Dietary Methionine Restriction Regulates Liver Protein Synthesis and Gene Expression Independently of Eukaryotic Initiation Factor 2 Phosphorylation in Mice

Background: The phosphorylation of eukaryotic initiation factor 2 (p-eIF2) during dietary amino acid insufficiency reduces protein synthesis and alters gene expression via the integrated stress response (ISR).Objective: We explored whether a Met-restricted (MR) diet activates the ISR to reduce body fat and regulate protein balance.Methods: Male and female mice aged 3-6 mo with either whole-body deletion of general control nonderepressible 2 (Gcn2) or liver-specific deletion of protein kinase R-like endoplasmic reticulum kinase (Perk) alongside wild-type or floxed control mice were fed an obesogenic diet sufficient in Met (0.86%) or an MR (0.12% Met) diet for ≤5 wk. Ala enrichment with deuterium was measured to calculate protein synthesis rates. The guanine nucleotide exchange factor activity of eIF2B was measured alongside p-eIF2 and hepatic mRNA expression levels at 2 d and 5 wk. Metabolic phenotyping was conducted at 4 wk, and body composition was measured throughout. Results were evaluated with the use of ANOVA (P < 0.05).Results: Feeding an MR diet for 2 d did not increase hepatic p-eIF2 or reduce eIF2B activity in wild-type or Gcn2-/- mice, yet many genes transcriptionally regulated by the ISR were altered in both strains in the same direction and amplitude. Feeding an MR diet for 5 wk increased p-eIF2 and reduced eIF2B activity in wild-type but not Gcn2-/- mice, yet ISR-regulated genes altered in both strains similarly. Furthermore, the MR diet reduced mixed and cytosolic but not mitochondrial protein synthesis in both the liver and skeletal muscle regardless of Gcn2 status. Despite the similarities between strains, the MR diet did not increase energy expenditure or reduce body fat in Gcn2-/- mice. Finally, feeding the MR diet to mice with Perk deleted in the liver increased hepatic p-eIF2 and altered body composition similar to floxed controls.Conclusions: Hepatic activation of the ISR resulting from an MR diet does not require p-eIF2. Gcn2 status influences body fat loss but not protein balance when Met is restricted

Physiologic Responses to Dietary Sulfur Amino Acid Restriction in Mice Are Influenced by Atf4 Status and Biological Sex

Background: Dietary sulfur amino acid restriction (SAAR) improves body composition and metabolic health across several model organisms in part through induction of the integrated stress response (ISR). Objective: We investigate the hypothesis that activating transcription factor 4 (ATF4) acts as a converging point in the ISR during SAAR. Methods: Using liver-specific or global gene ablation strategies, in both female and male mice, we address the role of ATF4 during dietary SAAR. Results: We show that ATF4 is dispensable in the chronic induction of the hepatokine fibroblast growth factor 21 while being essential for the sustained production of endogenous hydrogen sulfide. We also affirm that biological sex, independent of ATF4 status, is a determinant of the response to dietary SAAR. Conclusions: Our results suggest that auxiliary components of the ISR, which are independent of ATF4, are critical for SAAR-mediated improvements in metabolic health in mice

Disrupted Nitric Oxide Metabolism from Type II Diabetes and Acute Exposure to Particulate Air Pollution.

Type II diabetes is an established cause of vascular impairment. Particulate air pollution is known to exacerbate cardiovascular and respiratory conditions, particularly in susceptible populations. This study set out to determine the impact of exposure to traffic pollution, with and without particle filtration, on vascular endothelial function in Type II diabetes. Endothelial production of nitric oxide (NO) has previously been linked to vascular health. Reactive hyperemia induces a significant increase in plasma nitrite, the proximal metabolite of NO, in healthy subjects, while diabetics have a lower and more variable level of response. Twenty type II diabetics and 20 controls (ages 46-70 years) were taken on a 1.5 hr roadway traffic air pollution exposure as passengers. We analyzed plasma nitrite, as a measure of vascular function, using forearm ischemia to elicit a reactive hyperemic response before and after exposure to one ride with and one without filtration of the particle components of pollution. Control subjects displayed a significant increase in plasma nitrite levels during reactive hyperemia. This response was no longer present following exposure to traffic air pollution, but did not vary with whether or not the particle phase was filtered out. Diabetics did not display an increase in nitrite levels following reactive hyperemia. This response was not altered following pollution exposure. These data suggest that components of acute traffic pollution exposure diminish vascular reactivity in non-diabetic individuals. It also confirms that type II diabetics have a preexisting diminished ability to appropriately respond to a vascular challenge, and that traffic pollution exposure does not cause a further measureable acute change in plasma nitrite levels in Type II diabetics

Acute ride (air pollution) effects on resting vascular nitrite levels (pre-ischemia).

<p>(A) Controls resting nitrite levels pre and post ride. (B) Diabetic resting nitrite levels pre and post ride. Dash lines indicate decreasing nitrite values, while solid lines indicate increasing values from pre to post ride. Unfiltered and filtered rides are combined. ^adenotes mean values (thick black line).</p

Air pollution exposure effects on vascular nitrite response to reactive hyperemia in controls and diabetics.

<p>A) Control subjects’ change in plasma nitrite response to ischemia (resting nitrites subtracted from reactive hyperemia nitrites)before (pre ride) and following (post ride) exposure (n = 34). Mean changes in nitrite values (pre ride: 26.1nM; post ride:-4.1nM). B) Diabetic subjects’ change in plasma nitrite response to ischemia before and following exposure (n = 25). Mean change in nitrite values (pre ride: 8.7nM; post ride: 20.6nM) Dash lines indicate decreasing nitrite values, while solid lines indicate increasing values. Unfiltered and filtered rides are combined. ^<b>a</b>denotes (pre ischemia (resting) nitrite level) subtracted from (post ischemia (reactive hyperemia)nitrite level). ^<b>b</b>denotes mean values (thick black line).</p

Vascular nitrite concentrations at rest and following ischemia in control and diabetic subjects.

<p>Blood draws were taken at rest and 60sec following release (hyperemia) of the five minute upper arm occlusion (50mmHg above SBP) before and following the ride. Dash lines indicate decreasing nitrite values, while solid lines indicate increasing values. Unfiltered and filtered rides are combined (A)Nitrite concentration (nM) changes in control subjects before the ride(n = 40); (B) Nitrite concentration (nM) changes in diabetic subjects before the ride(n = 30); (C) Nitrite concentration (nM) changes in control subjects following the ride(n = 30); (D) Nitrite concentration (nM) changes in diabetic subjects following the ride(n = 25). ^adenotes mean values (thick black line).</p