MiR-122 acts as a tumor suppressor in hepatocarcinogenesis in vivo

Abstract

COMMENTARY ON:Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver. Hsu SH, Wang B, Kota J, Yu J, Costinean S, Kutay H, Yu L, Bai S, La Perle K, Chivukula RR, Mao H, Wei M, Clark KR, Mendell JR, Caligiuri MA, Jacob ST,Mendell JT, Ghoshal K. J Clin Invest. 2012 Aug 1;122(8):2871–83. Copyright (2012). Reprinted with permission of American Society for Clinical Investigation.http://www.ncbi.nlm.nih.gov/pubmed/22820288Abstract: miR-122, an abundant liver-specific microRNA (miRNA), regulates cholesterol metabolism and promotes hepatitis C virus (HCV) replication. Reduced miR-122 expression in hepatocellular carcinoma (HCC) correlates with metastasis and poor prognosis. Nevertheless, the consequences of sustained loss of function of miR-122 in vivo have not been determined. Here, we demonstrate that deletion of mouse Mir122 resulted in hepatosteatosis, hepatitis, and the development of tumors resembling HCC. These pathologic manifestations were associated with hyperactivity of oncogenic pathways and hepatic infiltration of inflammatory cells that produce pro-tumorigenic cytokines, including IL-6 and TNF. Moreover, delivery of miR-122 to a MYC-driven mouse model of HCC strongly inhibited tumorigenesis, further supporting the tumor suppressor activity of this miRNA. These findings reveal critical functions for miR-122 in the maintenance of liver homeostasis and have important therapeutic implications, including the potential utility of miR-122 delivery for selected patients with HCC and the need for careful monitoring of patients receiving miR-122 inhibition therapy for HCV.ANDMicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis. Tsai WC, Hsu SD, Hsu CS, Lai TC, Chen SJ, Shen R, Huang Y, Chen HC, Lee CH, Tsai TF, Hsu MT, Wu JC, Huang HD, Shiao MS, Hsiao M, Tsou AP. J Clin Invest. 2012 Aug 1;122(8):2884–97. Copyright 2012. Reprinted with permission of AMERICAN SOCIETY FOR CLINICAL INVESTIGATION.http://www.ncbi.nlm.nih.gov/pubmed/22820290Abstract: MicroRNA-122 (miR-122), which accounts for 70% of the liver’s total miRNAs, plays a pivotal role in the liver. However, its intrinsic physiological roles remain largely undetermined. We demonstrated that mice lacking the gene encoding miR-122a (Mir122a) are viable but develop temporally controlled steatohepatitis, fibrosis, and hepatocellular carcinoma (HCC). These mice exhibited a striking disparity in HCC incidence based on sex, with a male-to-female ratio of 3.9:1, which recapitulates the disease incidence in humans. Impaired expression of microsomal triglyceride transfer protein (MTTP) contributed to steatosis, which was reversed by in vivo restoration of Mttp expression. We found that hepatic fibrosis onset can be partially attributed to the action of a miR-122a target, the Klf6 transcript. In addition, Mir122a−/− livers exhibited disruptions in a range of pathways, many of which closely resemble the disruptions found in human HCC. Importantly, the reexpression of miR-122a reduced disease manifestation and tumor incidence in Mir122a−/− mice. This study demonstrates that mice with a targeted deletion of the Mir122a gene possess several key phenotypes of human liver diseases, which provides a rationale for the development of a unique therapy for the treatment of chronic liver disease and HCC.Hepatocellular carcinoma (HCC) is the fifth most prevalent and the third leading cause of cancer death worldwide. Although early-stage tumors can be curated using surgical approaches, treatment options for advanced HCC are limited. Virus- and alcohol-induced cirrhosis represent major causes of HCC, but the molecular mechanisms underlying HCC are still only incompletely understood [[1]El-Serag H.B. Epidemiology of viral hepatitis and hepatocellular carcinoma.Gastroenterology. 2012; 142 (e1261): 1264-1273Abstract Full Text Full Text PDF PubMed Scopus (2341) Google Scholar]. Several microRNAs (miRNAs) have been implicated in hepatocarcinogenesis and may represent promising new anti-cancer targets (reviewed in [[2]Wang X.W. Heegaard N.H. Orum H. MicroRNAs in liver disease.Gastroenterology. 2012; 142: 1431-1443Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar]). miRNAs are small non-coding regulatory RNAs resulting from Dicer-mediated processing of miRNA precursors. They act by modulating target messenger RNA translation and/or stabilization and are involved in a wide variety of biological processes. miR-122, a miRNA predominantly expressed in hepatocytes, accounts for approximately 70% of liver-expressed miRNAs. Evidence from sequestrating miR-122 in vivo suggests that this miRNA is involved in cholesterol biosynthesis and liver function [[3]Esau C. Davis S. Murray S.F. Yu X.X. Pandey S.K. Pear M. et al.MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.Cell Metab. 2006; 3: 87-98Abstract Full Text Full Text PDF PubMed Scopus (1737) Google Scholar]. It may also play a role in hepatocarcinogenesis as its expression is downregulated in HCC [[4]Bai S. Nasser M.W. Wang B. Hsu S.H. Datta J. Kutay H. et al.MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.J Biol Chem. 2009; 284: 32015-32027Crossref PubMed Scopus (427) Google Scholar] and hepatocyte-specific Dicer1 knockout (KO) mice spontaneously develop HCC [[5]Sekine S. Ogawa R. Ito R. Hiraoka N. McManus M.T. Kanai Y. et al.Disruption of Dicer1 induces dysregulated fetal gene expression and promotes hepatocarcinogenesis.Gastroenterology. 2009; 136 (e2301–2304): 2304-2315Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar]. However, its exact role in hepatocarcinogenesis and the underlying molecular mechanisms had not been investigated yet.Two independent laboratories now report arresting conclusions regarding the metabolic, anti-inflammatory, and anti-tumorigenic roles of miR-122, by generating germline KO, called either Mir122 KO [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar] or Mir122a−/− [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar], and liver-specific KO (Mir122 LKO) mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. These mice developed normally and were characterized by reduced serum cholesterol levels in line with previous studies reporting antisense miR-122 sequestration in vivo [[3]Esau C. Davis S. Murray S.F. Yu X.X. Pandey S.K. Pear M. et al.MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.Cell Metab. 2006; 3: 87-98Abstract Full Text Full Text PDF PubMed Scopus (1737) Google Scholar]. In Mir122 KO and Mir122 LKO mice, total serum cholesterol was reduced by 30% with concomitant decrease in LDL and HDL while TG levels remained unchanged [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Reduced serum cholesterol (total cholesterol, VLDL and HDL) was also observed in Mir122a−/− along with reduced TG levels [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. In contrast to antisense miR-122-treated mice, these Mir122 KO mice developed steatosis and hepatitis that progressed to fibrosis and HCC and thereby displayed a phenotype resembling liver-specific Dicer1 KO mice [[5]Sekine S. Ogawa R. Ito R. Hiraoka N. McManus M.T. Kanai Y. et al.Disruption of Dicer1 induces dysregulated fetal gene expression and promotes hepatocarcinogenesis.Gastroenterology. 2009; 136 (e2301–2304): 2304-2315Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar] (Table 1). Hepatic gene expression analysis indicated that numerous genes involved in lipid metabolism were differentially regulated in LKO mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Mir122a−/− mice also demonstrated impaired expression of several proteins involved in lipid metabolism, among which microsomal triglyceride transfer protein (MTTP). Although miR-122 does not directly target MTTP messenger RNA, prolonged injection of MTTP- or miR-122-expression constructs in Mir122a−/− mice allowed restoration of MTTP levels, increase of serum lipids and reduction of hepatic steatosis, inflammation and fibrosis [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. These functional experiments indicate that lack of MTTP and/or miR-122 expression leads to liver disease. In addition, both groups validated several novel direct targets of miR-122, among which Klf6 or Agpat1, that could also account for the observed phenotype [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar].Table 1Phenotype of Mir122 knockout mice and effect of Mir122 restoration. This table summarizes the characteristics of Mir122a−/− as well as Mir122 KO and Mir122 LKO mice 6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar. n.d., not determined; mo, months; wk, week. Open table in a new tab Moreover, in both models, prolonged lack of miR-122 in mouse hepatocytes associates with HCC [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. While spontaneous tumors were 3.9-fold higher in male Mir122a−/− mice than in females [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar], in line with observed HCC incidence in humans, Hsu et al. observed equal penetration in Mir122 KO males and females although males displayed higher tumor weight and grades [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. To ascertain that lack of miR-122 indeed accounts for HCC development, the authors elegantly demonstrated through in vivo rescue experiments that tumor development may be prevented by restoration of hepatic miR-122 expression [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. These experiments provide definite conclusions regarding the tumor suppressor function of miR-122 in the liver.Noteworthy, HCC-resembling tumors were associated with hepatic infiltration of inflammatory cells and increased levels of pro-inflammatory/pro-tumorigenic cytokines such as IL-6 and TNFα [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. In order to assess whether the anti-inflammatory function of miR-122 may be dissociated from its tumor suppressor function, Hsu et al. used a previously described non-inflammatory HCC mouse model (tet-o-MYC; LAP-tTA mice) [[8]Beer S. Zetterberg A. Ihrie R.A. McTaggart R.A. Yang Q. Bradon N. et al.Developmental context determines latency of MYC-induced tumorigenesis.PLoS Biol. 2004; 2: e332Crossref PubMed Scopus (118) Google Scholar]. Interestingly, miR-122 levels were markedly reduced in tumors from these mice and adenoviral-mediated delivery of miR-122 strongly reduced tumor burden in treated mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Thus, these experiments clearly demonstrate that miR-122 possesses an anti-tumorigenic function that is independent of its role in preventing liver disease and inflammation.In conclusion, these new mouse models provide an important step forward in the understanding of HCC development. These data clearly confirm an important role of miR-122 in hepatocarcinogenesis. Further studies are needed to explore the tumor suppressor function of miR-122. These results have also profound implications regarding the therapeutic potential of miR-122. While they indicate that complementing the low expression of miR-122 could be a valid strategy against HCC, it is important to note that miR-122 is also an essential host factor for hepatitis C virus (HCV) replication [[9]Jopling C.L. Yi M. Lancaster A.M. Lemon S.M. Sarnow P. Modulation of hepatitis C virus RNA abundance by a liver-specific MicroRNA.Science. 2005; 309: 1577-1581Crossref PubMed Scopus (2099) Google Scholar]. In contrast to the physiological role of miR-122 in negatively regulating expression of proteins involved in lipid metabolism, miR-122 positively regulates HCV translation and replication. Antiviral strategies targeting miR-122 are therefore in clinical development [[10]Janssen H.L. Reesink H.W. Zeuzem S. Lawitz E. Rodriguez-Torres M. Chen A. et al.A randomized, double-blind, placebo (plb) controlled safety and anti-viral proof-of-concept study of miravirsen (mir), an oligonucleotide targeting miR122, in treatment naive patients with genotype 1 (gt1) chronic HCV infection.Hepatology. 2011; 54 ([abstract LB106]): 101ACrossref PubMed Scopus (35) Google Scholar]. Given that chronic HCV infection is a major cause of HCC, the fact that miR-122 acts as a tumor suppressor in hepatocytes will have to be taken into account for the treatment of chronic HCV infection.Conflict of interestThe authors declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. COMMENTARY ON: Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver. Hsu SH, Wang B, Kota J, Yu J, Costinean S, Kutay H, Yu L, Bai S, La Perle K, Chivukula RR, Mao H, Wei M, Clark KR, Mendell JR, Caligiuri MA, Jacob ST,Mendell JT, Ghoshal K. J Clin Invest. 2012 Aug 1;122(8):2871–83. Copyright (2012). Reprinted with permission of American Society for Clinical Investigation. http://www.ncbi.nlm.nih.gov/pubmed/22820288 Abstract: miR-122, an abundant liver-specific microRNA (miRNA), regulates cholesterol metabolism and promotes hepatitis C virus (HCV) replication. Reduced miR-122 expression in hepatocellular carcinoma (HCC) correlates with metastasis and poor prognosis. Nevertheless, the consequences of sustained loss of function of miR-122 in vivo have not been determined. Here, we demonstrate that deletion of mouse Mir122 resulted in hepatosteatosis, hepatitis, and the development of tumors resembling HCC. These pathologic manifestations were associated with hyperactivity of oncogenic pathways and hepatic infiltration of inflammatory cells that produce pro-tumorigenic cytokines, including IL-6 and TNF. Moreover, delivery of miR-122 to a MYC-driven mouse model of HCC strongly inhibited tumorigenesis, further supporting the tumor suppressor activity of this miRNA. These findings reveal critical functions for miR-122 in the maintenance of liver homeostasis and have important therapeutic implications, including the potential utility of miR-122 delivery for selected patients with HCC and the need for careful monitoring of patients receiving miR-122 inhibition therapy for HCV. AND MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis. Tsai WC, Hsu SD, Hsu CS, Lai TC, Chen SJ, Shen R, Huang Y, Chen HC, Lee CH, Tsai TF, Hsu MT, Wu JC, Huang HD, Shiao MS, Hsiao M, Tsou AP. J Clin Invest. 2012 Aug 1;122(8):2884–97. Copyright 2012. Reprinted with permission of AMERICAN SOCIETY FOR CLINICAL INVESTIGATION. http://www.ncbi.nlm.nih.gov/pubmed/22820290 Abstract: MicroRNA-122 (miR-122), which accounts for 70% of the liver’s total miRNAs, plays a pivotal role in the liver. However, its intrinsic physiological roles remain largely undetermined. We demonstrated that mice lacking the gene encoding miR-122a (Mir122a) are viable but develop temporally controlled steatohepatitis, fibrosis, and hepatocellular carcinoma (HCC). These mice exhibited a striking disparity in HCC incidence based on sex, with a male-to-female ratio of 3.9:1, which recapitulates the disease incidence in humans. Impaired expression of microsomal triglyceride transfer protein (MTTP) contributed to steatosis, which was reversed by in vivo restoration of Mttp expression. We found that hepatic fibrosis onset can be partially attributed to the action of a miR-122a target, the Klf6 transcript. In addition, Mir122a−/− livers exhibited disruptions in a range of pathways, many of which closely resemble the disruptions found in human HCC. Importantly, the reexpression of miR-122a reduced disease manifestation and tumor incidence in Mir122a−/− mice. This study demonstrates that mice with a targeted deletion of the Mir122a gene possess several key phenotypes of human liver diseases, which provides a rationale for the development of a unique therapy for the treatment of chronic liver disease and HCC. Hepatocellular carcinoma (HCC) is the fifth most prevalent and the third leading cause of cancer death worldwide. Although early-stage tumors can be curated using surgical approaches, treatment options for advanced HCC are limited. Virus- and alcohol-induced cirrhosis represent major causes of HCC, but the molecular mechanisms underlying HCC are still only incompletely understood [[1]El-Serag H.B. Epidemiology of viral hepatitis and hepatocellular carcinoma.Gastroenterology. 2012; 142 (e1261): 1264-1273Abstract Full Text Full Text PDF PubMed Scopus (2341) Google Scholar]. Several microRNAs (miRNAs) have been implicated in hepatocarcinogenesis and may represent promising new anti-cancer targets (reviewed in [[2]Wang X.W. Heegaard N.H. Orum H. MicroRNAs in liver disease.Gastroenterology. 2012; 142: 1431-1443Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar]). miRNAs are small non-coding regulatory RNAs resulting from Dicer-mediated processing of miRNA precursors. They act by modulating target messenger RNA translation and/or stabilization and are involved in a wide variety of biological processes. miR-122, a miRNA predominantly expressed in hepatocytes, accounts for approximately 70% of liver-expressed miRNAs. Evidence from sequestrating miR-122 in vivo suggests that this miRNA is involved in cholesterol biosynthesis and liver function [[3]Esau C. Davis S. Murray S.F. Yu X.X. Pandey S.K. Pear M. et al.MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.Cell Metab. 2006; 3: 87-98Abstract Full Text Full Text PDF PubMed Scopus (1737) Google Scholar]. It may also play a role in hepatocarcinogenesis as its expression is downregulated in HCC [[4]Bai S. Nasser M.W. Wang B. Hsu S.H. Datta J. Kutay H. et al.MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.J Biol Chem. 2009; 284: 32015-32027Crossref PubMed Scopus (427) Google Scholar] and hepatocyte-specific Dicer1 knockout (KO) mice spontaneously develop HCC [[5]Sekine S. Ogawa R. Ito R. Hiraoka N. McManus M.T. Kanai Y. et al.Disruption of Dicer1 induces dysregulated fetal gene expression and promotes hepatocarcinogenesis.Gastroenterology. 2009; 136 (e2301–2304): 2304-2315Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar]. However, its exact role in hepatocarcinogenesis and the underlying molecular mechanisms had not been investigated yet. Two independent laboratories now report arresting conclusions regarding the metabolic, anti-inflammatory, and anti-tumorigenic roles of miR-122, by generating germline KO, called either Mir122 KO [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar] or Mir122a−/− [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar], and liver-specific KO (Mir122 LKO) mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. These mice developed normally and were characterized by reduced serum cholesterol levels in line with previous studies reporting antisense miR-122 sequestration in vivo [[3]Esau C. Davis S. Murray S.F. Yu X.X. Pandey S.K. Pear M. et al.MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.Cell Metab. 2006; 3: 87-98Abstract Full Text Full Text PDF PubMed Scopus (1737) Google Scholar]. In Mir122 KO and Mir122 LKO mice, total serum cholesterol was reduced by 30% with concomitant decrease in LDL and HDL while TG levels remained unchanged [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Reduced serum cholesterol (total cholesterol, VLDL and HDL) was also observed in Mir122a−/− along with reduced TG levels [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. In contrast to antisense miR-122-treated mice, these Mir122 KO mice developed steatosis and hepatitis that progressed to fibrosis and HCC and thereby displayed a phenotype resembling liver-specific Dicer1 KO mice [[5]Sekine S. Ogawa R. Ito R. Hiraoka N. McManus M.T. Kanai Y. et al.Disruption of Dicer1 induces dysregulated fetal gene expression and promotes hepatocarcinogenesis.Gastroenterology. 2009; 136 (e2301–2304): 2304-2315Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar] (Table 1). Hepatic gene expression analysis indicated that numerous genes involved in lipid metabolism were differentially regulated in LKO mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Mir122a−/− mice also demonstrated impaired expression of several proteins involved in lipid metabolism, among which microsomal triglyceride transfer protein (MTTP). Although miR-122 does not directly target MTTP messenger RNA, prolonged injection of MTTP- or miR-122-expression constructs in Mir122a−/− mice allowed restoration of MTTP levels, increase of serum lipids and reduction of hepatic steatosis, inflammation and fibrosis [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. These functional experiments indicate that lack of MTTP and/or miR-122 expression leads to liver disease. In addition, both groups validated several novel direct targets of miR-122, among which Klf6 or Agpat1, that could also account for the observed phenotype [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. Moreover, in both models, prolonged lack of miR-122 in mouse hepatocytes associates with HCC [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. While spontaneous tumors were 3.9-fold higher in male Mir122a−/− mice than in females [[7]Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar], in line with observed HCC incidence in humans, Hsu et al. observed equal penetration in Mir122 KO males and females although males displayed higher tumor weight and grades [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. To ascertain that lack of miR-122 indeed accounts for HCC development, the authors elegantly demonstrated through in vivo rescue experiments that tumor development may be prevented by restoration of hepatic miR-122 expression [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. These experiments provide definite conclusions regarding the tumor suppressor function of miR-122 in the liver. Noteworthy, HCC-resembling tumors were associated with hepatic infiltration of inflammatory cells and increased levels of pro-inflammatory/pro-tumorigenic cytokines such as IL-6 and TNFα [6Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar, 7Tsai W.C. Hsu S.D. Hsu C.S. Lai T.C. Chen S.J. Shen R. et al.MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.J Clin Invest. 2012; 122: 2884-2897Crossref PubMed Scopus (628) Google Scholar]. In order to assess whether the anti-inflammatory function of miR-122 may be dissociated from its tumor suppressor function, Hsu et al. used a previously described non-inflammatory HCC mouse model (tet-o-MYC; LAP-tTA mice) [[8]Beer S. Zetterberg A. Ihrie R.A. McTaggart R.A. Yang Q. Bradon N. et al.Developmental context determines latency of MYC-induced tumorigenesis.PLoS Biol. 2004; 2: e332Crossref PubMed Scopus (118) Google Scholar]. Interestingly, miR-122 levels were markedly reduced in tumors from these mice and adenoviral-mediated delivery of miR-122 strongly reduced tumor burden in treated mice [[6]Hsu S.H. Wang B. Kota J. Yu J. Costinean S. Kutay H. et al.Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.J Clin Invest. 2012; 122: 2871-2883Crossref PubMed Scopus (590) Google Scholar]. Thus, these experiments clearly demonstrate that miR-122 possesses an anti-tumorigenic function that is independent of its role in preventing liver disease and inflammation. In conclusion, these new mouse models provide an important step forward in the understanding of HCC development. These data clearly confirm an important role of miR-122 in hepatocarcinogenesis. Further studies are needed to explore the tumor suppressor function of miR-122. These results have also profound implications regarding the therapeutic potential of miR-122. While they indicate that complementing the low expression of miR-122 could be a valid strategy against HCC, it is important to note that miR-122 is also an essential host factor for hepatitis C virus (HCV) replication [[9]Jopling C.L. Yi M. Lancaster A.M. Lemon S.M. 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Given that chronic HCV infection is a major cause of HCC, the fact that miR-122 acts as a tumor suppressor in hepatocytes will have to be taken into account for the treatment of chronic HCV infection. Conflict of interestThe authors declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. The authors declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. Work in T.F.B.’s laboratory is supported by the European Union ( ERC-2008-AdG-233130-HEPCENT , INTERREG-IV-Rhin Supérieur-FEDER-Hepato-Regio-Net 2009 ), Laboratoire d’Excellence HEPSYS (Investissement d’Avenir; ANR-10-LAB-28 ), ANR-05-CEXC-008 , ANRS ( 2008/354, 2009/183, 2011/132, 2012/239 ), Région Alsace, Inserm, and University of Strasbourg . Work in S.P.’s laboratory is supported by the European Research Council ( ERC-2010-StG-260767-ncRNAVIR) , Agence Nationale de la Recherche ( ANR-08-MIEN-005 and ANR-10-LABX-36-netRNA ), the Institut National du Cancer ( INCa-PLBIO-2009-173 ) and CNRS .