónoma de Madrid 08.4/0031/1998, and by a grant-in-aid from the Agencia Española de Cooperación Internacional (AECI, n/ref 99CN0009). E.G.A.-S. received a fellowship from the Fundación ArecesPeer reviewe

Agonist-induced aggregation of Chinese hamster ovary cells coexpressing the human receptors for fibrinogen (integrin alphaIIbbeta3) and the platelet-activating factor: dissociation between adhesion and aggregation

9 páginas, 10 figuras, 1 tabla -- PAGS nros. 2819-2827This work reports the establishment of a Chinese hamster ovary (CHO) cell line stably coexpressing the human αIIbβ3 integrin and the platelet-activating factor receptor (PAFR). These cells aggregate in response to PAF in a Ca++, αIIbβ3, and soluble fibrinogen (Fg)–dependent manner that is prevented by PAF antagonists or αIIbβ3 blockade. The aggregating response is accompanied by enhanced binding of fibrinogen and the activation-dependent IgM PAC1. This model has permitted us to identify, for the first time, intracellular signals distinctly associated with either αIIbβ3-mediated adhesion or aggregation. Nonreceptor activation of protein kinase C (PKC) by phorbol ester produced cellular adhesion and spreading onto immobilized Fg, but it was not a sufficient signal to provoke cellular aggregation. Moreover, inhibition of PKC impeded the PAF stimulation of cellular adhesion, whereas the aggregation was not prevented. The PAF-induced cellular aggregation was distinctly associated with signaling events arising from the liganded Fg receptor and the agonist-induced stimulation of a calcium/calmodulin-dependent signaling pathway. Sustained tyrosine phosphorylation of both mitogen-activated protein kinase (MAPK) and an approximately 100-kd protein was associated with the PAF-induced aggregation, whereas phosphorylation of focal adhesion kinase (FAK) was preferably associated with cellular adherence and spreading onto immobilized Fg.Supported in part by grants from the Dirección General de Investigación Cientı́fica y Técnica (DGICYT PB97-1240, SAF 2000-0127 and DGICYT PM97-0016), Fondo de Investigaciones Sanitarias (96/2014), and Comunidad Autónoma de Madrid (08.4/0031/1998). L.S. was supported by a grant-in-aid from the Agencia Española de Cooperación Internacional (AECI)Peer reviewe

alpha-Adrenergic-mediated activation of human reconstituted fibrinogen receptor (integrin alphaIIbbeta3) in Chinese hamster ovary cells

9 páginas, 8 figuras -- PAGS nros. 1368-1376This work reports the functional studies of CHO cells coexpressinga-adrenergic (aAR) and human fibrinogen (Fg) receptors(integrin αIibβ3). Stimulation of these cells with a-agonistsproduced a transient rise in the free cytosolic calcium (Ca++)accompanied by enhanced binding to soluble Fg, and theseeffects were prevented by specific aAR antagonists. The a-adrenergic-induced activation of αIibβ3 in CHO-αIibβ3-aARincreased the rate of adhesion and extension of cells onto Fgcoated plates, and also induced a soluble Fg- and αLIIb&abeta;3-dependentformation of cell aggregates, whereas no effects wereobserved by the stimulation of CHO-αIibβ3 cells.a-Adrenergic antagonists, the ligand mimetic peptide RGDS, pertussis toxin(PTX), or EDTA, they all prevented the a-adrenergic stimulationof adhesion and aggregation. However, inhibition of PKCprevented the a-adrenergic stimulation of cell adherence,whereas blocking the intracellular Ca++ mobilization impededthe stimulation of cell aggregation.The α-adrenergic activationwas associated with phosphorylation of a protein of ~100 kDaand proteins of the MAPK family. The former was selectivelyphosphorylated by α-adrenergic stimulation whereas the latterwere phosphorylated by the binding of cells to Fg and markedlyintensified by a-adrenergic stimulationThis work has been supported in part by grants from the Direccion General de Investigacion (SAF 2000-0127, BMC2002-01053 and BMC2003-01409), Fondo de Investigaciones Sanitarias (FIS-PI021263) and Comunidad de Madrid (08.4/0029.1/2003). Nora Butta is recipient of a tenure track grant Ramon y Cajal from the Spanish Ministry of Science. Susana Larrucea was supported by a postdoctoral fellowships from the Comunidad de Madrid (08.4/0015.1/2001) and Sonia Alonso by a predoctoral fellowship from the Gobierno Vasco (BF101-40)Peer reviewe

Release of podocalyxin into the extracellular space. Role of metalloproteinases

7 páginas, 8 figuras, 3 figuras S, 1 tabla S -- PAGS nros. 1504-1510Podocalyxin (PODXL) is a type I membrane mucoprotein abundantly presented in the epithelial cells (podocytes) of kidney glomeruli where it plays an important role in maintaining the plasma filtration. PODXL is also expressed in other types of cells but its function is ignored. A recombinant soluble fragment of the PODXL ectodomain modifies the signaling of the membrane bound PODXL. Based on this antecedent, we aimed at investigating whether PODXL could be cleaved and released into the extracellular space as a soluble peptide. In this study, we used a fusion protein of human PODXL and green fluorescent protein expressed in CHO cells (CHO-PODXL-GFP) and a human tumor cell (Tera-1) inherently expressing PODXL. PODXL was detected by wide-field microscopy in the Golgi, the plasma membrane and in a vesicular form preferentially located at the leading edges of the cell and also progressing along the filopodium. We detected PODXL in the insoluble and soluble fractions of the extracellular medium of CHO-PODXL-GFP cells. Stimulation of protein kinase C (PKC) by Phorbol-12-myristate-13-acetate (PMA) enhanced the release of PODXL to the extracellular space whereas this effect was prevented either by inhibitors of PKC or specific inhibitors of matrix metalloproteinases. It is concluded that intact PODXL is released to the extracellular space as a cargo of microvesicles and also as a soluble cleaved fragment of ectodomainThis work was supported by grants from the Spanish Ministry of Science and Innovation (SAF2005-01261 and SAF2007-61701). The CIBER de Enfermedades Raras (CIBERER) is an initiative of the ISCIIIPeer reviewe

紀伊国 高野山 銀1匁

日本銀行金融研究所所蔵藩札等資料番号：ⅢAエドb1-57-1イ-43科学研究費助成事業（研究成果公開促進費）で電子化を実施データベースの名称：藩札等に関する統合データベース課題番号：20HP8030利用に関するお問い合わせ：画像の転載（出版物・HP等）に際しては、日本銀行貨幣博物館への申請手続きが必要です。詳しくは貨幣博物館ホームページ（http://www.imes.boj.or.jp/cm/service/）をご覧ください