Browsing by Author "Guzman, Leda"

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    Analytical detection of immunoglobulin heavy chain gene rearrangements in gastric lymphoid infiltrates by peak area analysis of the melting curve in the LightCycler System
    (ELSEVIER SCIENCE INC, 2007) Retamales, Eduardo; Rodriguez, Luis; Guzman, Leda; Aguayo, Francisco; Palma, Mariana; Backhouse, Claudia; Argandona, Jorge; Riquelme, Erick; Corvalan, Alejandro
    Because it is difficult to differentiate gastric mucosa-associated lymphoid tissue (MALT) lymphoma from chronic gastritis in gastric lymphoid infiltrates, molecular detection of monoclonality through immunoglobulin heavy chain (IgH) gene rearrangements is commonly performed. However, heterogeneity in the performance and results obtained from IgH gene rearrangements has been reported. To improve the accuracy in the diagnosis of gastric lymphoid infiltrates, we developed an analytical approach based on one-peak area analysis of the melting curve in the LightCycler System. Using a training-testing approach, the likelihood ratio method was selected to find a discriminative function of 4.64 in the training set (10 gastric MALT lymphomas and 10 chronic gastritis cases). This discriminative function was validated in the testing set (five gastric MALT lymphomas, six abnormal lymphocytic infiltrates with subsequently demonstrated gastric MALT lymphomas, and six cases of chronic gastritis). All but one case of gastric MALT lymphoma, as well as abnormal lymphocytic infiltrates, clustered under 4.64, and all chronic gastritis cases clustered above 4.64. These results were validated by conventional electrophoreses confirming one or two sharp bands in cases of gastric MALT lymphomas and a smear of multiple bands in cases of chronic gastritis. Analytical detection of IgH gene rearrangement in gastric lymphoid infiltrates by one-peak area analysis correctly distinguishes gastric MALT lymphomas from chronic gastritis, even in cases with diagnosis of abnormal lymphocytic infiltrates.
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    Gold@Silica Nanoparticles Functionalized with Oligonucleotides: A Prominent Tool for the Detection of the Methylated Reprimo Gene in Gastric Cancer by Dynamic Light Scattering
    (2019) Jose Marchant, Maria; Guzman, Leda; Corvalan, Alejandro H.; Kogan, Marcelo J.
    Reprimo (RPRM) is a tumor suppressor gene involved in the development of gastric cancer. Hypermethylation of the RPRM promoter region has been found in tumor tissue and plasma samples from patients with gastric cancer. These findings suggest that circulating methylated DNA of RPRM could be a candidate for a noninvasive detection of gastric cancer. We designed a nanosystem based on the functionalization of silica coated gold nanoparticles with oligonucleotides that recognize a specific DNA fragment of the RPRM promoter region. The functionality of the oligonucleotide on the surface of the nanoparticle was confirmed by polymerase chain reaction (PCR). The nanoparticles were incubated with a synthetic DNA fragment of methylated DNA of RPRM and changes in the size distribution after hybridization were evaluated by dynamic light scattering (DLS). A difference in the size distribution of nanoparticles hybridized with genomic DNA from the KATO III gastric cancer cell line was observed when was compared with DNA from the GES-1 normal cell line. These results showed that this nanosystem may be a useful tool for the specific and sensitive detection of methylated DNA of RPRM in patients at risk of developing gastric cancer.
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    In vitro evaluation and molecular docking of QS-21 and quillaic acid from Quillaja saponaria Molina as gastric cancer agents
    (NATURE RESEARCH, 2020) Guzman, Leda; Villalon, Katherine; Jose Marchant, Maria; Elena Tarnok, Maria; Cardenas, Pilar; Aquea, Gisela; Acevedo, Waldo; Padilla, Leandro; Bernal, Giuliano; Molinari, Aurora; Corvalan, Alejandro
    The cytotoxic mechanism of the saponin QS-21 and its aglycone quillaic acid (QA) was studied on human gastric cancer cells (SNU1 and KATO III). Both compounds showed in vitro cytotoxic activity with IC50 values: 7.1 mu M (QS-21) and 13.6 mu M (QA) on SNU1 cells; 7.4 mu M (QS-21) and 67 mu M (QA) on KATO III cells. QS-21 and QA induce apoptosis on SNU1 and KATO III, as demonstrated by TUNEL, Annexin-V and Caspase Assays. Additionally, we performed in silico docking studies simulating the binding of both triterpenic compounds to key proteins involved in apoptotic pathways. The binding energies (G(bin)) thus calculated, suggest that the pro-apoptotic protein Bid might be a plausible target involved in the apoptotic effect of both triterpenic compounds. Although QA shows some antiproliferative effects on SNU1 cells cultured in vitro, our results suggest that QS-21 is a more powerful antitumor agent, which merits further investigation regarding their properties as potential therapeutic agents for gastric cancer.
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    Involvement of several transcriptional regulators in the differential expression of tfd genes in Cupriavidus necator JMP134
    (2009) Trefault, Nicole; Guzman, Leda; Perez, Heidi; Godoy, Margarita; Gonzalez, Bernardo
    Cupriavidus necator JMP134 has been extensively studied because of its ability to degrade chloroaromatic compounds, including the herbicides 2,4-dichlorophenoxyacetic acid (2,4-D) and 3-chlorobenzoic acid (3-CB), which is achieved through the pJP4-encoded chlorocatechol degradation gene clusters: tfdC(I)D(I)E(I)F(I), and tfdD(II)C(II)E(II)F(II). The present work describes a different tfd-genes expression profile depending on whether C. necator cells were induced with 2,4-D or 3-CB. By contrast, in vitro binding assays of the Purified transcriptional activator TfdR showed similar binding to both tfd intergenic regions; these results were confirmed by in Vivo Studies of the expression of transcriptional lacZ fusions for these intergenic regions. Experiments aimed at investigating whether other pJP4 plasmid or chromosomal regulatory proteins could contribute to the differences in the response of both tfd promoters to induction by 2,4-D and 3-CB showed that the transcriptional regulators from the benzoate degradation pathway, CatR I and CatR2, affected 3-CB- and 2,4-D-related growth capabilities. It was also determined that the ISJP4-interrupted protein TfdT decreased growth on 3-CB. In addition, an ORF with 34% amino acid identity to IcIR-type transcriptional regulator members and located near the tfd(II) gene cluster module was shown to modulate the 2,4-D growth capability. Taken together, these results Suggest that tfd transcriptional regulation in C. necator JMP134 is far more complex than previously thought and that it involves proteins from different transcriptional regulator families. [Int Microbiol 2009; 12(2):97-106]