INTRODUCCIÓN: REVISIÓN CRITICA DEL PROBLEMA
INTRODUCCIÓN: REVISIÓN CRITICA DEL PROBLEMA
INTRODUCCIÓN: REVISIÓN CRITICA DEL PROBLEMA
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g) Data analysis<br />
ANEXOS<br />
Data are expressed as mean ± s.e.m. The mRNA levels of NOX4, P22PHOX and<br />
NOS3 were expressed as the fold change of each treatment with respect to HAS<br />
treatment. The activation of NF-kB was expressed as the fold change respect to<br />
HAS treatment of the optical density ratio between nucleus and cytoplasm. ROS<br />
production was expressed as the ratio between the cytochrome c reduction rate of<br />
each treatment and the HSA treatment. Two-group comparisons were analyzed by<br />
the 2-tailed Student’s t-test. Multiple comparisons were evaluated by ANOVA<br />
followed by Tukey test. Statistical significance was considered for p < 0.05.<br />
Results<br />
a) Role of the transcription factors in NOX4 and P22PHOX expression<br />
induced by gHSA.<br />
Firstly, we studied the role of two transcription factors, NF-κB and AP-1, which<br />
have been previously related to an expression change of NOX4 and P22PHOX.<br />
The expression of NOX4 and P22PHOX were quantified in HUVEC treated with<br />
HSA or gHSA (25 μg/mL) for 4 hours. The expression levels after these treatments<br />
were measured both in the presence and in the absence of BAY11-7082 (2μM) or<br />
SP600125 (20μM), NF-κB and AP-1 inhibitor, respectively, during 30 minutes<br />
before and during the 4 hours of treatment. Like in our previous work 5 , gHSA<br />
increased NOX4 and P22PHOX expression with respect to HSA (1.25 ± 0.10<br />
NOX4 fold change for gHSA, p < 0.05, n = 13; Figure 1a; 1.12 ± 0.05 P22PHOX<br />
fold change for gHSA, p < 0.05, n = 10; Figure 1b).<br />
The NF-κB inhibitor, BAY11-7082, suppressed the gHSA-enhanced NOX4<br />
expression (1.25 ± 0.10 NOX4 fold change for gHSA vs. 0.83 ± 0.10 fold change<br />
for gHSA+BAY, p > 0.05, n = 5; Figure 1a). This inhibitor tended to reduce<br />
P22PHOX expression but this reduction was not statistically significant (1.12 ±<br />
0.05 P22PHOX fold change for gHSA vs. 0.91 ± 0.16 fold change for gHSA+BAY,<br />
p > 0.05, n = 5; Figure 1b). On the other hand, the treatment with AP-1 inhibitor<br />
SP600125 markedly increased NOX4 expression, independently of the treatment<br />
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