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In vivo regulation of the heme oxygenase-1 gene in humanized transgenic mice.

ABSTRACT: Heme oxygenase-1 (HO-1) catalyzes the rate-limiting step in heme degradation, producing equimolar amounts of carbon monoxide, iron, and biliverdin. Induction of HO-1 is a beneficial response to tissue injury in diverse animal models of diseases including acute kidney injury. In vitro analysis has shown that the human HO-1 gene is transcriptionally regulated by changes in chromatin conformation, but whether such control occurs in vivo is not known. To enable such an analysis, we generated transgenic mice, harboring an 87-kb bacterial artificial chromosome expressing human HO-1 mRNA and protein and bred these mice with HO-1 knockout mice to generate humanized BAC transgenic mice. This successfully rescued the phenotype of the knockout mice including reduced birth rates, tissue iron overload, splenomegaly, anemia, leukocytosis, dendritic cell abnormalities, and survival after acute kidney injury induced by rhabdomyolysis or cisplatin nephrotoxicity. Transcription factors such as USF1/2, JunB, Sp1, and CTCF were found to associate with regulatory regions of the human HO-1 gene in the kidney following rhabdomyolysis. Chromosome conformation capture and ChIP-loop assays confirmed this in the formation of chromatin looping in vivo. Thus, these bacterial artificial chromosome humanized HO-1 mice are a valuable model to study the human HO-1 gene, providing insight to the in vivo architecture of the gene in acute kidney injury and other diseases.

SUBMITTER: Kim J

PROVIDER: S-EPMC3396739 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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In vivo regulation of the heme oxygenase-1 gene in humanized transgenic mice.

Kim Junghyun J Zarjou Abolfazl A Traylor Amie M AM Bolisetty Subhashini S Jaimes Edgar A EA Hull Travis D TD George James F JF Mikhail Fady M FM Agarwal Anupam A

Kidney international 20120411 3

Heme oxygenase-1 (HO-1) catalyzes the rate-limiting step in heme degradation, producing equimolar amounts of carbon monoxide, iron, and biliverdin. Induction of HO-1 is a beneficial response to tissue injury in diverse animal models of diseases including acute kidney injury. In vitro analysis has shown that the human HO-1 gene is transcriptionally regulated by changes in chromatin conformation, but whether such control occurs in vivo is not known. To enable such an analysis, we generated transge ...[more]

PMID: 22495295

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Project description:BackgroundHyperhomocysteinemia (HHcy) is an independent risk factor for liver diseases, such as fatty liver and hepatic fibrosis. However, the mechanisms underlying this pro-oxidative effect of homocysteine (Hcy) in hepatocytes remain largely unknown. Thus, we investigated the effect of Hcy on the gene expression of heme oxygenase-1 (HO-1), the primary rate-limiting enzyme in heme catabolism and a key anti-oxidant detoxification enzyme in maintaining cellular redox homeostasis.MethodsIn vivo, twenty male C57BL/6 mice at 8 weeks of age were randomly divided into two groups. One group was fed a chow diet (chow group; n = 10), the other group of mice was fed a methionine-supplemented diet (Met group, 1 mg kg(-1) day(-1) L-methionine in drinking water; n = 10) for 4 weeks. In vitro, HepG2 cells were stimulated with different doses of homocysteine (Hcy).ResultsFour weeks' methionine supplementation caused a significant increase of plasma Hcy concentration and a decrease of HO-1 expression in the liver of C57BL/6 mice than mice received chow diet. Besides, SOD enzyme activities were impaired and the level of oxidative stress markers, such as malondialdehyde (MDA) were elevated in the liver from mice supplemented with methionine compared with control mice. In cultured hepatocytes, Hcy treatment reduced both the mRNA and protein levels of HO-1 dose-dependently. However, Hcy had no effect on the gene expression of Nrf2, the major transcriptional regulator of HO-1. Instead, Hcy induced the expression of Bach1, a transcriptional repressor of HO-1. In addition, Hcy stimulated the nuclear localization of Bach1 but prevented that of Nrf2. Furthermore, we found that knockdown of Bach1 attenuated the suppression of the HO-1 expression by Hcy.ConclusionsCollectively, our results demonstrated that Bach1 plays an important role in Hcy-triggered ROS generations through inhibiting HO-1 expression, likely, resulting from the disturbed interplay between Bach1 and Nrf2.

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In vivo regulation of the heme oxygenase-1 gene in humanized transgenic mice.

Publications

In vivo regulation of the heme oxygenase-1 gene in humanized transgenic mice.

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