title: PP134. l-Arginine supplementation induces the expression of genes related to antioxidant defense in cultures of human vascular endothelial cells: Protective role for preeclampsia.
abstract: BACKGROUND
  Recently, it has been proposed that supplementation with l-Arginine reduces the incidence of preeclampsia in high risk women, but the molecular mechanisms involved in the protective effect need to be determined. In addition, a critical role of l-Arginine in endothelial cell survival during oxidative stress, and the participation of neutrophils in the induction of oxidative stress during preeclampsia have been suggested.
  
  
  OBJECTIVE
  To address if supplementation with l-arginine provides antioxidant defense in human vascular endothelial cells.
  
  
  METHODS
  Human vascular endothelial cells (HUVECs) were isolated from umbilical cord veins obtained from healthy women underwent cesarean sections at term, with no evidence of hypertension disorders through the pregnancy. HUVECs were cultured in EndoGro media with LS supplement kit and 1% antibiotic with (n=10) or without 200uM l-Arginine (n=10). Confluent HUVECs were stimulated with neutrophils activated with 50umol/L arachidonic acid (1:16 ratio of neutrophil/cells). After incubation, cells were rinsed in PBS and harvested for RNA and protein extraction. Reverse transcription was performed using the RT(2) First Strand kit, and expression gene profiling was generated using the RT(2) Profiler PCR Array Human Oxidative Stress and Antioxidant Defense that includes the expression profile of 84 genes related to the oxidative pathway. Expression results were analyzed with the RT(2) Profiler PCR Array Data Analysis Template v3.0 and two different lists of fold change in gene expression were generated: (1) HUVEC+neutrophils vs HUVEC+l-Arginine + neutrophils and (2) HUVEC vs HUVEC+neutrophils. Validation of the expression assays was performed using western blots or ELISAS for proteins expressed by selected genes.
  
  
  RESULTS
  Fold up- or down gene regulation are shown in Table 1. Forty six genes involved in oxidative stress defense were significantly up-regulated in HUVECs supplemented with l-arginine when were exposed to neutrophils. Interestingly, almost the same genes were down-regulated in non-supplemented HUVECs after neutrophil exposure.
  
  
  CONCLUSIONS
  Supplementation with l-Arginine upregulates the expression of genes related to antioxidant defense in primary cultures of endothelial cells. This finding provides a novel insight about the molecular mechanisms involved in the protective role of l-Arginine during preeclampsia.
authors.affiliation: Department of Biochemistry, Mexico City, Mexico.
authors.forename: A
authors.initials: A
authors.lastname: Espejel-Nuñez
authors.affiliation: Department of Biochemistry, Mexico City, Mexico.
authors.forename: A
authors.initials: A
authors.lastname: Flores-Pliego
authors.affiliation: Labor Room , Instituto Nacional de Perinatologia, Mexico City, Mexico.
authors.forename: S
authors.initials: S
authors.lastname: Espino Y Sosa
authors.affiliation: Department of Biochemistry, Mexico City, Mexico.
authors.forename: A
authors.initials: A
authors.lastname: Guzman-Grenfell
authors.affiliation: Department, Virginia Commonwealth University Medical Center, Richmond, VA, United States.
authors.forename: S W
authors.initials: SW
authors.lastname: Walsh
authors.affiliation: Department of Infectology, Instituto Nacional de Perinatologia, Mexico City, Mexico.
authors.forename: G
authors.initials: G
authors.lastname: Estrada-Gutierrez
authors.affiliation: Instituto de Ecologia, UNAM, Mexico City, Mexico.
authors.forename: R E
authors.initials: RE
authors.lastname: Cappello