Type I Interferon Responses Drive Intrahepatic T cells to Promote Metabolic Syndrome - PubMed (original) (raw)

. 2017 Apr 21;2(10):eaai7616.

doi: 10.1126/sciimmunol.aai7616.

Xavier S Revelo 1, Mark K Nøhr 1, Helen Luck 1 2, Kejing Zeng 1 3, Helena Lei 1, Sue Tsai 1, Stephanie A Schroer 1, Yoo Jin Park 1, Melissa Hui Yen Chng 4, Lei Shen 5, June Ann D'Angelo 6, Peter Horton 7 8, William C Chapman 9, Diane Brockmeier 10, Minna Woo 1, Edgar G Engleman 4, Oyedele Adeyi 11 12, Naoto Hirano 2 13, Tianru Jin 1, Adam J Gehring 6 14, Shawn Winer 1 12 15, Daniel A Winer 1 2 11 12

Affiliations

Type I Interferon Responses Drive Intrahepatic T cells to Promote Metabolic Syndrome

Magar Ghazarian et al. Sci Immunol. 2017.

Abstract

Obesity-related insulin resistance is driven by low-grade chronic inflammation of metabolic tissues. In the liver, non-alcoholic fatty liver disease (NAFLD) is associated with hepatic insulin resistance and systemic glucose dysregulation. However, the immunological factors supporting these processes are poorly understood. We found that the liver accumulates pathogenic CD8+ T cell subsets which control hepatic insulin sensitivity and gluconeogenesis during diet-induced obesity in mice. In a cohort of human patients, CD8+ T cells represent a dominant intrahepatic immune cell population which links to glucose dysregulation. Accumulation and activation of these cells are largely supported by type I interferon (IFN-I) responses in the liver. Livers from obese mice upregulate critical interferon regulatory factors (IRFs), interferon stimulatory genes (ISGs), and IFNα protein, while IFNαR1-/- mice, or CD8-specific IFNαR1-/- chimeric mice are protected from disease. IFNαR1 inhibitors improve metabolic parameters in mice, while CD8+ T cells and IFN-I responses correlate with NAFLD activity in human patients. Thus, IFN-I responses represent a central immunological axis that governs intrahepatic T cell pathogenicity during metabolic disease.

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Conflict of interest statement

COMPETING INTERESTS The authors have declared that they have no conflict of interest.

Figures

Figure 1

Figure 1. HFD-feeding promotes a pro-inflammatory shift in hepatic T cell populations in mice and hepatic CD8+ T cells correlate to glucose dysregulation in humans

Flow cytometry analysis of liver immune cells isolated from 16 wk HFD- or NCD-fed WT mice representing (A) total number of hepatic CD45+CD3+NK1.1- gated T cells per gram of tissue (n=16), (B) CD8+, CD4+, γδTCR+, or CD4+Foxp3+ (Treg) gated hepatic T cell subsets (n=5–16), (C) CD8+ or CD4+ CD62L− CD44+ gated effector memory T cell subsets (n=15), (D–G) intracellular T cell stains for (D) Ki-67 (n=4), (E) EdU (n=4), (F) IFNγ (n=9), and (G) TNFα (n=4) in CD8+, CD4+ or γδTCR+ gated T cells. Correlative analysis between (H) % CD8+ and % HbA1c (left), % CD4+ and HbA1c (right), and (I) % CD8+ and BMI (left), % CD4+ and BMI (right) from human intrahepatic mononuclear cells (HbA1c n=21, BMI n=30). Spearman r values and p values are denoting statistical significance are indicated in the figure graphs. Statistical significance is denoted by * (p<0.05), ** (p<0.01), and *** (p<0.001).

Figure 2

Figure 2. Intrahepatic CD8+ T cells drive hepatic insulin resistance

16 wk HFD-fed WT and CD8-deficient (CD8−/−) mice (A) representative H&E liver stains (Left, 200X magnification, Scale bar: 100μm; n=3) and liver triglyceride content (Right, n=5), (B) body weights and pyruvate tolerance test (PTT) (n=5), and (C) RT-PCR gene expression analysis of gluconeogenic enzymes (G6pc, Pepck1) and lipogenic factors (Srebp1, Pparγ) (n=6). (D) Body weights, glucose tolerance test (GTT) (n=6, 2 experiments), insulin tolerance test (ITT) (n=6, 2 experiments), and PTT (n=3) of 16 wk HFD-fed CD8−/− mice 1 week after receiving i.v. transfers of 5×106 negatively sorted intrahepatic or splenic CD8+ T cells, or PBS control. Statistical analysis is represented relative to PBS control group. (E) Representative western blot and analysis of liver lysates immunoblotted for phosphorylated and total Akt proteins after acute insulin injection of transfer mice (n=5), and (F) flow cytometry analysis of total CD8+ T cell and CD8+ effector memory T cells in liver, visceral adipose tissue (VAT), and spleens of transfer mice (n=4–6). (G) Representative western blot of pAkt/Akt and analysis of primary hepatocytes treated with 20 ng/ml IFNγ, 20 ng/ml TNFα, or both, or untreated for 24 hrs at 37°C and subsequently treated with 100 nM insulin for 15 min (n=4). Western blot analysis is represented as pAkt/Akt expression relative to control (insulin) group. Statistical significance is denoted by * (p<0.05), ** (p<0.01), and *** (p<0.001).

Figure 3

Figure 3. HFD-feeding induces a type I interferon response in the liver that promotes hepatic T cell inflammation and insulin resistance

(A) RT-PCR gene expression analysis of type I interferon (IFN-I) related interferon regulatory factors (IRFs) and interferon stimulatory genes (ISGs) (left, n=8–12), and quantification of IFNα protein (right, n=5) in liver lysates of 16 wk HFD- or NCD-fed WT mice (n=7–10). (B) Gene expression analysis of ISGs in CD8+ T cells from the livers of 16 wk HFD- or NCD-fed WT mice (n=5). (C–K) 16 wk HFD-fed WT and IFNαR1-deficient (IFNαR1−/−) mice. Representative H&E stain of livers (C, Left; 100X magnification, Scale bar: 100μm) (n=3), liver weights (C Middle, n=5), liver triglycerides (C Right, n=5), VAT weights (D Left), body weights (D Right), GTT (E Left; n=4 for WT, n=5 for KO), ITT (E Middle, n=5) and PTT (E Right, n=5), fasting glucose (F Left, n=4 for WT, n=5 for KO) and insulin (F Right, n=8–12), representative western blot and analysis of pAkt/Akt in liver lysates after acute insulin challenge in mice (G, n=4–5), flow cytometry analysis of T cell subsets (H, n=5), and intracellular T cell staining for (I) Ki-67, (J) IFNγ and (K) TNFα (n=3) using previously stated gating strategies. Western blot analysis is represented as pAkt/Akt expression relative to control group. Statistical significance is denoted by * (p<0.05), ** (p<0.01) and *** (p<0.001).

Figure 4

Figure 4. Intrahepatic type I interferon signaling and responsive CD8+ T cells promote metabolic disease in mice and correlate to worsened NAFLD in humans

(A) Body weights, GTT, ITT and PTT (n=5), and (B) flow cytometry analysis of IFNαR1 expression on CD8+ T cells (representative plot) and hepatic T cell subsets (n=3–4) in WT HFD-fed mice that were treated i.p. with 400 μg/wk of anti-IFNαR1 blocking antibody, or IgG control, for 5 wks beginning at 10 wks of HFD-feeding. (C) Body weight (n=10), GTT (n=10), ITT (n=5), PTT (n=10), (D) flow cytometry analysis of IFNαR1 expression on CD8+ T cells (representative plot) and (E) hepatic T cell subsets in 16 wk HFD-fed BM chimeric mice containing (WT CD8) or lacking (IFNαR1−/− CD8) IFNαR1 expression in CD8+ T cells (n=10). T cells were analyzed using previously stated gating strategies. (F) Representative immunohistochemical staining of human liver biopsies (n=18) given NAS scores and stained single stained for IRF3 (brown), or double stained for ISG15 (pink) and CD8 (brown) (200X magnification, Scale bar: 100μm), and correlative analysis between (G) NAS Score and IRF3 (top, n=19) or ISG15 (bottom, n=18) protein expression, and (H) NAS Score and lobular (top) or portal tract (bottom) CD8+ cells/high power field (HPF = 0.237 mm2) (n=18). Arrows indicate CD8+ cells. Spearman r values and p values denoting statistical significance are indicated in the figure graphs. Statistical significance is denoted by * (p<0.05), and ** (p<0.01).

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