Inhibition of DHCR24 activates LXRα to ameliorate hepatic steatosis and inflammation

Liver X receptor (LXR) agonism has theoretical potential for treating NAFLD/NASH, but synthetic agonists induce hyperlipidemia in preclinical models. Desmosterol, which is converted by Δ24-dehydrocholesterol reductase (DHCR24) into cholesterol, is a potent endogenous LXR agonist with anti-inflammatory properties. We aimed to investigate the effects of DHCR24 inhibition on NAFLD/NASH development. Here, by using APOE*3-Leiden. CETP mice, a well-established translational model that develops diet-induced human-like NAFLD/NASH characteristics, we report that SH42, a published DHCR24 inhibitor, markedly increases desmosterol levels in liver and plasma, reduces hepatic lipid content and the steatosis score, and decreases plasma fatty acid and cholesteryl ester concentrations. Flow cytometry showed that SH42 decreases liver inflammation by preventing Kupffer cell activation and monocyte infiltration. LXRα deficiency completely abolishes these beneficial effects of SH42. Together, the inhibition of DHCR24 by SH42 prevents diet-induced hepatic steatosis and inflammation in a strictly LXRα-dependent manner without causing hyperlipidemia. Finally, we also showed that SH42 treatment decreased liver collagen content and plasma alanine transaminase levels in an established NAFLD model. In conclusion, we anticipate that pharmacological DHCR24 inhibition may represent a novel therapeutic strategy for treatment of NAFLD/NASH.</p

Body weight, organ weight, and food intake characteristics.

<p>Body weight, organ weight, and food intake characteristics.</p

Innate immune cells, lymphocytes, and relative gene expression characteristics in liver.

<p>Innate immune cells, lymphocytes, and relative gene expression characteristics in liver.</p

MOS supplementation slightly affected hepatic monocytes and macrophage subsets.

<p>Hepatic extra-intestinal immune modulatory properties of MOS were assessed in mice fed a LFD or HFD with or without MOS for 17 weeks. Percentages of Ly6C^hi monocytes [A], macrophages [B], macrophage M1-like and M2-like subsets [C], eosinophils [D] and CD25+ CD8+ expressing T cells [E] within CD45+ cells in the liver. mRNA expression of the inflammatory markers <i>F4/80</i>, <i>CD11c</i>, <i>Ym1</i>, <i>Mcp1</i>, <i>Tnf-a</i>, <i>Il-6</i>, and <i>Il-10</i> was determined [F]. Values are presented as means ± SEM (n = 4–10 mice/group). Differences were evaluated for statistical significance by by two-way ANOVA, followed by a Tukey’s post hoc multiple comparison test and provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t003" target="_blank">Table 3</a>. For information on the immunological cell markers used in flow cytometry analysis, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#sec002" target="_blank">Method</a> section and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t003" target="_blank">Table 3</a>.</p

Innate immune cells, lymphocytes, and relative gene expression characteristics in mWAT.

<p>Innate immune cells, lymphocytes, and relative gene expression characteristics in mWAT.</p

MOS supplementation did not affect body weight, fat mass, organ weight, and food intake.

<p>Body weight [A], fat mass [B], mWAT weight [C], organ weight of liver, spleen, and thymus weight [D] of mice fed a LFD or HFD with or without MOS for 17 weeks. Values are presented as means ± SEM (n = 10 mice/group). Differences were evaluated for statistical significance by two-way ANOVA or two-way ANOVA for repeated measurements, both followed by a Tukey’s post hoc multiple comparison test and provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t001" target="_blank">Table 1</a>. mWAT, mesenteric white adipose tissue.</p

MOS supplementation did not improve whole-body glucose intolerance.

<p>Whole-body glucose homeostasis was assessed in mice fed a LFD or HFD with or without MOS for 17 weeks. Fasting plasma glucose [A] and insulin levels [B] were determined in 6-hour fasted mice in week 0, 4, and 8. An ipGTT was performed in 6-hour fasted mice at week 12. Blood glucose levels were measured at the indicated minutes [C], and the area under the curve (AUC) of the glucose excursion curve was calculated as a measure for glucose tolerance [D]. Values are presented as means ± SEM (n = 10 mice/group). Differences were evaluated for statistical significance by by two-way ANOVA or two-way ANOVA for repeated measurements, both followed by a Tukey’s post hoc multiple comparison test and provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t004" target="_blank">Table 4</a>.</p

Glucose, insulin, and ipGTT characteristics.

<p>Glucose, insulin, and ipGTT characteristics.</p

MOS supplementation reduced the abundance of M2-like monocytes and increased eosinophils in mWAT.

<p>Extra-intestinal immune modulatory properties of MOS were assessed in mWAT of mice fed a LFD or HFD with or without MOS for 17 weeks. Percentages of Ly6C^hi monocytes [A], YM1+ Ly6C^hi monocytes [B] macrophages [C], macrophage M1-like and M2-like subsets [D], and eosinophils [E] within CD45+ cells in SVF of mWAT. mRNA expression of the inflammatory markers <i>F4/80</i>, <i>CD11c</i>, <i>Ym1</i>, <i>Mcp1</i>, <i>Tnf-a</i>, <i>Il-6</i>, and <i>Il-10</i> was determined [F]. Values are presented as means ± SEM (n = 6–7 mice/group). Differences were evaluated for statistical significance by by two-way ANOVA, followed by a Tukey’s post hoc multiple comparison test and provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t002" target="_blank">Table 2</a>. For information on the immunological cell markers used in flow cytometry analysis, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#sec002" target="_blank">Method</a> section and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196165#pone.0196165.t002" target="_blank">Table 2</a>.</p

Myeloid ATP Citrate Lyase Regulates Macrophage Inflammatory Responses In Vitro Without Altering Inflammatory Disease Outcomes

Macrophages are highly plastic, key regulators of inflammation. Deregulation of macrophage activation can lead to excessive inflammation as seen in inflammatory disorders like atherosclerosis, obesity, multiple sclerosis and sepsis. Targeting intracellular metabolism is considered as an approach to reshape deranged macrophage activation and to dampen the progression of inflammatory disorders. ATP citrate lyase (Acly) is a key metabolic enzyme and an important regulator of macrophage activation. Using a macrophage-specific Acly-deficient mouse model, we investigated the role of Acly in macrophages during acute and chronic inflammatory disorders. First, we performed RNA sequencing to demonstrate that Acly-deficient macrophages showed hyperinflammatory gene signatures in response to acute LPS stimulation in vitro. Next, we assessed endotoxin-induced peritonitis in myeloid-specific Acly-deficient mice and show that, apart from increased splenic Il6 expression, systemic and local inflammation were not affected by Acly deficiency. Also during obesity, both chronic low-grade inflammation and whole-body metabolic homeostasis remained largely unaltered in mice with Acly-deficient myeloid cells. Lastly, we show that macrophage-specific Acly deletion did not affect the severity of experimental autoimmune encephalomyelitis (EAE), an experimental model of multiple sclerosis. These results indicate that, despite increasing inflammatory responses in vitro, macrophage Acly deficiency does not worsen acute and chronic inflammatory responses in vivo. Collectively, our results indicate that caution is warranted in prospective long-term treatments of inflammatory disorders with macrophage-specific Acly inhibitors. Together with our earlier observation that myeloid Acly deletion stabilizes atherosclerotic lesions, our findings highlight that therapeutic targeting of macrophage Acly can be beneficial in some, but not all, inflammatory disorders