| Abstrakt: |
Increased generation of reactive oxygen species is associated with endothelial dysfunction and targeting endogenous antioxidant defences has only recently been considered in the treatment of vascular diseases (Cheng et al., 2011). Offspring from gestational diabetic (GDM) pregnancies are predisposed to endothelial dysfunction in later life (Buchanan & Xiang, 2005), and endothelial cells from GDM umbilical cords exhibit abnormal nitric oxide production and reduced growth rates, indicating phenotypic changes caused by GDM (Sobrevia et al., 1995; Mann et al., 2003). To investigate the underlying mechanisms, we examined redox regulation in human umbilical vein endothelial cells (HUVEC) from GDM (n=40) and normal (n=47) pregnancies cultured in M199 (with 20% or 1% FCS). Mito-chondrial superoxide production measured by luminescence in GDM HUVEC was significantly higher than normal controls (210 ± 22 vs 81 ± 6 mean light units, mean ± SEM, n=6, P<0.001) whereas basal DNA fragmentation was comparable between groups. However, GDM HUVEC were more susceptible to the lipid peroxidation (4-hydroxynonenal, 20μmol/L) induced DNA fragmentation (n=5, P<0.05). Basal and HNE stimulated levels of glutathione (GSH), a key intracellular antioxidant, were also lower in GDM HUVEC. Moreover, HNE induced activation of the redox sensitive transcription factor NF-E2 related factor 2 (Nrf2), which mediates adaptive increases in GSH synthesis (Mann et al., 2003), was suppressed in GDM (n=5, p<0.05). Deficits in Nrf2 mediated redox signalling were further confirmed by abrogated HNE induced mRNA expression of the cystine/glutamate transporter (xCT) and phase II defence enzyme NAD(P)H quinone oxidoreductase 1 (NQO1) (n=6, P<0.05 and P<0.01 respectively). Nrf2 silencing in normal HUVEC partly reproduced the phenotypic changes of GDM HUVEC. To examine whether Nrf2 in GDM cells is epigenetically silenced, methylation of CpG islands in the promoter region of Nrf2 was measured by bisulphate restriction analysis. Although we could not detect differences in methylation between normal and GDM cells, protein expression of DJ-1, a known cytosolic Nrf2 stabilizer, was markedly reduced in GDM HUVEC (n=7, P<0.01). In summary, an altered vascular phenotype in utero may contribute to the increased risk of type 2 diabetes and cardiovascular disease in the offspring of gestational diabetic mothers in later life. [ABSTRACT FROM AUTHOR] |
