Mitsuhisa Tabata

Adiponectin and AdipoR1 regulate PGC-1α and mitochondria by Ca2+ and AMPK/SIRT1

Adiponectin is an anti-diabetic adipokine. Its receptors possess a seven-transmembrane topology with the amino terminus located intracellularly, which is the opposite of G-protein-coupled receptors. Here we provide evidence that adiponectin induces extracellular Ca2+ influx by adiponectin receptor 1 (AdipoR1), which was necessary for subsequent activation of Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ), AMPK and SIRT1, increased expression and decreased acetylation of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), and increased mitochondria in myocytes. Moreover, muscle-specific disruption of AdipoR1 suppressed the adiponectin-mediated increase in intracellular Ca2+ concentration, and decreased the activation of CaMKK, AMPK and SIRT1 by adiponectin. Suppression of AdipoR1 also resulted in decreased PGC-1α expression and deacetylation, decreased mitochondrial content and enzymes, decreased oxidative type I myofibres, and decreased oxidative stress-detoxifying enzymes in skeletal muscle, which were associated with insulin resistance and decreased exercise endurance. Decreased levels of adiponectin and AdipoR1 in obesity may have causal roles in mitochondrial dysfunction and insulin resistance seen in diabetes.

Angiopoietin-like Protein 2 Promotes Chronic Adipose Tissue Inflammation and Obesity-Related Systemic Insulin Resistance

Recent studies of obesity have provided new insights into the mechanisms underlying insulin resistance and metabolic dysregulation. Numerous efforts have been made to identify key regulators of obesity-linked adipose tissue inflammation and insulin resistance. We found that angiopoietin-like protein 2 (Angptl2) was secreted by adipose tissue and that its circulating level was closely related to adiposity, systemic insulin resistance, and inflammation in both mice and humans. Angptl2 activated an inflammatory cascade in endothelial cells via integrin signaling and induced chemotaxis of monocytes/macrophages. Constitutive Angptl2 activation in vivo induced inflammation of the vasculature characterized by abundant attachment of leukocytes to the vessel walls and increased permeability. Angptl2 deletion ameliorated adipose tissue inflammation and systemic insulin resistance in diet-induced obese mice. Conversely, Angptl2 overexpression in adipose tissue caused local inflammation and systemic insulin resistance in nonobese mice. Thus, Angptl2 is a key adipocyte-derived inflammatory mediator that links obesity to systemic insulin resistance.

Hepatic AdipoR2 signaling plays a protective role against progression of nonalcoholic steatohepatitis in mice

It is unclear how hepatic adiponectin resistance and sensitivity mediated by the adiponectin receptor, AdipoR2, contributes to the progression of nonalcoholic steatohepatitis (NASH). The aim of this study was to examine the roles of hepatic AdipoR2 in NASH, using an animal model. We fed C57BL/6 mice a methionine‐deficient and choline‐deficient (MCD) diet for up to 8 weeks and analyzed changes in liver pathology caused by either an AdipoR2 short hairpin RNA–expressing adenovirus or an AdipoR2‐overexpressing adenovirus. Inhibition of hepatic AdipoR2 expression aggravated the pathological state of NASH at all stages: fatty changes, inflammation, and fibrosis. In contrast, enhancement of AdipoR2 expression in the liver improved NASH at every stage, from the early stage to the progression of fibrosis. Inhibition of AdipoR2 signaling in the liver diminished hepatic peroxisome proliferator activated receptor (PPAR)‐α signaling, with decreased expression of acyl‐CoA oxidase (ACO) and catalase, leading to an increase in lipid peroxidation. Hepatic AdipoR2 overexpression had the opposite effect. Reactive oxygen species (ROS) accumulation in liver increases hepatic production of transforming growth factor (TGF)‐β1 at all stages of NASH; adiponectin/AdipoR2 signaling ameliorated TGF‐β–induced ROS accumulation in primary cultured hepatocytes, by enhancing PPAR‐α activity and catalase expression. Conclusion: The adiponectin resistance and sensitivity mediated by AdipoR2 in hepatocytes regulated steatohepatitis progression by changing PPAR‐α activity and ROS accumulation, a process in which TGF‐β signaling is implicated. Thus, the liver AdipoR2 signaling pathway could be a promising target in treating NASH. (HEPATOLOGY 2008;48:458–473.)

Angiopoietin‐like proteins: emerging targets for treatment of obesity and related metabolic diseases

Obesity and related metabolic diseases, such as type 2 diabetes, hypertension and hyperlipidemia are an increasingly prevalent medical and social problem in developed and developing countries. These conditions are associated with increased risk of cardiovascular disease, the leading cause of death. Therefore, it is important to understand the molecular basis underlying obesity and related metabolic diseases in order to develop effective preventive and therapeutic approaches against these conditions. Recently, a family of proteins structurally similar to the angiogenic‐regulating factors known as angiopoietins was identified and designated ‘angiopoietin‐like proteins’ (ANGPTLs). Encoded by seven genes, ANGPTL1–7 all possess an N‐terminal coiled‐coil domain and a C‐terminal fibrinogen‐like domain, both characteristic of angiopoietins. ANGPTLs do not bind to either the angiopoietin receptor Tie2 or the related protein Tie1, indicating that these ligands function differently from angiopoietins. Like angiopoietins, some ANGPTLs potently regulate angiogenesis, but ANGPTL3, ‐4 and ANGPTL6/angiopoietin‐related growth factor (AGF) directly regulate lipid, glucose and energy metabolism independent of angiogenic effects. Recently, we found that ANGPTL2 is a key adipocyte‐derived inflammatory mediator that links obesity to systemic insulin resistance. In this minireview, we focus on the roles of ANGPTL2 and ANGPTL6/AGF in obesity and related metabolic diseases, and discuss the possibility that both could function as molecular targets for the prevention and treatment of obesity and metabolic diseases.

Synoviocyte-Derived Angiopoietin-Like Protein 2 Contributes to Synovial Chronic Inflammation in Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by symmetrical polyarticular synovitis of the diarthrodial joints. Several proinflammatory cytokines derived from both infiltrating inflammatory cells and activated resident cells within the RA joint play a fundamental role in the processes that cause inflammation. However, anticytokine treatment is beneficial but not curative, the effects are only partial, and nonresponses are common. Therefore, an effort has been made to identify other key regulators of inflammation in articular structures to develop new therapies to suppress synovial inflammation and joint destruction in RA. Adipose tissue-derived angiopoietin-like protein 2 (Angptl2) activates an inflammatory cascade in endothelial cells and induces chemotaxis of monocytes/macrophages in obesity, resulting in initiation and propagation of inflammation within adipose tissues and obesity-related metabolic diseases. Angptl2 mRNA and protein are abundantly expressed in hyperplastic rheumatoid synovium of RA patients, especially in fibroblast-like and macrophage-like synoviocytes, but not in B and T lymphocytes. Angptl2 concentration in joints of RA patients was also significantly increased in comparison with patients with osteoarthritis, which in comparison with RA represents a significantly lower inflammatory grade form of arthritis. Notably, Angptl2 promoted increased chemotactic activities of CD14+CD16- monocytes from synovial fluid of RA patients. Therefore, Angptl2 acts as an important rheumatoid synovium-derived inflammatory mediator in RA pathogenesis.

Role of Endothelial Cell–Derived Angptl2 in Vascular Inflammation Leading to Endothelial Dysfunction and Atherosclerosis Progression

Objective—Cardiovascular disease (CVD), the most common morbidity resulting from atherosclerosis, remains a frequent cause of death. Efforts to develop effective therapeutic strategies have focused on vascular inflammation as a critical pathology driving atherosclerosis progression. Nonetheless, molecular mechanisms underlying this activity remain unclear. Here, we ask whether angiopoietin-like protein 2 (Angptl2), a proinflammatory protein, contributes to vascular inflammation that promotes atherosclerosis progression. Approach and Results—Histological analysis revealed abundant Angptl2 expression in endothelial cells and macrophages infiltrating atheromatous plaques in patients with cardiovascular disease. Angptl2 knockout in apolipoprotein E–deficient mice (ApoE−/−/Angptl2−/−) attenuated atherosclerosis progression by decreasing the number of macrophages infiltrating atheromatous plaques, reducing vascular inflammation. Bone marrow transplantation experiments showed that Angptl2 deficiency in endothelial cells attenuated atherosclerosis development. Conversely, ApoE−/− mice crossed with transgenic mice expressing Angptl2 driven by the Tie2 promoter (ApoE−/−/Tie2-Angptl2 Tg), which drives Angptl2 expression in endothelial cells but not monocytes/macrophages, showed accelerated plaque formation and vascular inflammation because of increased numbers of infiltrated macrophages in atheromatous plaques. Tie2-Angptl2 Tg mice alone did not develop plaques but exhibited endothelium-dependent vasodilatory dysfunction, likely because of decreased production of endothelial cell–derived nitric oxide. Conversely, Angptl2−/− mice exhibited less severe endothelial dysfunction than did wild-type mice when fed a high-fat diet. In vitro, Angptl2 activated proinflammatory nuclear factor-&kgr;B signaling in endothelial cells and increased monocyte/macrophage chemotaxis. Conclusions—Endothelial cell–derived Angptl2 accelerates vascular inflammation by activating proinflammatory signaling in endothelial cells and increasing macrophage infiltration, leading to endothelial dysfunction and atherosclerosis progression.

Angptl 4 deficiency improves lipid metabolism, suppresses foam cell formation and protects against atherosclerosis

Angiopoietin-like protein family 4 (Angptl 4) has been shown to regulate lipoprotein metabolism through the inhibition of lipoprotein lipase (LPL). We generated ApoE(-/-)Angptl 4(-/-) mice to study the effect of Angptl 4 deficiency on lipid metabolism and atherosclerosis. Fasting and postolive oil-loaded triglyceride (TG) levels were largely decreased in ApoE(-/-)Angptl 4(-/-) mice compared with and ApoE(-/-)Angptl 4(+/+) mice. There was a significant (75+/-12%) reduction in atherosclerotic lesion size in ApoE(-/-)Angptl 4(-/-) mice compared with ApoE(-/-) Angptl 4(+/+) mice. Peritoneal macrophages, isolated from Angptl 4(-/-) mice to investigate the foam cell formation, showed a significant decrease in newly synthesized cholesteryl ester (CE) accumulation induced by acetyl low-density lipoprotein (acLDL) compared with those from Angptl 4(+/+) mice. Thus, genetic knockout of Angptl 4 protects ApoE(-/-) mice against development and progression of atherosclerosis and strongly suppresses the ability of the macrophages to become foam cells in vitro.

Obesity resistance and increased hepatic expression of catabolism‐related mRNAs in Cnot3+/− mice

Obesity is a life‐threatening factor and is often associated with dysregulation of gene expression. Here, we show that the CNOT3 subunit of the CCR4–NOT deadenylase complex is critical to metabolic regulation. Cnot3+/− mice are lean with hepatic and adipose tissues containing reduced levels of lipids, and show increased metabolic rates and enhanced glucose tolerance. Cnot3+/− mice remain lean and sensitive to insulin even on a high‐fat diet. Furthermore, introduction of Cnot3 haplodeficiency in ob/ob mice ameliorated the obese phenotype. Hepatic expression of most mRNAs is not altered in Cnot3+/− vis‐à‐vis wild‐type mice. However, the levels of specific mRNAs, such as those coding for energy metabolism‐related PDK4 and IGFBP1, are increased in Cnot3+/− hepatocytes, having poly(A) tails that are longer than those seen in control cells. We provide evidence that CNOT3 is involved in recruitment of the CCR4–NOT deadenylase to the 3′ end of specific mRNAs. Finally, as CNOT3 levels in the liver and white adipose tissues decrease upon fasting, we propose that CNOT3 responds to feeding conditions to regulate deadenylation‐specific mRNAs and energy metabolism.

Angiopoietin-Related Growth Factor Enhances Blood Flow Via Activation of the ERK1/2-eNOS-NO Pathway in a Mouse Hind-Limb Ischemia Model

Objective—Transgenic mice overexpressing angiopoietin-related growth factor (AGF) exhibit enhanced angiogenesis, suggesting that AGF may be a useful drug target in ischemic disease. Our goal was to determine whether AGF enhances blood flow in a mouse hind-limb ischemia model and to define molecular mechanisms underlying AGF signaling in endothelial cells. Methods and Results—Intramuscular injection of adenovirus harboring AGF into the ischemic limb increased AGF production, which increased blood flow through induction of angiogenesis and arteriogenesis, thereby reducing the necessity for limb amputation. In vitro analysis showed that exposing human umbilical venous endothelial cells to AGF increased nitric oxide (NO) production through activation of an ERK1/2-endothelial NO synthetase (eNOS) signaling pathway. AGF-stimulated eNOS phosphorylation, NO production, and endothelial cell migration were all abolished by specific MEK1/2 inhibitors. Moreover, AGF did not restore blood flow to ischemic hind-limbs of either mice receiving NOS inhibitor L-NAME or eNOS knockout mice. Conclusion—Activation of an ERK1/2-eNOS-NO pathway is a crucial signaling mechanism by which AGF increases blood flow through induction of angiogenesis and arteriogenesis. Further investigation of the regulation underlying AGF signaling pathway may contribute to develop a new clinical strategy for ischemic vascular diseases.

Effects of epalrestat, an aldose reductase inhibitor, on diabetic peripheral neuropathy in patients with type 2 diabetes, in relation to suppression of Nɛ-carboxymethyl lysine

OBJECTIVE We investigated the efficacy of epalrestat, an aldose reductase inhibitor, for diabetic peripheral neuropathy in Japanese patients with type 2 diabetes. METHODS A total of 38 type 2 diabetic patients (22 men and 16 women; mean ± S.E.M. age 63.3 ± 1.0 years; duration of diabetes 9.6 ± 0.8 years) with diabetic neuropathy were newly administered 150 mg/day epalrestat (EP group). Motor nerve conduction velocity (MCV), sensory nerve conduction velocity (SCV), and minimum F-wave latency were evaluated before administration of epalrestat and after 1 and 2 years. Serum N(ɛ)-carboxymethyl lysine (CML) as a parameter of advanced glycation end products (AGEs), lipid peroxide, and soluble vascular cell adhesion molecule (sVCAM)-1 as a parameter of angiopathy were measured before administration and after 1 year. We compared the results with those of 36 duration of diabetes-matched type 2 diabetic patients (mean ± S.E.M. duration of diabetes 8.2 ± 0.7 years) as control (C group). RESULTS The EP group showed significant suppression of deterioration of MCV (P<.01) and minimum F-wave latency (P<.01) in the tibial nerve and SCV (P<.05) in the sural nerve compared to those in the C group after 2 years. There was a significant difference in change in CML level between groups (-0.18 ± 0.13 mU/ml in the EP group vs. +0.22 ± 0.09 mU/ml in the C group, P<.05) after 1 year. CONCLUSIONS Epalrestat suppressed the deterioration of diabetic peripheral neuropathy, especially in the lower extremity. Its effects might be mediated by improvement of the polyol pathway and suppression of production of AGEs.