The concept of a hepato-splenic axis has recently been put forward. We aimed to investigate whether hepatic and splenic metabolic activities are linked, and if splenic metabolic activity is increased in non-alcoholic fatty liver disease (NAFLD). Blood clearance rates of phosphorylated F-fluorodeoxyglucose were measured in the spleen and liver from dynamic PET using Gjedde-Patlak-Rutland graphical analysis and abdominal aorta for input function in 59 patients undergoing routine PET/CT. Plot gradient (Ki), which represents blood clearance, was divided by intercept (V(0)), which represents tissue FDG distribution volume, and multiplied by blood glucose to give glucose uptake rate per unit tracer distribution volume (MRglu). In addition, liver-to-spleen raw count rate ratio was plotted against time, and gradient (b) divided by intercept (A) to obtain hepatic-to-splenic blood clearance ratio independent of aortic input function. Hepatic steatosis was inferred when hepatic CT density was ≤40 HU. There was no difference in splenic MRglu between 8 patients with inactive lympho-proliferative disease (LPD) as identified by negative PET/CT, 25 with non-haematological malignancy and 13 with normal PET/CT. It was significantly increased in 13 with active LPD, who were therefore excluded, along with 3 more with type-2 diabetes mellitus. Splenic MRglu was higher in patients with hepatic steatosis (4.0±1.6; n = 12) than without (2.6±1.7 μmol/min/100 ml; P = 0.02) and correlated inversely with hepatic CT density (r = -0.49; P<0.001). Hepatic and splenic Ki/V(0) correlated (r = 0.52; P<0.01) in 22 patients in whom the correlation coefficient between b/A and hepatic-to-splenic Ki/V(0) ratio was 0.99 and in whom, therefore, input function errors in graphical analysis could be discounted. In men, splenic longitudinal diameter correlated significantly with hepatic CT density (r = -0.35; P = 0.046), hepatic MRglu (r = 0.44; P = 0.005) and splenic MRglu (r = 0.35; P = 0.046). Splenic Ki/V(0) correlated positively with blood glucose, suggesting sensitivity to insulin. We conclude that hepatic and splenic metabolic activities are linked and that a speculative mechanism, which deserves further investigation, is shared insulin sensitivity. Splenic MRglu and spleen size are increased in NAFLD.