Tetrahydrobiopterin-dependent preservation of nitric oxide–mediated endothelial function in diabetes by targeted transgenic GTP–cyclohydrolase I overexpression
J. Clin. Invest. Nicholas J. Alp, et al. 112:725 doi:10.1172/JCI17786 [Go to this article.]

Figure 2
(a) Evaluation of transgenic and native GTPCH mRNA expression by RT-PCR. The top panel shows human-specific primers to detect transgenic GTPCH mRNA. A PCR product of 181 bp (arrowhead) was detected in lung and liver, and aorta from GCH-Tg but not WT littermates, and only after the RT step (RT+). The middle panel shows mouse-specific primers used to detect native GTPCH mRNA. A PCR product of 181 bp (arrowhead) was detected in lung, liver, and aorta from both GCH-Tg and WT mice. (b) Quantification of GTPCH expression by real-time fluorescent RT-PCR. Shown are means and SEMs of three samples, each quantified in triplicate. Note that arbitrary units are independent among the three graphs. In the top panel, transgenic GTPCH mRNA expression was 10-fold higher in lung than in liver and aorta among GCH-Tg animals but was not detected in WT littermates (**P < 0.01 for comparisons). In the middle panel, native GTPCH mRNA expression in liver was approximately 10-fold higher than in lung and 100-fold higher than in aorta, but similar between GCH-Tg and WT mice for each organ. In the bottom panel, total GTPCH mRNA expression in the lung was approximately 20-fold higher in GCH-Tg than in WT mice (*P < 0.05); in the liver there was no difference in GCH-Tg as compared with WT mice; in the aorta, there was a 5-fold increase in total GTPCH mRNA expression in GCH-Tg as compared with WT mice (**P < 0.01). (c) Immunoblotting with a rabbit anti-human GTPCH polyclonal antibody to detect transgenic GTPCH protein identified a specific 35-kDa band (arrowhead) in lysates from lung, liver, and aorta from GCH-Tg but not WT animals.