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Molecular regulation of urea cycle function by the liver glucocorticoid receptor.

Okun JG, Conway S, Schmidt KV, Schumacher J, Wang X, de Guia R, Zota A, Klement J, Seibert O, Peters A, Maida A, Herzig S, Rose AJ - Mol Metab (2015)

Bottom Line: Serum urea levels were consistently affected by GC/GR gain- (∼+32%) and loss (∼-30%) -of-function.Combined liver-specific GR loss-of-function with DEX treatment revealed a tissue-autonomous role for the GR to coordinate an upregulation of liver urea production rate in vivo and ex vivo, and prevent hyperammonaemia and associated neuromuscular dysfunction in vivo.Liver mRNA expression profiling and GR-cistrome mining identified Arginase I (ARG1) a urea cycle gene targeted by the liver GR.

View Article: PubMed Central - PubMed

Affiliation: Division of Neuropediatrics and Metabolic Medicine, University Children's Hospital, Heidelberg, Germany.

ABSTRACT

Objective: One of the major side effects of glucocorticoid (GC) treatment is lean tissue wasting, indicating a prominent role in systemic amino acid metabolism. In order to uncover a novel aspect of GCs and their intracellular-receptor, the glucocorticoid receptor (GR), on metabolic control, we conducted amino acid and acylcarnitine profiling in human and mouse models of GC/GR gain- and loss-of-function.

Methods: Blood serum and tissue metabolite levels were determined in Human Addison's disease (AD) patients as well as in mouse models of systemic and liver-specific GR loss-of-function (AAV-miR-GR) with or without dexamethasone (DEX) treatments. Body composition and neuromuscular and metabolic function tests were conducted in vivo and ex vivo, the latter using precision cut liver slices.

Results: A serum metabolite signature of impaired urea cycle function (i.e. higher [ARG]:[ORN + CIT]) was observed in human (CTRL: 0.45 ± 0.03, AD: 1.29 ± 0.04; p < 0.001) and mouse (AAV-miR-NC: 0.97 ± 0.13, AAV-miR-GR: 2.20 ± 0.19; p < 0.001) GC/GR loss-of-function, with similar patterns also observed in liver. Serum urea levels were consistently affected by GC/GR gain- (∼+32%) and loss (∼-30%) -of-function. Combined liver-specific GR loss-of-function with DEX treatment revealed a tissue-autonomous role for the GR to coordinate an upregulation of liver urea production rate in vivo and ex vivo, and prevent hyperammonaemia and associated neuromuscular dysfunction in vivo. Liver mRNA expression profiling and GR-cistrome mining identified Arginase I (ARG1) a urea cycle gene targeted by the liver GR.

Conclusions: The liver GR controls systemic and liver urea cycle function by transcriptional regulation of ARG1 expression.

No MeSH data available.


Related in: MedlinePlus

The liver glucocorticoid receptor coordinates the enhanced urea cycle function upon dexamethasone treatment. Urea production rate (A) as well as ornithine (B) and arginine (C) balance in mouse liver slices treated with dexamethasone, RU486 and corresponding vehicle treatments ex vivo (N = 4/group). Serum urea levels (A) and urea area under the curve (AUC; B) in response to an intraperitoneal alanine tolerance test (ipATT) in GR-floxed mice pretreated with adenoviral constructs expressing (Ad-CRE) or not (Ad-NC) Cre-recombinase with (Dex) or without (Veh) chronic dexamethasone treatment (N = 6–7/group). Serum (F) and liver (G) amino acid levels in the same mice as in D. ARG: arginine, ORN: ornithine, CIT: citrulline. Data are mean ± SEM. Effect of Dex: *p < 0.05, **p < 0.01, ***p < 0.001. Effect of Ad-CRE: #p < 0.05, ##p < 0.01, ###p < 0.001.
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fig3: The liver glucocorticoid receptor coordinates the enhanced urea cycle function upon dexamethasone treatment. Urea production rate (A) as well as ornithine (B) and arginine (C) balance in mouse liver slices treated with dexamethasone, RU486 and corresponding vehicle treatments ex vivo (N = 4/group). Serum urea levels (A) and urea area under the curve (AUC; B) in response to an intraperitoneal alanine tolerance test (ipATT) in GR-floxed mice pretreated with adenoviral constructs expressing (Ad-CRE) or not (Ad-NC) Cre-recombinase with (Dex) or without (Veh) chronic dexamethasone treatment (N = 6–7/group). Serum (F) and liver (G) amino acid levels in the same mice as in D. ARG: arginine, ORN: ornithine, CIT: citrulline. Data are mean ± SEM. Effect of Dex: *p < 0.05, **p < 0.01, ***p < 0.001. Effect of Ad-CRE: #p < 0.05, ##p < 0.01, ###p < 0.001.

Mentions: Hepatic glucocorticoid resistance impairs the upregulation of urea cycle resulting in hyperammonaemia and neuromuscular dysfunction upon dexamethasone treatment. Conceivably, changes in urea cycle function in vivo upon GR gain- and loss-of-function could result from tissue autonomous or tissue non-autonomous mechanisms. To investigate this further, we performed studies using mouse liver slices ex vivo, which, unlike primary hepatocytes, maintain glucocorticoid sensitivity and more closely resemble fully differentiated liver in vivo[23]. We treated mouse liver slices with DEX and the GR-antagonist RU486, and combinations thereof, and could demonstrate that while DEX promoted liver urea production rate, as well as net ORN output and net ARG and TYR uptake, RU486 completely inhibited this effect (Figure 3A–C; Figure S3A), demonstrating that the liver GR action per se is sufficient to confer effects of DEX on urea cycle function. Given this result, we wanted to test whether the liver GR could confer systemic effects of DEX treatment in vivo. Indeed, liver GR loss-of-function (Figure S3B) had expected effects on blood glucose (Figure S3C), but also dramatically reduced DEX-induced in vivo urea production rate, as judged by an alanine tolerance test (Figure 3D–E). Importantly, serum (Figure S3F) and liver (Figure S3G) amino acid profiling revealed that while DEX promoted lower ARG and higher ORN and CIT levels, these effects were blunted by liver-specific GR knockdown, highlighting a key role for the liver in mediating these changes. We then examined the potential functional consequences of GR loss-of-function in this study, and could demonstrate that the liver GR does not affect the reduction of body mass that typically occurs with DEX treatment in vivo (Figure 4A–B). In particular, ECHO-MRI revealed that DEX reduced lean mass (Figure 4C) but not fat mass (Figure S4A), with lower skeletal muscle (Figure S4B) and liver (Figure S4C) mass upon DEX treatment. We hypothesised that if the urea cycle regulation were dysfunctional, this would lead to hyperammonaemia upon DEX treatment, and, indeed, this was the case (Figure 4D). Given that hyperammonaemia typically results in a manifestation of neuromuscular dysfunction [34], we tested this possibility and could demonstrate that both forelimb grip strength (Figure 4E) as well as balance control (Figure 4F) were negatively affected by DEX treatment in liver GR knockout mice. Taken together, these data demonstrate that the liver GR coordinated effective amino acid disposal via tissue-autonomous upregulation of urea cycle function upon DEX-induced lean tissue wasting.


Molecular regulation of urea cycle function by the liver glucocorticoid receptor.

Okun JG, Conway S, Schmidt KV, Schumacher J, Wang X, de Guia R, Zota A, Klement J, Seibert O, Peters A, Maida A, Herzig S, Rose AJ - Mol Metab (2015)

The liver glucocorticoid receptor coordinates the enhanced urea cycle function upon dexamethasone treatment. Urea production rate (A) as well as ornithine (B) and arginine (C) balance in mouse liver slices treated with dexamethasone, RU486 and corresponding vehicle treatments ex vivo (N = 4/group). Serum urea levels (A) and urea area under the curve (AUC; B) in response to an intraperitoneal alanine tolerance test (ipATT) in GR-floxed mice pretreated with adenoviral constructs expressing (Ad-CRE) or not (Ad-NC) Cre-recombinase with (Dex) or without (Veh) chronic dexamethasone treatment (N = 6–7/group). Serum (F) and liver (G) amino acid levels in the same mice as in D. ARG: arginine, ORN: ornithine, CIT: citrulline. Data are mean ± SEM. Effect of Dex: *p < 0.05, **p < 0.01, ***p < 0.001. Effect of Ad-CRE: #p < 0.05, ##p < 0.01, ###p < 0.001.
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Related In: Results  -  Collection

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fig3: The liver glucocorticoid receptor coordinates the enhanced urea cycle function upon dexamethasone treatment. Urea production rate (A) as well as ornithine (B) and arginine (C) balance in mouse liver slices treated with dexamethasone, RU486 and corresponding vehicle treatments ex vivo (N = 4/group). Serum urea levels (A) and urea area under the curve (AUC; B) in response to an intraperitoneal alanine tolerance test (ipATT) in GR-floxed mice pretreated with adenoviral constructs expressing (Ad-CRE) or not (Ad-NC) Cre-recombinase with (Dex) or without (Veh) chronic dexamethasone treatment (N = 6–7/group). Serum (F) and liver (G) amino acid levels in the same mice as in D. ARG: arginine, ORN: ornithine, CIT: citrulline. Data are mean ± SEM. Effect of Dex: *p < 0.05, **p < 0.01, ***p < 0.001. Effect of Ad-CRE: #p < 0.05, ##p < 0.01, ###p < 0.001.
Mentions: Hepatic glucocorticoid resistance impairs the upregulation of urea cycle resulting in hyperammonaemia and neuromuscular dysfunction upon dexamethasone treatment. Conceivably, changes in urea cycle function in vivo upon GR gain- and loss-of-function could result from tissue autonomous or tissue non-autonomous mechanisms. To investigate this further, we performed studies using mouse liver slices ex vivo, which, unlike primary hepatocytes, maintain glucocorticoid sensitivity and more closely resemble fully differentiated liver in vivo[23]. We treated mouse liver slices with DEX and the GR-antagonist RU486, and combinations thereof, and could demonstrate that while DEX promoted liver urea production rate, as well as net ORN output and net ARG and TYR uptake, RU486 completely inhibited this effect (Figure 3A–C; Figure S3A), demonstrating that the liver GR action per se is sufficient to confer effects of DEX on urea cycle function. Given this result, we wanted to test whether the liver GR could confer systemic effects of DEX treatment in vivo. Indeed, liver GR loss-of-function (Figure S3B) had expected effects on blood glucose (Figure S3C), but also dramatically reduced DEX-induced in vivo urea production rate, as judged by an alanine tolerance test (Figure 3D–E). Importantly, serum (Figure S3F) and liver (Figure S3G) amino acid profiling revealed that while DEX promoted lower ARG and higher ORN and CIT levels, these effects were blunted by liver-specific GR knockdown, highlighting a key role for the liver in mediating these changes. We then examined the potential functional consequences of GR loss-of-function in this study, and could demonstrate that the liver GR does not affect the reduction of body mass that typically occurs with DEX treatment in vivo (Figure 4A–B). In particular, ECHO-MRI revealed that DEX reduced lean mass (Figure 4C) but not fat mass (Figure S4A), with lower skeletal muscle (Figure S4B) and liver (Figure S4C) mass upon DEX treatment. We hypothesised that if the urea cycle regulation were dysfunctional, this would lead to hyperammonaemia upon DEX treatment, and, indeed, this was the case (Figure 4D). Given that hyperammonaemia typically results in a manifestation of neuromuscular dysfunction [34], we tested this possibility and could demonstrate that both forelimb grip strength (Figure 4E) as well as balance control (Figure 4F) were negatively affected by DEX treatment in liver GR knockout mice. Taken together, these data demonstrate that the liver GR coordinated effective amino acid disposal via tissue-autonomous upregulation of urea cycle function upon DEX-induced lean tissue wasting.

Bottom Line: Serum urea levels were consistently affected by GC/GR gain- (∼+32%) and loss (∼-30%) -of-function.Combined liver-specific GR loss-of-function with DEX treatment revealed a tissue-autonomous role for the GR to coordinate an upregulation of liver urea production rate in vivo and ex vivo, and prevent hyperammonaemia and associated neuromuscular dysfunction in vivo.Liver mRNA expression profiling and GR-cistrome mining identified Arginase I (ARG1) a urea cycle gene targeted by the liver GR.

View Article: PubMed Central - PubMed

Affiliation: Division of Neuropediatrics and Metabolic Medicine, University Children's Hospital, Heidelberg, Germany.

ABSTRACT

Objective: One of the major side effects of glucocorticoid (GC) treatment is lean tissue wasting, indicating a prominent role in systemic amino acid metabolism. In order to uncover a novel aspect of GCs and their intracellular-receptor, the glucocorticoid receptor (GR), on metabolic control, we conducted amino acid and acylcarnitine profiling in human and mouse models of GC/GR gain- and loss-of-function.

Methods: Blood serum and tissue metabolite levels were determined in Human Addison's disease (AD) patients as well as in mouse models of systemic and liver-specific GR loss-of-function (AAV-miR-GR) with or without dexamethasone (DEX) treatments. Body composition and neuromuscular and metabolic function tests were conducted in vivo and ex vivo, the latter using precision cut liver slices.

Results: A serum metabolite signature of impaired urea cycle function (i.e. higher [ARG]:[ORN + CIT]) was observed in human (CTRL: 0.45 ± 0.03, AD: 1.29 ± 0.04; p < 0.001) and mouse (AAV-miR-NC: 0.97 ± 0.13, AAV-miR-GR: 2.20 ± 0.19; p < 0.001) GC/GR loss-of-function, with similar patterns also observed in liver. Serum urea levels were consistently affected by GC/GR gain- (∼+32%) and loss (∼-30%) -of-function. Combined liver-specific GR loss-of-function with DEX treatment revealed a tissue-autonomous role for the GR to coordinate an upregulation of liver urea production rate in vivo and ex vivo, and prevent hyperammonaemia and associated neuromuscular dysfunction in vivo. Liver mRNA expression profiling and GR-cistrome mining identified Arginase I (ARG1) a urea cycle gene targeted by the liver GR.

Conclusions: The liver GR controls systemic and liver urea cycle function by transcriptional regulation of ARG1 expression.

No MeSH data available.


Related in: MedlinePlus