Reply
Authors: Leon A. Adams, Darrell H. Crawford, Tim K. St. Pierre, John K. Olynyk
Journal: Hepatology (2015)

This study was supported by a Clinical Research Grant from the Raine Medical Research Foundation and the Western Australian Government Department of Health. Resonance Health Ltd (Claremont, Australia) provided support for the magnetic resonance scans. Potential conflict of interest: Dr. St. Pierre is employed by, owns stock in, is on the speakers' bureau for, and received grants from Resonance Health. We would like to thank Ryan et al. for their interest and comments regarding our examining the impact of iron reduction by phlebotomy in patients with nonalcoholic fatty liver disease (NAFLD).1 As pointed out by Ryan et al., the potential benefit of iron treatment for NAFLD and metabolic syndrome is based on observational data and in vitro studies with very little high‐quality evidence. For example, the study referenced by Ryan et al. as being controlled was actually a single‐arm study.2 Our prospective, multicenter, randomized, controlled trial, being the largest study of this type, was unable to detect any effect of iron reduction on liver steatosis, serum markers of liver injury, lipid peroxidation, insulin resistance, or quality‐of‐health measures. Ryan et al. point out that iron reduction therapy might be expected to benefit a subgroup of NAFLD patients with NASH, iron overload and insulin resistance. Among hospital‐based cohorts of NAFLD patients, an elevated serum ferritin concentration is present in only one third of subjects, with less than half of these having nonalcoholic steatohepatitis (NASH). A hepatic iron concentration above the upper limit of normal (1.5 mg/g) is even more uncommon, being present in less than 10% of individuals overall.3 Thus, the proportion of NAFLD subjects with NASH and true iron overload represent only a small subset of the wider population of patients with NAFLD. Our inclusion criteria did not require an elevated serum ferritin, iron overload, or histologically proven (NASH) to be present. Nevertheless, half of our cohort (n = 32) had an elevated serum ferritin concentration at baseline; however, no benefit of venesection was observed in this subgroup in any endpoint, including hepatic steatosis, alanine aminotransaminase (ALT), and insulin resistance. Additionally, in our cohort, 25 (34%) and 26 (36%) subjects had NASH defined by cytokeratin (CK)18 value >275 IU/L and significant insulin resistance defined as a homeostasis model of assessment for insulin resistance >4.0 (see Table 1). Although interpretation of post‐hoc analyses needs to be performed with caution, we could detect no impact of venesection on study endpoints, apart from serum ferritin, in patients with presumed NASH or insulin resistance. Biopsies were not performed at study entry or finish, limiting the ability to examine the treatment effect on histological endpoints. However, given the lack of change in any validated serum marker of liver injury (ALT, CK18, and Hepascore) or level of steatosis, as determined by magnetic resonance imaging, we feel it would be unlikely that phlebotomy would have produced a significant histological effect. The phlebotomy intervention resulted in a significantly greater reduction in serum ferritin levels in the treatment group, compared to the control population, with the intervention group undergoing a median number of seven venesections up to a maximum of 19 sessions. Despite this, only 10 of 33 subjects in the treatment group reached a target ferritin level of <45 ng/mL, with the median ferritin level at end of treatment being 111 ng/mL. This reflects the invasiveness and inconvenience of phlebotomy treatment. No dose‐response relationship was observed between size of ferritin reduction or number of venesection sessions and change in endpoints over the duration of the study. Table 1 - Differences in End of Study – Baseline Outcomes Between Control and Venesection Groups According to Baseline Insulin Resistance HOMA ≤ 4.0 HOMA > 4.0 Characteristic Control Group (n = 30) Venesection Group (n = 18) P Value Control Group (n = 11) Venesection Group (n = 15) P Value ALT (IU/l) −1.5 (−17, 9) −11 (−27, 2) 0.3 −5 (−55, 1) −7 (−32, 9) 0.7 CK‐18 (U/l) −3.5 (−30, 12) −18 (−30, ‐.2) 0.3 −18 (−116, 111) −14 (−98, 133) 0.7 HOMA 0.1 (−0.8, 0.8) 0.2 (−0.6, 0.8) 0.7 0.3 (−7.0, 1.8) −0.4 (−2.2, 0.7) 1.0 Insulin Sensitivity Index −0.03 (−0.31, 0.87) 0.28 (−1.3, 2.6) 0.9 0.38 (−0.25, 0.68) 0.15 (−0.31, 0.76) 0.8 Ferritin (ng/ml) −99 (167) −275 (308) 0.02 −40 (299) −351 (255) 0.03 Hepatic steatosis (%) −1.1 (5.1) −1.4 (5.8) 0.8 −3.3 (8.3) −1.6 (3.4) 0.6 CK‐18 < 275 CK‐18 ≥ 275 Control Group (n = 29) Venesection Group (n = 20) P Value Control Group (n = 12) Venesection Group (n = 13) P Value ALT (IU/l) −1 (−17, 7.5) 10.5 (−23, −2.5) 0.2 −6.5 (−68, 6) −7 (−31, 13) 0.6 CK‐18 (U/l) −2 (−18, 10) −10 (−27, 7) 0.4 −66 (−173, 84) −37 (−98, 103) 0.4 HOMA −0.2 (−0.7, 0.4) −0.1 (−0.8, 0.6) 1.0 0.7 (−2.7, 1.9) 0.1 (−1.4, 1.7) 0.8 Insulin Sensitivity Index −0.03 (‐.27, .92) 0.98 (−1.36, 2.64) 0.3 0.48 (‐.31, .75) −0.08 (‐.43, .19) 0.3 Ferritin (ng/ml) −35 (57) −128 (141) 0.02 −42 (138) −147 (111) 0.05 Hepatic steatosis (%) −0.3 (4.4) −2.1 (5.6) 0.5 −3.1 (8.0) −1.0 (3.4) 0.4 In summary, our study has demonstrated phlebotomy has no significant impact on liver steatosis, liver injury, or insulin resistance across a spectrum of patients with NAFLD. As mentioned in our discussion, we agree with Ryan et al. that we have not excluded a therapeutic effect of phlebotomy on the small subgroup of NAFLD patients with histological evidence of NASH, significant hepatic iron overload, and insulin resistance. Overall, this subgroup represents a minority of subjects with NAFLD, and dedicated adequately powered randomized, controlled intervention trials examining venesection in this population remain to be performed.