Fig 5

A. The relative amounts of NO released from NOR-4 in the RPMI-1640 medium was measured using the DAF-2 fluorescence assay method in relation to the incubation times. A.U., arbitrary unit. B. Live or dead C. sinensis adults were incubated for 24 h in the RIMI-1640 medium supplemented with NOR-4 and DAF-2 under 1% or 20% oxygen conditions. The fluorescence values of the conditioned media, which were measured using the DAF-2 method, were serially subtracted from those of NOR-4- and DAF-2-only media (n = 3, mean ± SD). The percentages on the histograms of the live-worm groups indicate the degree of fluorescence reduction compared to those in the dead-worm groups. ^☆P < 0.01; ^★P < 0.001. The results of western blot analysis of the conditioned media (20 μL/well) with an anti-CsMb1 antibody are shown in the inset image. C. The expression levels of the globin genes were examined in the C. sinensis worms described in panel B by performing qRT-PCR. The fold increases were calculated by comparing the values of the experimental groups with those in the 20% worm-only control group (n = 3, mean ± SD). ^○P < 0.05; ^☆P < 0.01.