Iron content and hepcidin expression in Nos2^+/+ and Nos2^−/− hepatocytes. (A) Cellular iron content of primary Nos2^+/+ and Nos2^−/− hepatocytes was measured by atomic absorption spectrometry. Results were normalized for protein content and compared by Student’s t test (cells isolated from n = 12 individual mice per group). (B) Primary Nos2^+/+ and Nos2^−/− hepatocytes were isolated and stimulated with 100 ng/ml LPS, 10 ng/ml IL-6, 25 ng/ml Bmp-6, or 50 µM Nor5 for 6 h. Hepcidin mRNA levels were determined by qRT-PCR. Data, normalized for mRNA levels of the housekeeping gene Hprt, were compared by ANOVA using Bonferroni’s correction (n = 4 independent experiments). *, P < 0.05; ***, P < 0.001.