Browsing by Author "Aguayo, C."

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    Differential expression of functional nucleoside transporters in non-differentiated and differentiated human endothelial progenitor cells
    (W B SAUNDERS CO LTD, 2010) Guzman Gutierrez, E.; Sandoval, C.; Nova, E.; Castillo, J. L.; Vera, J. C.; Lamperti, L.; Krause, B.; Salomon, C.; Sepulveda, C.; Aguayo, C.; Sobrevia, L.
    Extracellular adenosine removal is via human equilibrative nucleoside transporters 1 (hENT1) and 2 (hENT2) in the endothelium, thus regulating adenosine-induced revascularization and angiogenesis. Since human endothelial progenitor cells (hEPCs) promote revascularization, we hypothesize differential expression of nucleoside transporters in hEPCs. hEPCs were cultured 3 (hEPC-3d) or 14 (hEPC-14d) days. RT-PCR for prominin 1, CD34, octamer-4, kinase insert domain receptor, oxidized low-density lipoprotein (lectin-like) receptor 1 and tyrosine endothelial kinase was used to evaluate phenotypic differentiation. Flow cytometry was used to estimate CD34(+)/KDR- (non-differentiated), CD34(-)/KDR+ (differentiated) or CD34(+)/KDR+ (mixed) cell populations. Adenosine transport was measured in absence or presence of sodium, S-(4-nitrobenzyl)-6-thio-inosine (NBTI, 1-10 mu M), inosine, hypoxanthine or guanine (0.1-5 mM), hENTs protein abundance by western blot, and hENTs, hCNT1, hCNT2 and hCNT3 mRNA expression by real time RT-PCR. hEPC-3d cells were CD34(+)/KDR- compared with hEPC-14d cells that were CD34(-)/KDR+ hEPC-3d cells exhibit hENT1-like adenosine transport (NBTI-sensitive, Na+-independent), which is absent in hEPC-14d cells. hEPC-14d cells exhibit two transport components: component 1 (NBTI insensitive, Na+-independent) and component 2 (NBTI insensitive, Na+-dependent, Hill coefficient similar to 1.8), the latter resembling CNT3-like transport. hEPC-3d cells express hENT1 protein and mRNA, which is reduced (similar to 90%) in hEPC-14d cells, but instead only hCNT3 mRNA is expressed in this cell type. hENT2, hCNT1 and hCNT2 were undetectable in hEPCs. Thus, hEPCs exhibit a differential expression of hENT1 and hCNT3 functional nucleoside transporters, which could be related with its differentiation stage. (c) 2010 Elsevier Ltd. All rights reserved.
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    Bchex virulence gene of Botrytis cinerea: characterization and functional analysis
    (2011) Aguayo, C.; Riquelme, J.; Valenzuela, P. D. T.; Hahn, M.; Silva Moreno, E.
    We previously identified Bchex as a highly expressed gene during filamentous growth in Botrytis cinerea. The gene encodes the principal protein of the Woronin body and has been shown to seal septal pores in response to cellular damage. In the present study, Southern blot analysis of genomic DNA indicated that the gene exists as a single copy in the B. cinerea genome. The gene was differentially expressed during various developmental stages: expression was high in germinating conidia and the mycelial stage and lower in resting conidia and the appressorial stage. For functional analyses, homologous recombination was used to obtain a Delta Bchex knockout mutant. Growth of the mutant was strongly reduced growth in complete medium and in defined media with sucrose, fructose or pectin as the carbon source. After detached tomato leaves were inoculated with the Bchex mutant, lesion development was markedly reduced compared to the control, suggesting that Bchex participates in normal growth, germination and virulence of this fungus.