Euthyroidism was defined as TSH, free T4 and free T3 levels within the reference range as provided by the manufacturer. We additionally excluded subjects with positive anti-thyroid peroxidase auto-antibodies. Applying these selection criteria, 1854 subjects were eligible for the current analyses. Patient characteristics, including age, sex, alcohol use, smoking status, body mass index, systolic and diastolic blood pressure, and waist circumference were obtained. Blood was drawn after an overnight fasting period for measurement of free T4, free T3, TSH, bilirubin, glucose, insulin, total cholesterol, high density lipoprotein cholesterol, triglycerides, aspartate aminotransferase, and alanine aminotransferase. To our knowledge, this is the first report on an independent positive relationship of total bilirubin with free T4 in a large group of euthyroid, non-diabetic individuals recruited from the general population. Of note, multivariable linear regression analyses demonstrated a significant positive modification of the effect of free T4 on bilirubin by insulin resistance as quantified by HOMAIR. This effect-modification remained essentially unaltered after controlling for potential confounders, including smoking, alcohol, transaminases, and total cholesterol. Our results, therefore, are in concert with the hypothesis that low-normal thyroid function may confer lower circulating bilirubin levels, especially in insulin resistant individuals. In addition, the effect of free T4 on bilirubin was modified by the HDL cholesterol concentration. We recently Ganoderic-acid-G documented a positive relationship of circulating levels of bilirubin with free T4 in euthyroid T2DM subjects. In that report, bilirubin was not significantly correlated with free T4 in non-diabetic subjects, possibly due to the limited number of participants. Moreover, it could not be determined what was the driving force behind this association, i.e. insulin resistance or hyperglycemia resulting from b-cell dysfunction. Our previous findings, therefore, endorsed our main rationale to investigate the association between bilirubin and free T4 in a large group of non-diabetic subjects. In the current study, we found bilirubin to be more strongly associated with HOMA-IR and insulin than with Albaspidin-AA glucose in age- and sex-adjusted analysis. Furthermore, a positive modification of the effect of free T4 on bilirubin by HOMA-IR was observed in such a way that the effect of free T4 on bilirubin was most pronounced in the most insulin resistant subjects. This effect modification was not observed with plasma glucose, raising the possibility that insulin 
resistance rather than hyperglycemia per se could represent a mechanism linking low bilirubin to low normal thyroid function. In view of the strong anti-oxidative properties of the HDL fraction, and the modification of HDL antioxidative capacity by thyroid function, it is also of potential relevance that the effect of free T4 on bilirubin was modified by HDL cholesterol. The interaction of free T4 with insulin resistance on bilirubin may have pathophysiological relevance since lower thyroid functional status, impaired insulin sensitivity, and low bilirubin are all characterized by enhanced oxidative stress.