Reduced NO synthesis and eNOS mRNA expression in endothelial cells from newborns with a strong family history of type 2 diabetes.
BACKGROUND
A deficient synthesis of nitric oxide (NO) may play a role in the early endothelial dysfunction of healthy humans with a strong family history of type 2 diabetes (DM2). In this study, we evaluate the intracellular synthesis of NO and the expression of eNOS transcripts in human umbilical vein endothelial cells (HUVECs), exposed to high glucose concentrations, of healthy newborns with (experimental) and without (control) a strong family history of DM2.


METHODS
HUVECs were incubated in M-199 culture media (containing a 5 mmol/L physiological glucose concentration) or supraphysiological glucose concentrations (15 or 30 mmol/L), for 48 h. Flow cytometry, reactive of Griess and RT-PCR were used to determine intracellular NO synthesis, presence of NO metabolites, and expression of eNOS, GLUT1 or p53 transcripts.


RESULTS
NO synthesis in experimental HUVECs showed a progressive reduction in the presence of increasing glucose concentration (11% for 5 mmol to 8% for 30 mmol; p < 0.01), whereas control HUVECs showed an increase in NO synthesis (3% for 5 mmol to 31% for 30 mmol; p < 0.001). In experimental HUVECs, we found a diminished expression of eNOS and p53, and also an enhanced expression of GLUT1 mRNA transcripts. Control HUVECs showed an increase in eNOS, and no modifications in p53 or GLUT1 mRNA transcripts.


CONCLUSIONS
Our results show how HUVECs, isolated from healthy newborns with a strong family history of DM2, have an abnormal intracellular synthesis of NO and an impaired expression of eNOS, GLUT1 and p53 genes, all associated with NO synthesis.
Alvarado-Vásquez
Noé
N
Departamento de Bioquímica, Instituto Nacional de Enfermedades Respiratorias, México. nnooee@gmail.com
Zapata
Estrella
E
Alcázar-Leyva
Susana
S
Massó
Felipe
F
Montaño
Luis F
LF
D000328:Adult; D002455:Cell Division; D003924:Diabetes Mellitus, Type 2 / Q000235:genetics*; D004730:Endothelium, Vascular / Q000201:enzymology*; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007231:Infant, Newborn; D008487:Medical History Taking; D008928:Mitochondria / Q000201:enzymology; D009569:Nitric Oxide / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000235:genetics*; D011247:Pregnancy; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014471:Umbilical Veins / Q000201:enzymology
D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III; D005947:Glucose