Hypoxia-inducible factor–1α–dependent induction of miR122 enhances hepatic ischemia tolerance
Cynthia Ju, … , Kalpana Ghoshal, Holger K. Eltzschig
Cynthia Ju, … , Kalpana Ghoshal, Holger K. Eltzschig
Published April 1, 2021
Citation Information: J Clin Invest. 2021;131(7):e140300. https://doi.org/10.1172/JCI140300.
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Hypoxia-inducible factor–1α–dependent induction of miR122 enhances hepatic ischemia tolerance

Abstract

Hepatic ischemia and reperfusion (IR) injury contributes to the morbidity and mortality associated with liver transplantation. microRNAs (miRNAs) constitute a family of noncoding RNAs that regulate gene expression at the posttranslational level through the repression of specific target genes. Here, we hypothesized that miRNAs could be targeted to enhance hepatic ischemia tolerance. A miRNA screen in a murine model of hepatic IR injury pointed us toward the liver-specific miRNA miR122. Subsequent studies in mice with hepatocyte-specific deletion of miR122 (miR122loxP/loxP Alb-Cre+ mice) during hepatic ischemia and reperfusion revealed exacerbated liver injury. Transcriptional studies implicated hypoxia-inducible factor–1α (HIF1α) in the induction of miR122 and identified the oxygen-sensing prolyl hydroxylase domain 1 (PHD1) as a miR122 target. Further studies indicated that HIF1α-dependent induction of miR122 participated in a feed-forward pathway for liver protection via the enhancement of hepatic HIF responses through PHD1 repression. Moreover, pharmacologic studies utilizing nanoparticle-mediated miR122 overexpression demonstrated attenuated liver injury. Finally, proof-of-principle studies in patients undergoing orthotopic liver transplantation showed elevated miR122 levels in conjunction with the repression of PHD1 in post-ischemic liver biopsies. Taken together, the present findings provide molecular insight into the functional role of miR122 in enhancing hepatic ischemia tolerance and suggest the potential utility of pharmacologic interventions targeting miR122 to dampen hepatic injury during liver transplantation.

Authors

Cynthia Ju, Meng Wang, Eunyoung Tak, Boyun Kim, Christoph Emontzpohl, Yang Yang, Xiaoyi Yuan, Huban Kutay, Yafen Liang, David R. Hall, Wasim A. Dar, J. Steve Bynon, Peter Carmeliet, Kalpana Ghoshal, Holger K. Eltzschig

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Figure 2

Role of HIF in miR122 induction.

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Role of HIF in miR122 induction. (A) The promoter for miR122 includes 2 ...
(A) The promoter for miR122 includes 2 putative HIF-binding sites — hypoxia response elements (HREs). Maps indicate the promoter constructs generated for analysis of promoter activity. (B) ChIP analysis demonstrated binding of HIF1α to the putative HRE within the miR122 promoter (2-way ANOVA. n = 4/group from 4independent experiments). (C) Effective knockdown of HIF1A mRNA following lentivirus-mediated HIF1α shRNA delivery into HepG2 cells (2-tailed, unpaired Student’s t test. n = 8 and n = 7 for the control [Ctrl] and shRNA groups, respectively). (D and E) HIF1α shRNA abolished HIF1α induction in HepG2 cell nuclei following 6-hour hypoxia culturing in 1% oxygen (results are representative of 3 independent experiments). (G–I) Experiments analogous to those depicted in D and E demonstrate preserved induction of miR122 following shRNA-mediated repression of HIF2α. (G) Two-tailed, unpaired Student’s t test; n = 8 and 11 for control and shRNA groups, respectively. (H and I) Results are representative of 4 independent experiments. (F and J) Time-dependent induction of miR122 in HepG2 cells exposed to hypoxia was abolished following HIF1α, but not HIF2α, knockdown compared with the same control shRNA group (2-way ANOVA, n = 4, and 6 at 0, 2, 4, 8 hours for the control group shared by F and J, and n = 6 for all shRNA groups, respectively, at 0, 2, 4, and 8 hours). (K–N) In mice with hepatocyte-specific deletion of HIF1α (HIF1Afl/fl Alb-Cre+), HIF1A mRNA and protein were abolished, and induction of miR122 was abolished following ischemia and reperfusion (2-tailed, unpaired Student’s t test). (K) n = 5 and 8 in WT and KO groups, respectively. (M) n = 3/group. (N) n = 8 and 6 in WT and KO groups, respectively. All data are shown as the mean ± SEM.