CORRESPONDENCE Ethnic Differences in Viral Dominance Patterns in Patients with Hepatitis B Virus and Hepatitis C Virus Dual Infection To the Editor: We read with great interest the article entitled ‘‘Ethnic Differen- ces in Viral Dominance Patterns in Patients with Hepatitis B Virus and Hepatitis C Virus Dual Infection,’’ recently published by Nguyen et al. in this journal. 1 The study was designed to evaluate and compare the demo- graphic, clinical, and viral characteristics of multiethnic patients infected by hepatitis virus admitted at two large liver centers in the United States. The investigators defined the study design as a matched case-control study, in which the case group included patients with hepatitis B virus (HBV) and hepatitis C virus (HCV) dual infection, and the control group comprised patients with HBV monoinfection. However, in our opinion, there is an important methodological issue in the reported data: The main conclusion of the article was not about the comparison between cases and controls, but rather it underscored a secondary analysis with cases only in which ethnic differences were examined in terms of viral dominance patterns among HBV and HCV dual-infected patients. It is our opinion that the investigators deviated from the scope of the case-control study originally proposed, because it is clear that the investigation was not delineated to test the hypothesis that ended up becoming the core conclusion, including the title of the article. Therefore, the investigators’ conclusions regarding viral dominance were not consistent with the a priori study aims proposed. GABRIELE ROCKENBACH, M.Sc. ALEXANDRE JOSE ´ DE MELO NETO, M.D. NE ˆ MORA TREGNAGO BARCELLOS, M.D., Ph.D. FERNANDO HERZ WOLFF , M.D., Ph.D. Postgraduate Studies in Epidemiology School of Medicine Universidade Federal do Rio Grande do Sul Ramiro Barcelos, 2600, Rio Grande do Sul, Brazil Reference 1. Nguyen LH, Ko S, Wong SS, Tran PS, Trinh HN, Garcia RT, et al. Ethnic differences in viral dominance patterns in patients with hepatitis B virus and hepatitis C virus dual infection. HEPATOLOGY 2011;53: 1839-1845. Copyright V C 2012 by the American Association for the Study of Liver Diseases. View this article online at wileyonlinelibrary.com. DOI 10.1002/hep.24533 Potential conflict of interest: Nothing to report. Reply: We appreciate Ronkenback et al.’s interest in our study. 1 The aim of the study was to further characterize hepatitis B virus/hepatitis C virus (HBV-HCV) dual infection, including viral dominance pat- tern, which is one of the most important topics in the study of HBV- HCV dual-infected patients. In addition to examining the clinical characteristics of HBV-HCV dual-infected cases, in comparison with HBV monoinfected controls, an analysis of viral dominance was suit- ably part of our main aims, even if it did not require the comparison of HBV-HCV cases and HBV monoinfected controls. The major findings and primary conclusions of the study are that HBV-HCV dual-infected and HBV monoinfected patients generally had similar clinical characteristics, Asian ethnicity is a major independent predic- tor of HBV-dominant disease, and HCV dominance with undetect- able HBV DNA is more common in non-Asians, all of which were within the scope of our major aims. LONG H. NGUYEN, B.A. MINDIE NGUYEN, M.D., M.A.S. Stanford University School of Medicine Stanford, CA Reference 1. Nguyen LH, Ko S, Wong SS, Tran PS, Trinh HN, Garcia RT, et al. Eth- nic differences in viral dominance patterns in patients with hepatitis B vi- rus and hepatitis C virus dual infection. HEPATOLOGY 2011;53: 1839–1845. Copyright V C 2012 by the American Association for the Study of Liver Diseases. View this article online at wileyonlinelibrary.com. DOI 10.1002/hep.25719 Potential conflict of interest: Nothing to report. Identification of Endogenous Normalizers for Serum MicroRNAs by Microarray Profiling: U6 Small Nuclear RNA Is Not a Reliable Normalizer To the Editor: We read with great interest the results that the expression of circulating microRNA (miRNA) miR-122 was substantially higher in acetaminophen-induced acute liver injury (APAP-ALI) patients, compared to healthy controls, using quantitative real- time polymerase chain reaction (PCR) assays with U6 small nu- clear RNA (snRNA) as an internal control, as reported by Starkey Lewis et al. 1 However, serum miRNA expression profiles gener- ated from a large number of human samples by our laboratory indicate that circulating U6 snRNA is not a reliable internal normalizer. The accuracy of circulating miRNA expression analysis critically depends on proper normalization of the data. Endogenous normal- izer specific for circulating miRNAs have not yet been well defined. Although some cell/tissue miRNA normalizers, including U6 and miR-16, have been used in circulating miRNA data analyses, recent studies suggest that cell/tissue normalizers may not serve as circulat- ing normalizers. 2 To identify the miRNAs with the most stable expression in human serum, we have evaluated 117 serum miRNA expression profiles from young, aging, and different disease condi- tions using a real-time PCR array system, based on a global expres- sion mean normalization strategy. 3 Of 332 miRNAs detected in se- rum, 58 displayed consistent expression across all samples (Fig. 1A). 1640