Interleukin-1 Receptor-Associated Kinase-3 Is a Key Inhibitor of Inflammation in Obesity and Metabolic Syndrome

BACKGROUND: Visceral obesity is associated with the rising incidence of type 2 diabetes and metabolic syndrome. Low-grade chronic inflammation and oxidative stress synergize in obesity and obesity-induced disorders. OBJECTIVE: We searched a cluster of molecules that support interactions between these stress conditions in monocytes. METHODS: RNA expressions in blood monocytes of two independent cohorts comprising 21 and 102 obese persons and 46 age-matched controls were determined by microarray and independently validated by quantitative RT-PCR analysis. The effect of three-month weight loss after bariatric surgery was determined. The effect of RNA silencing on inflammation and oxidative stress was studied in human monocytic THP-1 cells. RESULTS: Interleukin-1 receptor-associated kinase-3 (IRAK3), key inhibitor of IRAK/NFκB-mediated chronic inflammation, is downregulated in monocytes of obese persons. Low IRAK3 was associated with high superoxide dismutase-2 (SOD2), a marker of mitochondrial oxidative stress. A comparable expression profile was also detected in visceral adipose tissue of the same obese subjects. Low IRAK3 and high SOD2 was associated with a high prevalence of metabolic syndrome (odds ratio: 9.3; sensitivity: 91%; specificity: 77%). By comparison, the odds ratio of high-sensitivity C-reactive protein, a widely used marker of systemic inflammation, was 4.3 (sensitivity: 69%; specificity: 66%). Weight loss was associated with an increase in IRAK3 and a decrease in SOD2, in association with a lowering of systemic inflammation and a decreasing number of metabolic syndrome components. We identified the increase in reactive oxygen species in combination with obesity-associated low adiponectin and high glucose and interleukin-6 as cause of the decrease in IRAK3 in THP-1 cells in vitro. CONCLUSION: IRAK3 is a key inhibitor of inflammation in association with obesity and metabolic syndrome. Our data warrant further evaluation of IRAK3 as a diagnostic and prognostic marker, and as a target for intervention

Weight-loss-associated induction of peroxisome proliferator-activated receptor-alpha and peroxisome proliferator-activated receptor-gamma correlate with reduced atherosclerosis and improved cardiovascular function in obese insulin-resistant mice.

BACKGROUND: Weight loss in obese insulin-resistant but not in insulin-sensitive persons reduces coronary heart disease risk. To what extent changes in gene expression are related to atherosclerosis and cardiovascular function is unknown. METHODS AND RESULTS: We studied the effect of diet restriction-induced weight loss on gene expression in the adipose tissue, the heart, and the aortic arch and on atherosclerosis and cardiovascular function in mice with combined leptin and LDL-receptor deficiency. Obesity, hypertriglyceridemia, and insulin resistance are associated with hypertension, impaired left ventricular function, and accelerated atherosclerosis in those mice. Compared with lean mice, peroxisome proliferator-activated receptors (PPAR)-alpha and PPAR-gamma expression was downregulated in obese double-knockout mice. Diet restriction caused a 45% weight loss, an upregulation of PPAR-alpha and PPAR-gamma, and a change in the expression of genes regulating glucose transport and insulin sensitivity, lipid metabolism, oxidative stress, and inflammation, most of which are under the transcriptional control of these PPARs. Changes in gene expression were associated with increased insulin sensitivity, decreased hypertriglyceridemia, reduced mean 24-hour blood pressure and heart rate, restored circadian variations of blood pressure and heart rate, increased ejection fraction, and reduced atherosclerosis. PPAR-alpha and PPAR-gamma expression was inversely related to plaque volume and to oxidized LDL content in the plaques. CONCLUSIONS: Induction of PPAR-alpha and PPAR-gamma in adipose tissue, heart, and aortic arch is a key mechanism for reducing atherosclerosis and improving cardiovascular function resulting from weight loss. Improved lipid metabolism and insulin signaling is associated with decreased tissue deposition of oxidized LDL that increases cardiovascular risk in persons with the metabolic syndrome

Weight-loss-associated induction of peroxisome proliferator-activated receptor-alpha and peroxisome proliferator-activated receptor-gamma correlate with reduced atherosclerosis and improved cardiovascular function in obese insulin-resistant mice

BACKGROUND: Weight loss in obese insulin-resistant but not in insulin-sensitive persons reduces coronary heart disease risk. To what extent changes in gene expression are related to atherosclerosis and cardiovascular function is unknown. METHODS AND RESULTS: We studied the effect of diet restriction-induced weight loss on gene expression in the adipose tissue, the heart, and the aortic arch and on atherosclerosis and cardiovascular function in mice with combined leptin and LDL-receptor deficiency. Obesity, hypertriglyceridemia, and insulin resistance are associated with hypertension, impaired left ventricular function, and accelerated atherosclerosis in those mice. Compared with lean mice, peroxisome proliferator-activated receptors (PPAR)-alpha and PPAR-gamma expression was downregulated in obese double-knockout mice. Diet restriction caused a 45% weight loss, an upregulation of PPAR-alpha and PPAR-gamma, and a change in the expression of genes regulating glucose transport and insulin sensitivity, lipid metabolism, oxidative stress, and inflammation, most of which are under the transcriptional control of these PPARs. Changes in gene expression were associated with increased insulin sensitivity, decreased hypertriglyceridemia, reduced mean 24-hour blood pressure and heart rate, restored circadian variations of blood pressure and heart rate, increased ejection fraction, and reduced atherosclerosis. PPAR-alpha and PPAR-gamma expression was inversely related to plaque volume and to oxidized LDL content in the plaques. CONCLUSIONS: Induction of PPAR-alpha and PPAR-gamma in adipose tissue, heart, and aortic arch is a key mechanism for reducing atherosclerosis and improving cardiovascular function resulting from weight loss. Improved lipid metabolism and insulin signaling is associated with decreased tissue deposition of oxidized LDL that increases cardiovascular risk in persons with the metabolic syndrome.status: publishe

Characteristics and gene expressions before and after weight loss in obese patients (1^st cohort).

<p>Data shown are means ± SEM.</p><p>*<i>P</i><0.05,</p><p>**<i>P</i><0.01 and</p><p>***<i>P</i><0.001 obese compared with lean controls;</p>$<p><i>P</i><0.05,</p>$$<p><i>P</i><0.01 and</p>$$$<p><i>P</i><0.001 compared with before weight loss;</p><p>Abbreviations: BMI, body mass index; C, cholesterol; DBP, diastolic blood pressure; HOMA-IR, homeostasis model assessment of insulin resistance; hs-CRP, high sensitivity C-reactive protein; ox-LDL, oxidized LDL; SBP, systolic blood pressure.</p

Exposure of IRAK3-depleted THP-1 cells to additional stress results in more inflammation and ROS.

<p>Gene expression was analyzed using qRT-PCR and mROS production was determined by flow cytometry in THP-1 cells exposed to (<b>A</b>) 5.5 mM D-glucose and 9.5 mM D-mannitol (osmotic control) or 15 mM D-glucose (n = 6), and (<b>B</b>) 100 ng/ml IL-6 (n = 6). Data shown are means ± SEM. ^*<i>P</i><0.05, ^**<i>P</i><0.01 and ^***<i>P</i><0.001 compared with THP-1 cells exposed to 5.5 mM D-glucose or PBS vehicle. (<b>C</b>) Gene/protein expression and ROS production in THP-1 cells transiently transfected with siRNA targeting <i>IRAK3</i> (n = 10) or in THP-1 cells exposed to 100 mU/ml glucose oxidase with (n = 4) or without (n = 5) silencing of <i>IRAK3</i>. Data shown are means ± SEM. ^*<i>P</i><0.05, ^**<i>P</i><0.01 and ^***<i>P</i><0.001 compared with THP-1 control cells or THP-1 cells transfected with negative control siRNA; ^$$<i>P</i><0.01 and ^$$$<i>P</i><0.001 compared with THP-1 cells transfected with <i>IRAK3</i> siRNA; ^##<i>P</i><0.01 and ^###<i>P</i><0.001 compared with THP-1 cells exposed to glucose oxidase. Abbreviations: iROS, intracellular ROS; mROS, mitochondrial ROS; ROS, reactive oxygen species.</p

Regulation of IRAK3 expression in THP-1 monocytes.

<p>(<b>A</b>) Gene expression was analyzed by measuring relative <i>RNA</i> levels using qRT-PCR, protein expression and ROS production were determined by flow cytometry in THP-1 cells exposed to 1 or 10 µg/ml gADIPOQ (n = 6) or in <i>IRAK3</i>-depleted THP-1 cells exposed to 10 µg/ml gADIPOQ (n = 4) for 6 h and 24 h. Data shown are means ± SEM of 24 h exposed cells normalized to 6 h exposed cells. ^*<i>P</i><0.05, ^**<i>P</i><0.01 and ^***<i>P</i><0.001 compared with THP-1 cells exposed to high gADIPOQ; ^$$<i>P</i><0.01 and ^$$$<i>P</i><0.001 compared with THP-1 cells exposed to low gADIPOQ. (<b>B</b>) Gene/protein expression in THP-1 cells exposed to 10 µg/ml gADIPOQ and 10 µg/ml ox-LDL (n = 6), 1 µg/ml gADIPOQ and 25 µg/ml ox-LDL (n = 6) or 10 µg/ml gADIPOQ and 25 µg/ml ox-LDL (n = 6). Data are expressed as means ± SEM. ^**<i>P</i><0.01 compared with THP-1 cells exposed to 10 µg/ml gADIPOQ and 10 µg/ml ox-LDL; ^$<i>P</i><0.05 and ^$$<i>P</i><0.01 compared with THP-1 cells exposed to 1 µg/ml gADIPOQ and 25 µg/ml ox-LDL. Abbreviations: gADIPOQ, globular adiponectin, iROS, intracellular ROS; mROS, mitochondrial ROS; ox-LDL, oxidized LDL; ROS, reactive oxygen species.</p

Association of <i>RNA</i> expressions in monocytes and blood levels with occurrence of metabolic syndrome.

<p>*Cut points were determined by ROC curve analysis. Data are means (and 95% confidence intervals). Abbreviations: OR, odds ratio; NPV, negative predictive value; PPV, positive predictive value.</p

Gene expressions of the <i>IRAK3</i>-related pathway and adipocyte differentiation markers in visceral adipose tissue.

<p>(<b>A</b>) Gene expression in visceral adipose tissue was analyzed by measuring relative <i>RNA</i> levels using qRT-PCR for key molecules in the TLR2/NFκB inflammatory pathway. The adipose tissue specific antioxidant gene <i>SOD3</i> instead of <i>SOD2</i> was used as oxidative stress marker in visceral adipose tissue. (<b>B</b>) Relative <i>RNA</i> levels of markers of adipocyte differentiation (<i>PPARs</i> and <i>ADIPOQ</i>), insulin signaling (<i>INSR</i>) and glucose uptake (<i>GLUT4</i>) in visceral adipose tissue as determined by qRT-PCR. Data shown are means. ^*<i>P</i><0.05 and ^**<i>P</i><0.01 obese persons compared with lean controls; lean controls (n = 7), obese patients (n = 21).</p

Characteristics of obese women with and without T2DM for validation (2^nd cohort).

<p>Data shown are means ± SEM.</p><p>*<i>P</i><0.05,</p><p>**<i>P</i><0.01 and</p><p>***<i>P</i><0.001 obese compared with lean controls;</p>$<p><i>P</i><0.05,</p>$$<p><i>P</i><0.01 and</p>$$$<p><i>P</i><0.001 compared with obese women without T2DM;</p><p>Abbreviations: BMI, body mass index; C, cholesterol; DBP, diastolic blood pressure; HOMA-IR, homeostasis model assessment of insulin resistance; hs-CRP, high sensitivity C-reactive protein; ox-LDL, oxidized LDL; SBP, systolic blood pressure; T2DM, type 2 diabetes.</p