Visualized macrophage dynamics and significance of S100A8 in obese fat.

Chronic low-grade inflammation of adipose tissue plays a crucial role in the pathophysiology of obesity. Immunohistological microscopic analysis in obese fat tissue has demonstrated the infiltration of several immune cells such as macrophages, but dynamics of immune cells have not been fully elucidated and clarified. Here, by using intravital multiphoton imaging technique, to our knowledge for the first time, we analyzed and visualized the inflammatory processes in adipose tissue under high-fat and high-sucrose (HF/HS) diet with lysozyme M-EGFP transgenic (LysM(EGFP)) mice whose EGFP was specifically expressed in the myelomonocytic lineage. Mobility of LysM(EGFP)-positive macrophages was shown to be activated just 5 d after HF/HS diet, when the distinct hypertrophy of adipocytes and the accumulation of macrophages still have not become prominent. Significant increase of S100A8 was detected in mature adipocyte fraction just 5 d after HF/HS diet. Recombinant S100A8 protein stimulated chemotactic migration in vitro and in vivo, as well as induced proinflammatory molecules, both macrophages and adipocytes, such as TNF-α and chemokine (C-C motif) ligand 2. Finally, an antibody against S100A8 efficiently suppressed the HF/HS diet-induced initial inflammatory change, i.e., increased mobilization of adipose LysM(EGFP)-positive macrophages, and ameliorated HF/HS diet-induced insulin resistance. In conclusion, time-lapse intravital multiphoton imaging of adipose tissues identified the very early event exhibiting increased mobility of macrophages, which may be triggered by increased expression of adipose S100A8 and results in progression of chronic inflammation in situ.

Pioglitazone suppresses neuronal and muscular degeneration caused by polyglutamine-expanded androgen receptors.

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disease caused by the expansion of a CAG repeat in the androgen receptor (AR) gene. Mutant AR has been postulated to alter the expression of genes important for mitochondrial function and induce mitochondrial dysfunction. Here, we show that the expression levels of peroxisome proliferator-activated receptor-γ (PPARγ), a key regulator of mitochondrial biogenesis, were decreased in mouse and cellular models of SBMA. Treatment with pioglitazone (PG), an activator of PPARγ, improved the viability of the cellular model of SBMA. The oral administration of PG also improved the behavioral and histopathological phenotypes of the transgenic mice. Furthermore, immunohistochemical and biochemical analyses demonstrated that the administration of PG suppressed oxidative stress, nuclear factor-κB (NFκB) signal activation and inflammation both in the spinal cords and skeletal muscles of the SBMA mice. These findings suggest that PG is a promising candidate for the treatment of SBMA.

Silencing neuronal mutant androgen receptor in a mouse model of spinal and bulbar muscular atrophy.

Spinal and bulbar muscular atrophy (SBMA), an adult-onset neurodegenerative disease that affects males, results from a CAG triplet repeat/polyglutamine expansions in the androgen receptor (AR) gene. Patients develop progressive muscular weakness and atrophy, and no effective therapy is currently available. The tissue-specific pathogenesis, especially relative pathological contributions between degenerative motor neurons and muscles, remains inconclusive. Though peripheral pathology in skeletal muscle caused by toxic AR protein has been recently reported to play a pivotal role in the pathogenesis of SBMA using mouse models, the role of motor neuron degeneration in SBMA has not been rigorously investigated. Here, we exploited synthetic antisense oligonucleotides to inhibit the RNA levels of mutant AR in the central nervous system (CNS) and explore its therapeutic effects in our SBMA mouse model that harbors a mutant AR gene with 97 CAG expansions and characteristic SBMA-like neurogenic phenotypes. A single intracerebroventricular administration of the antisense oligonucleotides in the presymptomatic phase efficiently suppressed the mutant gene expression in the CNS, and delayed the onset and progression of motor dysfunction, improved body weight gain and survival with the amelioration of neuronal histopathology in motor units such as spinal motor neurons, neuromuscular junctions and skeletal muscle. These findings highlight the importance of the neurotoxicity of mutant AR protein in motor neurons as a therapeutic target.

Paeoniflorin eliminates a mutant AR via NF-YA-dependent proteolysis in spinal and bulbar muscular atrophy.

The accumulation of abnormal proteins is a common characteristic of neurodegenerative diseases. This accumulation reflects a severe disturbance of cellular homeostasis in pathogenic protein clearance. Here, we demonstrated that the activation of the two major proteolytic machineries, the molecular chaperone-ubiquitin proteasome system (UPS) and the autophagy system, were simultaneously enhanced by paeoniflorin (PF), a major component of Paeonia plants, and exerted therapeutic effects in models of spinal and bulbar muscular atrophy (SBMA). PF significantly increased the expression of nuclear factor-YA (NF-YA), which strongly upregulated the molecules involved in the proteolytic machinery [molecular chaperones, carboxyl terminus of Hsc70-interacting protein and transcription factor EB], which thus mitigated the behavioral and pathological impairments in an SBMA mouse model through the upregulation of pathogenic androgen receptor protein clearance in motor neurons and muscles. These findings demonstrated that PF is able to enhance both the UPS and autophagy systems by upregulating the expression of NF-YA, which promotes therapeutic effects in an SBMA model.

Overexpression of hepatocyte growth factor in SBMA model mice has an additive effect on combination therapy with castration.

Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease caused by the expansion of a polyglutamine (polyQ)-encoding tract within the androgen receptor (AR) gene. The pathologic features of SBMA are motor neuron loss in the spinal cord and brainstem and diffuse nuclear accumulation and nuclear inclusions of mutant AR in residual motor neurons and certain visceral organs. Hepatocyte growth factor (HGF) is a polypeptide growth factor which has neuroprotective properties. To investigate whether HGF overexpression can affect disease progression in a mouse model of SBMA, we crossed SBMA transgenic model mice expressing an AR gene with an expanded CAG repeat with mice overexpressing HGF. Here, we report that high expression of HGF induces Akt phosphorylation and modestly ameliorated motor symptoms in an SBMA transgenic mouse model treated with or without castration. These findings suggest that HGF overexpression can provide a potential therapeutic avenue as a combination therapy with disease-modifying therapies in SBMA.

Uric acid secretion from adipose tissue and its increase in obesity.

Obesity is often accompanied by hyperuricemia. However, purine metabolism in various tissues, especially regarding uric acid production, has not been fully elucidated. Here we report, using mouse models, that adipose tissue could produce and secrete uric acid through xanthine oxidoreductase (XOR) and that the production was enhanced in obesity. Plasma uric acid was elevated in obese mice and attenuated by administration of the XOR inhibitor febuxostat. Adipose tissue was one of major organs that had abundant expression and activities of XOR, and adipose tissues in obese mice had higher XOR activities than those in control mice. 3T3-L1 and mouse primary mature adipocytes produced and secreted uric acid into culture medium. The secretion was inhibited by febuxostat in a dose-dependent manner or by gene knockdown of XOR. Surgical ischemia in adipose tissue increased local uric acid production and secretion via XOR, with a subsequent increase in circulating uric acid levels. Uric acid secretion from whole adipose tissue was increased in obese mice, and uric acid secretion from 3T3-L1 adipocytes was increased under hypoxia. Our results suggest that purine catabolism in adipose tissue could be enhanced in obesity.

Exercise attenuates polyglutamine-mediated neuromuscular degeneration in a mouse model of spinal and bulbar muscular atrophy.

BACKGROUND: Spinal and bulbar muscular atrophy (SBMA) is a hereditary neuromuscular disorder caused by the expansion of trinucleotide cytosine-adenine-guanine (CAG) repeats, which encodes a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. Recent evidence suggests that, in addition to motor neuron degeneration, defective skeletal muscles are also the primary contributors to the pathogenesis in SBMA. While benefits of physical exercise have been suggested in SBMA, underlying mechanism remains elusive. METHODS: We investigated the effect of running exercise in a transgenic mouse model of SBMA carrying human AR with 97 expanded CAGs (AR97Q). We assigned AR97Q mice to exercise and sedentary control groups, and mice in the exercise group received 1-h forced running wheel (5 m/min) 5 days a week for 4 weeks during the early stage of the disease. Motor function (grip strength and rotarod performance) and survival of each group were analysed, and histopathological and biological features in skeletal muscles and motor neurons were evaluated. RESULTS: AR97Q mice in the exercise group showed improvement in motor function (~40% and ~50% increase in grip strength and rotarod performance, respectively, P < 0.05) and survival (median survival 23.6 vs. 16.7 weeks, P < 0.05) with amelioration of neuronal and muscular histopathology (~1.4-fold and ~2.8-fold increase in motor neuron and muscle fibre size, respectively, P < 0.001) compared to those in the sedentary group. Nuclear accumulation of polyQ-expanded AR in skeletal muscles and motor neurons was suppressed in the mice with exercise compared to the sedentary mice (~50% and ~30% reduction in 1C2-positive cells in skeletal muscles and motor neurons, respectively, P < 0.05). We found that the exercise activated 5'-adenosine monophosphate-activated protein kinase (AMPK) signalling and inhibited mammalian target of rapamycin pathway that regulates protein synthesis in skeletal muscles of SBMA mice. Pharmacological activation of AMPK inhibited protein synthesis and reduced polyQ-expanded AR proteins in C2C12 muscle cells. CONCLUSIONS: Our findings suggest the therapeutic potential of exercise-induced effect via AMPK activation in SBMA.

Serum C1q- binding adiponectin in maintenance hemodialysis patients.

BACKGROUND: Patients on maintenance hemodialysis (HD) have much higher levels of adiponectin (Total-APN). Adiponectin and C1q form a protein complex in human blood, and serum C1q-binding adiponectin (C1q-APN) can be measured. We recently reported that C1q-APN/Total-APN ratio rather than Total-APN correlated with atherosclerosis in diabetics. However, the characteristics of C1q-APN in HD patients remain unclear. The preset study investigated the characteristics of the adiponectin parameters including C1q-APN and also to clarify the relationship between various serum adiponectin parameters and atherosclerotic cardiovascular diseases (ACVD) in HD patients. METHODS: The single cross-sectional study subjects were 117 Japanese patients (males/females = 61/56) on regular HD. Blood Total-APN, high molecular weight-adiponectin (HMW-APN), C1q-APN and C1q concentrations were measured by enzyme-linked immunosorbent assays. ACVD were defined as stroke, coronary and peripheral artery diseases, thoracic and abdominal aneurysms. RESULTS: Stepwise regression analysis identified high-density lipoprotein-cholesterol (HDL-C) as the only significant and independent determinant of C1q-APN in males, and duration of HD as the only significant and independent determinant of C1q-APN in females. Stepwise regression analysis identified uric acid, low-density lipoprotein-cholesterol and triglyceride as significant and independent determinants of C1q-APN/Total-APN ratio in males, and leukocyte count and HDL-C as significant and independent determinants of C1q-APN/Total-APN ratio in females. Multiple logistic regression analysis identified inorganic phosphorus and C1q-APN or C1q-APN/C1q ratio as significant determinants of ACVD. CONCLUSIONS: Low serum C1q-APN and C1q-APN/C1q ratio, but not C1q-APN/Total-APN ratio, correlated with ACVD in HD patients. TRIAL REGISTRATION: ClinicalTrials.gov: UMIN http://000004318.

High circulating levels of S100A8/A9 complex (calprotectin) in male Japanese with abdominal adiposity and dysregulated expression of S100A8 and S100A9 in adipose tissues of obese mice.

S100A8/A9 complex, calprotectin, which serves as an endogenous ligand for immune pathways, is associated with atherosclerosis. These proteins are reported to have several functions such as activating NADPH oxidase, binding toll-like receptor 4 and associated with the receptor for advanced glycation end-products. We recently reported S100A8 mRNA was highly expressed in mouse white adipose tissues and differentiated 3T3-L1 adipocytes. However, regulation of S100A9 expression in murine adipose tissue remains to be elucidated. The results of our studies in male Japanese, obese and control mice and cultured cells showed: (1) serum levels of S100A8/A9 complex, calprotectin, correlated with visceral fat area, body mass index, subcutaneous fat area, and leukocyte count in 500 Japanese men, and (2) higher mRNA expression levels of S100A8 in mature adipocyte fraction and S100A9 in stromal vascular cell fraction of obese mice, compared with those of lean mice. Overexpression of S100A8 and S100A9 in obese adipose tissue may be involved, at least partly, in not only high circulating levels of S100A8/A9 complex in abdominal obesity but also adipose and systemic tissue inflammation.

Contribution of glucocorticoid-mineralocorticoid receptor pathway on the obesity-related adipocyte dysfunction.

AIMS: Mineralocorticoid receptor (MR) blockade ameliorated insulin resistance with improvements in adipocytokine dysregulation, inflammation, and excess of reactive oxygen species (ROS) in obese adipose tissue and adipocytes, but its mechanism has not been clarified. The 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1), producing active glucocorticoids, is highly expressed in adipocytes and glucocorticoids bind to MR with higher affinity than to glucocorticoid receptor (GR). We investigated whether glucocorticoids effect on adipocytokines and ROS through MR in adipocytes. In addition, fat distributions of MR and GR were investigated in human subjects. METHODS AND RESULTS: Corticoid receptors and their target genes were examined in adipose tissue of obese db/db mice. 3T3-L1 adipocytes were treated with glucocorticoids, H(2)O(2), MR antagonist eplerenone (EP), GR antagonist RU486 (RU), MR-siRNA, and/or N-acetylcysteine. Human adipose tissues were obtained from seven patients who underwent abdominal surgery. The mRNA levels of MR and its target gene were higher in db/db mice than in control db/m+mice. In 3T3-L1 adipocytes, glucocorticoids, similar to H(2)O(2), caused the dysregulation of mRNA levels of various genes related to adipocytokines and the increase of intracellular ROS. Such changes were rectified by MR blockade, not by GR antagonist. In human fat, MR mRNA level was increased in parallel with the increase of body mass index (BMI) and its increase was more significant in visceral fat, while there were no apparent correlations of GR mRNA level to BMI or fat distribution. CONCLUSION: Glucocorticoid-MR pathway may contribute to the obesity-related adipocytokine dysregulation and adipose ROS.

Hyperinsulinemia correlates with low levels of plasma B-type natriuretic peptide in Japanese men irrespective of fat distribution.

BACKGROUND: B-type natriuretic peptide (BNP), a member of the natriuretic peptide family, is a cardiac-derived secretory hormone with natriuretic, diuretic, and vasorelaxant activities. Intraabdominal fat accumulation is associated with atherosclerotic cardiovascular diseases and cardiac dysfunction. Circulating BNP levels are relatively low (within the normal limits) in obesity and the metabolic syndrome. However, the relationship between plasma BNP levels and visceral fat accumulation in general population has not been reported. The present study analyzed the relationships between plasma BNP levels and various clinical variables, including insulin, visceral and subcutaneous fat area (VFA and SFA, respectively), in normal Japanese men. METHODS: The study (Victor-J study) subjects were consecutive 500 Japanese male workers, who underwent a health checkup and were measured VFA and SFA by computed tomography. RESULTS: Age-adjusted simple linear regression analysis showed that log-BNP correlated positively with HDL-cholesterol, and negatively with VFA, log-immunoreactive insulin (IRI), log-triglyceride, and LDL-cholesterol, but not body mass index or SFA. Stepwise multiple regression analysis identified log-IRI and HDL-cholesterol as significant determinants of log-BNP. Subjects with IRI ≥5.5 µIU/mL had lower plasma BNP levels than those with IRI < 5.5 µIU/mL, irrespective of obesity (body mass index, cutoff value 25 kg/m2), visceral fat accumulation (VFA, cutoff value 100 cm2) and subcutaneous fat accumulation (SFA, cutoff value 128 cm2). CONCLUSIONS: Our study showed that hyperinsulinemia correlated with low levels of plasma BNP in general men, irrespective of fat distribution. TRIAL REGISTRATION: UMIN 000004318.

Dynamic changes of adiponectin and S100A8 levels by the selective peroxisome proliferator-activated receptor-gamma agonist rivoglitazone.

OBJECTIVE: Accumulating evidence indicates that the regimen to increase adiponectin will provide a novel therapeutic strategy for metabolic syndrome. Here, we tested the effect of a potent and selective peroxisome proliferator-activated receptor-γ agonist, rivoglitazone (Rivo), a newly synthesized thiazolidinedione derivative, on adiponectin, insulin resistance, and atherosclerosis. METHODS AND RESULTS: ob/ob mice, apolipoprotein E knockout (apoE KO) mice, and apoE and adiponectin double knockout mice were administered pioglitazone, Rivo, or no compound. Remarkable elevation of plasma adiponectin was observed, especially in Rivo-treated ob/ob mice. Rivo ameliorated insulin resistance in ob/ob mice and reduced atherosclerotic areas in apoE KO mice compared with the pioglitazone group but failed to decrease atherosclerotic areas in double knockout mice. Among adipose mRNAs, adipose S100A8, which activates Toll-like receptor 4-dependent signal transduction cascades and locates upstream of inflammation, was markedly increased in ob/ob mice, and its increase was completely reversed by Rivo treatment. In RAW264.7 macrophage cells and 3T3-L1 adipocytes, Rivo significantly reduced S100A8 mRNA levels. CONCLUSIONS: The peroxisome proliferator-activated receptor-γ agonist Rivo remarkably enhanced adiponectin in plasma and decreased adipose S100A8 mRNA levels in obese mice. Rivo treatment apparently ameliorated insulin resistance in ob/ob mice and reduced atherosclerosis in apoE KO mice, partly through adiponectin.

The Combined Efficacy of a Two-Year Period of Cybernic Treatment With a Wearable Cyborg Hybrid-Assistive Limb and Leuprorelin Therapy in a Patient With Spinal and Bulbar Muscular Atrophy: A Case Report.

Spinal and bulbar muscular atrophy (SBMA), also known as Kennedy's disease, is a rare, slowly progressive, incurable, and hereditary neurodegenerative disease caused by the testosterone-dependent accumulation of pathogenic polyglutamine-expanded androgen receptor protein. After extensive review, two treatments for SBMA have recently been approved in Japan; this decision was based on the results of randomized controlled trials: First, anti-androgen therapy using leuprorelin acetate (leuprorelin), a disease-modifying drug that can inhibit the progression of dysphagia but has not yet been proved to improve gait function; second, cybernic treatment with a wearable cyborg hybrid assistive limb (HAL®) (Cyberdyne Inc. Tsukuba, Japan). The HAL is an innovative walking exercise system that has been shown to significantly improve gait function in eight neuromuscular diseases without reduction in muscle function, including SBMA. It is possible that the combination of these two approaches might yield better outcomes. However, the long-term effects of such a combined approach have yet to be clinically evaluated. Here, we describe the case of a 39-year-old male with SBMA who commenced anti-androgen therapy with leuprorelin 1 year previously; this was followed by cybernic treatment with HAL. The duration of walking exercise with HAL was 20-30 min a day in one session. Over 2 weeks, the patient underwent nine sessions (one course). The efficacy of HAL was evaluated by gait function tests before and after one course of cybernic treatment. Then, leuprorelin treatment was combined with cybernic sessions every 2 months for 2 years (13 courses in total). Walking ability, as evaluated by the 2-min walk test, improved by 20.3% in the first course and peaked 10 months after the commencement of combined therapy (a 59.0% improvement). Walking function was maintained throughout the period. Generally, SBMA is characterized by moderately increased serum levels of creatine kinase (CK), reflecting neuromuscular damage; interestingly, the patient's CK levels decreased dramatically with combined therapy, indicating remarkable functional improvement. Long-term combined therapy improved the patient's gait function with a steady reduction in CK levels. The combination of leuprorelin with cybernic treatment can, therefore, improve and maintain gait function without damaging the motor unit and may also suppress disease progression.

Reciprocal regulation of natriuretic peptide receptors by insulin in adipose cells.

Atrial- and brain-type natriuretic peptides (ANP and BNP, respectively) have been shown to exert potent lipolytic action in adipocytes. A family of natriuretic peptide receptors (NPRs), NPR-1, NPR-2, and NPR-3, mediates their physiologic effects. NPR-1 and NPR-2 are receptor guanylyl cyclases, while NPR-3 lacks enzymatic activity and functions primarily as a clearance receptor for natriuretic peptides. ANP has a high affinity for NPR-1 and NPR-3 than other natriuretic peptides. There is a possibility that ANP may exhibit its lipolytic effect through the balance of NPR-1 and NPR-3 expressions in adipocytes. However, the regulation of adipose NPRs has not been fully elucidated. We here examined the regulation of mouse adipose NPRs by insulin, an anti-lipolytic hormone. Among the insulin target organs, NPR-1 mRNA levels were higher in white adipose tissue (WAT) than in liver and skeletal muscle. NPR-3 mRNA was expressed most abundantly in WAT. Fasting condition induced NPR-1 mRNA level while suppressed NPR-3 mRNA level in WAT. Administration of streptozotocin resulted in the increase of NPR-1 mRNA level while the decrease of NPR-3 mRNA level in WAT. In ob/ob mice, hyperinsulinemic model, NPR-1 mRNA level was lower whereas NPR-3 mRNA level was higher compared to lean control mice. In 3T3-L1 adipocytes, insulin significantly reduced NPR-1 mRNA level while increased NPR-3 mRNA levels both through phosphatidylinositol 3-kinase (PI3-kinase) pathway. In summary, NPR-1 and NPR-3 were highly expressed in WAT and adipose NPR-1 and NPR-3 were reciprocally regulated by insulin. This study suggests that insulin may efficiently promote lipogenesis partly by reducing the lipolytic action of ANP through the opposite regulation of NPR-1 and NPR-3.

DNA methylation inhibitor attenuates polyglutamine-induced neurodegeneration by regulating Hes5.

Spinal and bulbar muscular atrophy (SBMA) is a polyglutamine-mediated neuromuscular disease caused by a CAG repeat expansion in the androgen receptor (AR) gene. While transcriptional dysregulation is known to play a critical role in the pathogenesis of SBMA, the underlying molecular pathomechanisms remain unclear. DNA methylation is a fundamental epigenetic modification that silences the transcription of various genes that have a CpG-rich promoter. Here, we showed that DNA methyltransferase 1 (Dnmt1) is highly expressed in the spinal motor neurons of an SBMA mouse model and in patients with SBMA. Both genetic Dnmt1 depletion and treatment with RG108, a DNA methylation inhibitor, ameliorated the viability of SBMA model cells. Furthermore, a continuous intracerebroventricular injection of RG108 mitigated the phenotype of SBMA mice. DNA methylation array analysis identified hairy and enhancer of split 5 (Hes5) as having a CpG island with hyper-methylation in the promoter region, and the Hes5 expression was strongly silenced in SBMA. Moreover, Hes5 over-expression rescued the SBMA cells possibly by inducing Smad2 phosphorylation. Our findings suggest DNA hyper-methylation underlies the neurodegeneration in SBMA.

Src inhibition attenuates polyglutamine-mediated neuromuscular degeneration in spinal and bulbar muscular atrophy.

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disease caused by an expanded CAG repeat in the androgen receptor (AR) gene. Here, we perform a comprehensive analysis of signaling pathways in a mouse model of SBMA (AR-97Q mice) utilizing a phosphoprotein assay. We measure the levels of 17 phosphorylated proteins in spinal cord and skeletal muscle of AR-97Q mice at three stages. The level of phosphorylated Src (p-Src) is markedly increased in the spinal cords and skeletal muscles of AR-97Q mice prior to the onset. Intraperitoneal administration of a Src kinase inhibitor improves the behavioral and histopathological phenotypes of the transgenic mice. We identify p130Cas as an effector molecule of Src and show that the phosphorylated p130Cas is elevated in murine and cellular models of SBMA. These results suggest that Src kinase inhibition is a potential therapy for SBMA.

Correlation of insulin resistance and motor function in spinal and bulbar muscular atrophy.

This study aimed to evaluate various metabolic parameters in patients with spinal and bulbar muscular atrophy (SBMA), to investigate the association between those indices and disease severity, and to explore the underlying molecular pathogenesis. We compared the degree of obesity, metabolic parameters, and blood pressure in 55 genetically confirmed SBMA patients against those in 483 age- and sex-matched healthy control. In SBMA patients, we investigated the correlation between these factors and motor functional indices. SBMA patients had lower body mass index, blood glucose, and Hemoglobin A1c, but higher blood pressure, homeostasis model assessment of insulin resistance (HOMA-IR, a marker of insulin resistance), total cholesterol, and adiponectin levels than the control subjects. There were no differences in visceral fat areas, high-density lipoprotein-cholesterol (HDL-C), or triglyceride levels in two groups. Revised amyotrophic lateral sclerosis functional rating scale (ALSFRS-R) correlated positively with HDL-C, but negatively with HOMA-IR. Through stepwise multiple regression analysis, we identified HOMA-IR as a significant metabolic determinant of ALSFRS-R. In biochemical analysis, we found that decreased expressions of insulin receptors, insulin receptor substrate-1 and insulin receptor-ß, in autopsied muscles and fibroblasts of SBMA patients. This study demonstrates that SBMA patients have insulin resistance, which is associated with the disease severity. The expressions of insulin receptors are attenuated in the skeletal muscle of SBMA, providing a possible pathomechanism of metabolic alterations. These findings suggested that insulin resistance is a metabolic index reflecting disease severity and pathogenesis as well as a potential therapeutic target for SBMA.

BIIB021, a synthetic Hsp90 inhibitor, induces mutant ataxin-1 degradation through the activation of heat shock factor 1.

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited neurodegenerative disease caused by the expansion of a polyglutamine (polyQ) tract in ataxin-1 (ATXN1). The pathological hallmarks of SCA1 are the loss of cerebellar Purkinje cells and neurons in the brainstem and the presence of nuclear aggregates containing the polyQ-expanded ATXN1 protein. Heat shock protein 90 (Hsp90) inhibitors have been shown to reduce polyQ-induced toxicity. This study was designed to examine the therapeutic effects of BIIB021, a purine-scaffold Hsp90 inhibitor, on the protein homeostasis of polyQ-expanded mutant ATXN1 in a cell culture model of SCA1. Our results demonstrated that BIIB021 activated heat shock factor 1 (HSF1) and suppressed the abnormal accumulation of ATXN1 and its toxicity. The pharmacological degradation of mutant ATXN1 via activated HSF1 was dependent on both the proteasome and autophagy systems. These findings indicate that HSF1 is a key molecule in the regulation of the protein homeostasis of the polyQ-expanded mutant ATXN1 and that Hsp90 has potential as a novel therapeutic target in patients with SCA1.

Tracing the movement of adiponectin in a parabiosis model of wild-type and adiponectin-knockout mice.

Adiponectin is exclusively synthesized by adipocytes and exhibits anti-diabetic, anti-atherosclerotic and anti-inflammatory properties. Hypoadiponectinemia is associated in obese individuals with insulin resistance and atherosclerosis. However, the mechanisms responsible for hypoadiponectinemia remain unclear. Here, we investigated adiponectin movement using hetero parabiosis model of wild type (WT) and adiponectin-deficient (KO) mice. WT mice were parabiosed with WT mice (WT-WT) or KO mice (WT-KO) and adiponectin levels were measured serially up to 63 days after surgery. In the WT-KO parabiosis model, circulating adiponectin levels of the WT partners decreased rapidly, on the other hand, those of KO partners increased, and then these reached comparable levels each other at day 7. Circulating adiponectin levels decreased further to the detection limit of assay, and remained low up to day 63. However, adiponectin protein was detected in the adipose tissues of not only the WT partner but also WT-KO mice. In the diet-induced obesity model, high adiponectin protein levels were detected in adipose stromal vascular fraction of diet-induced obese KO partner, without changes in its binding proteins. The use of parabiosis experiments shed light on movement of native adiponectin among different tissues such as the state of hypoadiponectinemia in obesity.

Accumulation of adiponectin in inflamed adipose tissues of obese mice.

OBJECTIVE: Adipose tissue inflammation plays an important role in the pathogenesis of obesity-associated complications, such as atherosclerosis. Adiponectin secreted from adipocytes has various beneficial effects including anti-inflammatory effect. Obesity often presents with hypoadiponectinemia. However, the mechanism and adiponectin movement in obesity remain uncharacterized. Here we investigated tissue distribution of adiponectin protein in lean and obese mice. METHODS: Adiponectin protein levels were evaluated by enzyme-linked immunosorbent assay and western blotting. Adipose tissues were fractionated into mature adipocyte fraction (MAF) and stromal vascular fraction (SVF). RESULTS: Adiponectin protein was detected not only in MAF but also in SVF, which lacks adiponectin mRNA expression, of adipose tissue remarkably. SVF adiponectin protein level was higher in obese mice than in lean mice. The mechanism of adiponectin accumulation was investigated in adiponectin-deficient (APN-KO) mice after injection of plasma from wild-type mice. These mice showed accumulation of exogenous adiponectin, which derived from wild type mice, in adipose tissues, and the adiponectin was more observed in SVF of diet induced obese APN-KO mice than lean APN-KO mice. Among the adiponectin binding proteins, T-cadherin mRNA and protein levels in SVF of obese mice were remarkably higher than in lean mice. Oxidative stress levels were also significantly higher in SVF of obese mice than lean mice. Mechanistically, H2O2 up-regulated T-cadherin mRNA level in murine macrophages. CONCLUSIONS: The results demonstrated adiponectin targets to adipose SVF of obese mice. These findings should shed a new light on the pathology of adipose tissue inflammation and hypoadiponectinemia of obesity.