Adiponectin Treatment Research Articles

Effect of chronic intermittent hypoxia on genioglossus and its intervention by adiponectin

To investigate the effects of chronic intermittent hypoxia (CIH) on electromyograph (EMG) and ultrastructure of genioglossus (GG) and the interventive effects with adiponectin supplement. Forty-two healthy male Wistar rats were randomly divided into normal control (A), CIH (B) and adiponectin treatment (C) groups with 14 rats in each. CIH was performed 8 hours per day for 5 weeks in both group B and C. In group C, transvenous injection of adiponectin of 10 microg dosage each time, twice a week for 5 weeks. While in group A and B, transvenous injection of saline was performed twice a week for 5 weeks. At the beginning of 6th week the GG EMG voltages were measured before, during and following hypoxia stimulation by inserted bipolar needle electrodes and compared among three groups. Transmission electron microscope was used for observation of ultrastructure of GG. The serum adiponectin level in group B (1226.0 +/- 112.0) ng/ml (x(-) +/- s) was significantly lower than that in group A (2491.8 +/- 117.9) ng/ml, q = 38.2, P < 0.01), and adiponectin level in group C (1988.3 +/- 114.7) ng/ml was significantly higher than that in group B (q = 23.0, P < 0.01). Comparison of GG EMG activity showed that the baseline amplitude of GG EMG before hypoxia stimulation was significantly lower in group B than that in both group A and group C (all P < 0.01). At the 5th min of hypoxia stimulation the GG EMG activities were significantly enhanced among three groups (all P < 0.01). Such an enhancement was the most evident in group A but the least remarkable in group B, with a significant difference among three groups (q(ab) = 17.5; q(ac) = 8.9; q(bc) = 8.6, all P < 0.01). 15 min, 30 min and 45 min after hypoxia stimulation the amplitude of GG EMG remained at relative higher levels in group A and C, significantly higher than that in group B (all P < 0.01). CIH could cause significant ultrastructural pathological changes such as myofibril discontinuities, lysis of myofilament, edema of mitochondria and disruption of cristae, vacuoles and lysis of some mitochondria in group B. Venous supplement of adiponectin could improve pathological changes resulting from CIH. CIH could resulted in pathological changes in EMG and ultrastructure of GG, which might be associated with hypoadiponectinemia caused by CIH.

Adiponectin increases motility of human prostate cancer cells via adipoR, p38, AMPK, and NF‐κB pathways

Prostate cancer is the most commonly diagnosed malignancy in men. Prostate cancer shows a predilection for metastasis to the bone. Adiponectin is a protein hormone secreted predominantly by differentiated adipocytes and involved in energy homeostasis. The aim of this study was to investigate whether adiponectin is associated with migration of prostate cancer cells. Cancer cells migration activity was examined using the Transwell assay. The p38 and AMPK phosphorylation was examined by using Western blot method. The cell surface expression of integrins was examined by using flow cytometry. The qPCR was used to examine the mRNA expression of integrin. A transient transfection protocol was used to examine NF-kappaB activity. We found that adiponectin increased the migration and the expression of alpha5beta1 integrin of human prostate cancer cells. Adiponectin-mediated migration and integrins expression was attenuated by p38 inhibitor (SB203580), p38 mutant, AMPK siRNA, AMPK inhibitor (araA and compound C). Activations of p38, AMPK and NF-kappaB pathways after adiponectin treatment was demonstrated, and adiponectin-induced expression of integrins and migration activity was inhibited by the specific inhibitor and mutant of p38, AMPK, and NF-kappaB cascades. This study showed for the first time that the adiponectin mediates migration of human prostate cancer cells. One of the mechanisms underlying adiponectin directed migration was transcriptional up-regulation of alpha5beta1 integrin and activation of AdipoR1 receptor, p38, AMPK, and NF-kappaB pathways.

Involvement of adiponectin and leptin in breast cancer: clinical and in vitro studies

Obesity is a risk factor for breast cancer development. A recent hypothesis suggests that the adipokines, adiponectin and leptin, are involved in breast cancer development. This prompted us to investigate the role of adiponectin and leptin in mammary carcinogenesis. Adiponectin receptors (AdipoR1 and AdipoR2) and leptin receptor (Ob-Rt, representing all the isoforms of Ob-R) proteins were detected by immunohistochemistry in in situ ductal carcinoma, invasive ductal malignancy, and healthy adjacent tissue. In addition, mRNA expression of adiponectin, AdipoR1, AdipoR2, leptin, Ob-Rt, and Ob-Rl (the long isoform of Ob-R) was observed in MCF-7 breast cancer cells. Interestingly, leptin mRNA expression was 34.7-fold higher than adiponectin mRNA expression in the MCF-7 cell line. Moreover, adiponectin (10 microg/ml) tended to decrease the mRNA expression of leptin (-36%) and Ob-Rl (-28%) and significantly decreased Ob-Rt mRNA level (-26%). In contrast, leptin treatment (1 microg/ml) significantly decreased AdipoR1 mRNA (-23%). Adiponectin treatment (10 microg/ml) inhibited the proliferation of MCF-7 cells, whereas leptin (1 microg/ml) stimulated the growth of cancer cells. In addition, adiponectin inhibited leptin-induced cell proliferation (both 1 microg/ml). Using microarray analysis, we found that adiponectin reduced the mRNA levels of genes involved in cell cycle regulation (mitogen-activated protein kinase 3 and ATM), apoptosis (BAG1, BAG3, and TP53), and potential diagnosis/prognosis markers (ACADS, CYP19A1, DEGS1, and EVL), whereas leptin induced progesterone receptor mRNA expression. In conclusion, the current study indicates an interaction of leptin- and adiponectin-signaling pathways in MCF-7 cancer cells whose proliferation is stimulated by leptin and suppressed by adiponectin.

Adiponectin suppresses tumorigenesis in Apc Min/+ mice

Recent reports have shown that adiponectin has a suppressive effect on various types of malignancy. In order to clarify the role of adiponectin in colorectal carcinogenesis, we examined the effect of exogenous administration of adiponectin on intestinal polyp formation in C57BL/6 J- Apc Min /+ mice, which possess a point mutation in the Apc gene. And we found that adiponectin treatment significantly decreased the number of adenomatous polyps, especially polyps larger than 2 mm in diameter, in the small intestine. Two major receptors for adiponectin, AdipoR1 and AdipoR2, were expressed in adenomatous polyps, and their expression levels were not altered by adiponectin injection. In conclusion, adiponectin suppresses the growth of intestinal adenomas in the Apc Min /+ mice. Increasing the adiponectin level may be a new strategy for the prevention of colorectal cancer at an early step of carcinogenesis.

Subcellular Localization of Cyclic AMP-Responsive Element Binding Protein-Regulated Transcription Coactivator 2 Provides a Link between Obesity and Breast Cancer in Postmenopausal Women

Epidemiologic evidence supports a correlation between obesity and breast cancer in women. AMP-activated protein kinase plays an important role in energy homeostasis and inhibits the actions of cyclic AMP-responsive element binding protein-regulated transcription coactivator 2 (CRTC2). In postmenopausal women, the cyclic AMP-responsive element binding protein-dependent regulation of aromatase is a determinant of breast tumor formation through local production of estrogens. The present work aimed to examine the effect of adipokines on aromatase expression and identify additional mechanisms by which prostaglandin E(2) causes increased aromatase expression in human breast adipose stromal cells. Treatment of human adipose stromal cells with forskolin and phorbol 12-myristate 13-acetate (PMA), to mimic prostaglandin E(2), resulted in nuclear translocation of CRTC2. Aromatase promoter II (PII) activity assays showed that CRTC2 in addition to forskolin/PMA treatment significantly increased PII-induced activity. CRTC2 binding to PII was examined by chromatin immunoprecipitation, and forskolin/PMA treatment was associated with increased binding to PII. Treatment of human adipose stromal cells with leptin significantly up-regulated aromatase expression associated with nuclear translocation of CRTC2 and increased binding of CRTC2 to PII. Adiponectin treatment significantly decreased forskolin/PMA-stimulated aromatase expression, consistent with the decreased nuclear translocation of CRTC2 and the decreased binding of CRTC2 to PII. The expression and activity of the AMP-activated protein kinase LKB1 was examined and found to be significantly decreased following either forskolin/PMA or leptin treatment. In contrast, adiponectin significantly increased LKB1 expression and activity. In conclusion, the regulation of aromatase by CRTC2, in response to the altered hormonal milieu associated with menopause and obesity, provides a critical link between obesity and breast cancer.

Involvement of AdipoR receptor in adiponectin-induced motility and α2β1 integrin upregulation in human chondrosarcoma cells

Chondrosarcoma is a type of highly malignant tumor with a capacity to invade locally and cause distant metastasis. Chondrosarcoma shows a predilection for metastasis to the lungs. Adiponectin is a protein hormone secreted predominantly by differentiated adipocytes and is involved in energy homeostasis. However, the effect of adiponectin on migration activity in human chondrosarcoma cells is mostly unknown. We found that adiponectin increased the migration and expression of alpha2beta1 integrin in human chondrosarcoma cells. The protein and messenger RNA expression of adiponectin receptor (AdipoR1 and AdipoR2) in chondrosarcoma patients and chondrosarcoma cell lines were significantly higher than the normal cartilage. Moreover, primary chondrosarcoma and chondrosarcoma cell lines (SW1353 and JJ012) were more invasive than normal chondrocytes. Adiponectin-mediated migration and integrin expression was attenuated by 5'-adenosine monophosphate-activated protein kinase (AMPK) small interfering RNA and an AMPK inhibitor (Ara A and compound C). Activation of p38 and nuclear factor-kappa B (NF-kappaB) pathways after adiponectin treatment was demonstrated, and adiponectin-induced expression of integrins and migration activity was inhibited by the specific inhibitor and mutant of p38 and NF-kappaB cascades. This study showed for the first time that adiponectin mediates the migration of human chondrosarcoma cells. One mechanism underlying adiponectin-directed migration was transcriptional upregulation of alpha2beta1 integrin and activation of AdipoR receptor, AMPK, p38 and NF-kappaB pathways.

LKB1 is required for adiponectin-mediated modulation of AMPK–S6K axis and inhibition of migration and invasion of breast cancer cells

Adiponectin is widely known as an adipocytokine with therapeutic potential for its markedly protective function in the pathogenesis of obesity-related disorders, metabolic syndrome, systemic insulin resistance, cardiovascular disease and more recently carcinogenesis. In the present study, we show that adiponectin inhibits adhesion, invasion and migration of breast cancer cells. Further analysis of the underlying molecular mechanisms revealed that adiponectin treatment increased AMP-activated protein kinase (AMPK) phosphorylation and activity as evident by increased phosphorylation of downstream target of AMPK, acetyl-coenzyme A carboxylase and inhibition of p70S6 kinase (S6K). Intriguingly, we discovered that adiponectin treatment increases the expression of tumor suppressor gene LKB1 in breast cancer cells. Overexpression of LKB1 in breast cancer cells further increased adiponectin-mediated phosphorylation of AMPK. Using isogenic LKB1 knockdown cell line pair, we found that LKB1 is required for adiponectin-mediated modulation of AMPK-S6K axis and more importantly, inhibition of adhesion, migration and invasion of breast cancer cells. Taken together these data present a novel mechanism involving specific upregulation of tumor suppressor gene LKB1 by which adiponectin inhibits adhesion, invasion and migration of breast cancer cells. Our findings indicate the possibility of using adiponectin analogues to inhibit invasion and migration of breast cancer cells.

Adiponectin gene single-nucleotide polymorphisms and treatment response to obesity

In the adiponectin gene polymorphisms, single-nucleotide polymorphism (SNP)-45 and SNP276 have reportedly been associated with obesity, Type 2 diabetes, and other features of metabolic syndrome. Whether these adiponectin SNP affect obesity-related parameters during caloric restriction in obese subjects. Thirty- two obese Japanese women were treated by meal replacement with a low calorie diet for 8 weeks and asked to maintain their habitual lifestyle. Obesity-related parameters were measured before and after the treatment period. We determined four SNP (T45G, I164T, G276T, and C-11377G) using a fluorescent allele-specific DNA primer assay systemand FRET probe assay system. After the treatment, the extent of decrease in waist circumference was greater in the subjects with the G/G or G/T genotype of SNP276 than in those with the T/T genotype (p=0.026). As for SNP45, the extent of decrease in triglyceride levels was greater in the subjects with the T/T genotype than in those with the T/G genotype (p=0.003). For SNP-11377, the extent of decrease in systolic blood pressure and fasting plasma glucose was greater in the subjects with the C/G or G/G genotype than in those with the C/C genotype (p=0.044). Our findings indicate that each SNP in the adiponectin gene might modify the change in obesity-related parameters during meal replacement with a low calorie diet.

Modulation of fatty acid induced oxidative stress by adiponectin in 3T3 L1 adipocytes

Stimulation of toll‐like receptor signaling by palmitate increases inflammation in adipocytes. Adiponectin is an anti‐inflammatory adipokine. We hypothesized that treatment of adipocytes with palmitate (0.5 mM) would increase production of ROS and transcript levels of iNOS, TLR2 and 4 while decreasing transcript levels of IL10, and that co‐treatment with adiponectin (30 ?g/mL Adn + Palm) would alleviate inflammation and normalize gene expression. We used 10‐day post differentiation mouse 3T3 L1 adipocytes for our studies. Adiponectin decreased ROS production (p=0.0395) while palmitate did not affect ROS production. Treatment with Adn + Palm decreased ROS production compared to palmitate (Palm) alone (p=0.0156). Adiponectin treatment did not affect iNOS, TLR2, or TLR4 transcript levels. Palmitate treatment did not affect TLR2 transcript levels, but there was a trend toward an increase in both iNOS and TLR4 transcript expression (p=0.0616 and 0.0697, respectively). Adiponectin did not alter IL10 expression. Palmitate reduced IL10 transcript abundance as compared with the vehicle control (p=0.0111), and adiponectin reversed this effect. These data suggest that palmitate reduces anti‐inflammatory cytokine production and increases oxidative stress in adipocytes. Adiponectin may have an important role in attenuating the adverse effects of palmitate.

The Cardioprotective Effects of Adiponectin in the ZDF Heart Following Ischemia Are Associated with a Significant Attenuation of Myocardial ICAM‐1 Expression

Ischemic heart disease (IHD) is significantly enhanced in type 2 diabetes (T2D) and inflammation is associated with IHD and T2D. The adipocytokine, adiponectin (APN), which has anti‐inflammatory actions, is chronically reduced in type 2 diabetes. APN may link inflammation, T2D and IHD. We examined if a globular form of adiponectin was cardioprotective in the Zucker Diabetic Fatty (ZDF) rodent model of T2D and if the cardioprotection was associated with myocardial expression of ICAM‐1, a key adhesion molecule and marker of coronary microvascular dysfunction. ZDF (n=14) rats and their lean controls ZLC (n=13) were subjected to 30 minutes of LAD occlusion and 120 minutes of reperfusion. Animals were treated with either PBS or recombinant globular adiponectin (1.0 μg/g). At R120, the extent of myocardial infarction was measured. Myocardial ICAM‐1 expression was also quantified immunohistochemically. We found that adiponectin treatment significantly attenuated the enhanced myocardial infarction in the ZDF model of T2D (p&lt;0.05). The reduced infarction was associated with a significant reduction in myocardial ICAM‐1 expression (p&lt;0.05). These findings suggest that adiponectin treatment protects against I/R injury in the T2D heart. The cardioprotective action may be related to reduced coronary microvascular inflammation enhancing PMN sequestration and PMN‐mediated tissue injury. Supported by NIH HL 58859.

Adiponectin Improves Cardiomyocyte Contractile Function in db/db Diabetic Obese Mice

Low levels of adiponectin, a fat-derived hormone, are found to be correlated with coronary heart disease, type 2 diabetes, obesity, and insulin resistance. Conversely, high adiponectin levels are predictive of reduced coronary risk in long-term epidemiologic studies. However, the precise role of adiponectin in cardiomyocyte function is still not clear. This study was designed to examine the role of adiponectin in cardiac contractile function in the db/db model of diabetic obesity. Mechanical properties and intracellular Ca(2+) transients were evaluated in cardiomyocytes from lean control and db/db mice with or without adiponectin (10 microg/ml) treatment. Expression and phosphorylation of IRS-1, Akt, c-Jun, and c-Jun N terminal kinase (JNK) as well as markers of endoplasmic reticulum (ER) stress were evaluated using western blotting. Cardiomyocytes from db/db mice exhibited greater cross-sectional area, depressed peak shortening (PS), and maximal velocity of shortening/re-lengthening as well as prolonged duration of re-lengthening. Consistently, myocytes from db/db mice displayed a reduced electrically stimulated rise in intracellular Ca(2+) and prolonged intracellular Ca(2+) decay, which were abrogated by adiponectin treatment. Ratios between phosphorylated c-Jun and c-Jun as well as phosphorylated IRS-1 and IRS-1 were increased in db/db mice, the effect of which was attenuated by adiponectin. Levels of the phosphorylated ER stress makers PERK (Thr980), IRE-1, and eIF2alpha were significantly elevated in db/db mice compared with lean controls, although the effect was unaffected by adiponectin. Collectively, our data suggest that adiponectin improves cardiomyocyte dysfunction in db/db diabetic obese mice through a mechanism possibly related to c-Jun and IRS-1.

Adiponectin activates tumor suppressor LKB1 and AMP-activated protein kinase signaling to inhibit malignant properties of breast cancer cells.

Abstract Abstract #3067 Introduction: The prevalence of obesity in the developed world has reached epidemic proportions in recent years. Obese breast cancer patients exhibit a higher risk for metastasis, larger tumor burden and poorer response to endocrine therapy. Therefore, it is very important to understand the adverse effects of obesity on breast cancer in order to devise appropriate new approaches to their treatment. Obesity is considered an endocrine disorder and serum levels of adiponectin, an adipocytokine proposed to have therapeutic potential, get significantly lowered with obesity. More recently, low serum adiponectin levels were associated with increased risk and more aggressive phenotype (larger tumor size and high histological grade) of breast cancer in women. Recently, anti-proliferative and pro-apoptotic response of adiponectin was shown in breast cancer cells but the molecular mechanisms governing the anti-cancer effects of adiponectin largely remain unknown. In the present study, we found a novel mechanism underlying adiponectin action involving upregulation of tumor suppressor LKB1 and activation of AMPK (AMP-activated protein kinase) signaling to inhibit malignant properties of breast cancer cells.&amp;#x2028; Methods and Results: Adiponectin inhibited malignant properties such as invasion and migration of breast cancer cells in matrigel invasion, scratch migration and quantitative Electric Cell-Substrate Impedance Sensing (ECIS) invasion and migration assays. Interestingly, adiponectin induced phosphorylation of AMPK at Thr 172 in breast cancer cells. Adiponectin treatment also affected downstream targets of mTOR signaling pathway, including decreased phosphorylation of p70S6K showing the involvement of mTOR-S6K axis. Intriguingly, we found that adiponectin increased the expression of tumor suppressor LKB1 in breast cancer cells. Co-immunoprecipitation assays revealed that LKB1 co-immunoprecipitated with tumor suppressor TSC2. Increase in expression of upstream kinase LKB1 might directly regulate adiponectin induced activation of AMPK. We also found that breast cancer cells transfected with wild-type LKB1 plasmids increased phosphorylation of AMPK and decreased phosphorylation of p70S6K showing decrease in mTOR activity. In contrast, breast cancer cells transfected with kinase dead LKB1 decreased phosphorylation of AMPK and increased activation of p70S6K.&amp;#x2028; Discussion: Taken together these data indicate a novel crosstalk between adiponectin and tumor suppressor LKB1 that modulates AMPK-S6K axis and inhibits metastatic properties, invasion and migration of breast cancer cells. The influence of adipocytokine, adiponectin on breast carcinogenesis is just beginning to be elucidated and our study is the first to show the direct involvement of tumor suppressor genes LKB1 and TSC2 in anti-cancer effects of adiponectin. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3067.

Taurine supplementation prevents ethanol-induced decrease in serum adiponectin and reduces hepatic steatosis in rats #

Chronic ethanol feeding decreases expression of adiponectin by adipocytes and circulating adiponectin. Adiponectin treatment during chronic ethanol feeding prevents liver injury in mice. Chronic ethanol feeding also increases oxidative and endoplasmic reticulum (ER) stress in adipose tissue. Here we tested the hypothesis that supplemental taurine, an amino acid that functions as a chemical chaperone/osmolyte and enhances cellular antioxidant activity, would prevent ethanol-induced decreases in adiponectin expression and attenuate liver injury. Serum adiponectin concentrations decreased as early as 4 to 7 days after feeding rats a 36% ethanol diet. This rapid decrease was associated with increased oxidative, but not ER, stress in subcutaneous adipose tissue. Taurine prevented ethanol-induced oxidative stress and increased inflammatory cytokine expression in adipose tissue. Ethanol feeding also rapidly decreased expression of transcription factors regulating adiponectin expression (CCAAT/enhancer binding protein alpha; peroxisome proliferator-activated receptor alpha/gamma) in subcutaneous adipose tissue. Taurine prevented the ethanol-induced decrease in CCAAT/enhancer binding protein alpha and peroxisome proliferator-activated receptor alpha, normalizing adiponectin messenger (m)RNA and serum adiponectin concentrations. In the liver, taurine prevented ethanol-induced oxidative stress and attenuated tumor necrosis factor alpha expression and steatosis, at least in part, by increasing expression of genes involved in fatty acid oxidation. In subcutaneous adipose tissue, taurine decreased ethanol-induced oxidative stress and cytokine expression, as well as normalized expression of adiponectin mRNA. Taurine prevented ethanol-induced decreases in serum adiponectin; normalized adiponectin was associated with a reduction in hepatic oxidative stress, tumor necrosis factor alpha expression, and steatosis. Taken together, these data demonstrate that taurine has important protective effects against ethanol-induced tissue injury in both adipose and liver tissue.

Adiponectin increases insulin-like growth factor I-induced progesterone and estradiol secretion in human granulosa cells

To identify adiponectin and its receptors (AdipoR1 and AdipoR2) in human granulosa cells (GC) and to study the effects of recombinant human adiponectin on P and E(2) secretion from these cells. The effects of recombinant human adiponectin on the secretion of P and E(2) by cultured human GCs were investigated. Academic institutions. Seventeen infertile and healthy women undergoing IVF. Primary human GC cultures stimulated with human recombinant adiponectin (5 microg/mL). Determination of messenger RNA (mRNA) and protein expression of adiponectin and its receptors AdipoR1 and AdipoR2 in fresh human GCs by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblot, respectively. Measurement of P and E(2) levels in the conditioned media by RIA and determination of cell proliferation by tritied thymidine incorporation. Human GCs express adiponectin receptors AdipoR1 and AdipoR2 but not adiponectin. In primary human GCs, adiponectin increases P and E(2) secretion in response to insulin-like growth factor I (IGF-I). This was associated with an increase in the p450 aromatase protein level but not those of p450scc, 3 beta HSD, or StAR. Adiponectin treatment does not affect IGF-1-induced cell proliferation and basal steroidogenesis (no IGF-1 or FSH stimulation). Adiponectin rapidly stimulates MAPK ERK1/2 and p38 phosphorylation in primary human GCs. Adiponectin receptors AdipoR1 and AdipoR2, but not adiponectin, are present in human GCs. Adiponectin increases IGF-1-induced P and E(2) secretion in primary human GCs.

Adenovirus-mediated gene transfer of adiponectin reduces the severity of collagen-induced arthritis in mice

Adiponectin (APN) is a hormone released by adipose tissue with anti-inflammatory properties. The purpose of this study was to examine the therapeutic effects of systemic delivery of APN in murine arthritis model. Collagen-induced arthritis (CIA) was induced in male DBA1/J mice, and adenoviral vectors encoding human APN (Ad-APN) or beta-galactosidase (Ad-β-gal) as control were injected either before or during arthritis progression. Systemic APN delivery at both time points significantly decreased clinical disease activity scores of CIA. In addition, APN treatment before arthritis progression significantly decreased histological scores of inflammation and cartilage damage, bone erosion, and mRNA levels of pro-inflammatory cytokines in the joints, without altering serum anti-collagen antibodies levels. Immunohistochemical staining showed significant inhibition of complement C1q and C3 deposition in the joints of Ad-APN infected CIA mice. These results provide novel evidence that systemic APN delivery prevents inflammation and joint destruction in murine arthritis model.

Endothelial dysfunction in adiponectin deficiency and its mechanisms involved

Endothelial dysfunction is the earliest pathologic alteration in diabetic vascular injury and plays a critical role in the development of atherosclerosis. Plasma levels of adiponectin (APN), a novel vasculoprotective adipocytokine, are significantly reduced in diabetic patients, but its relationship with endothelial dysfunction remains unclear. The present study aims to determine whether APN deficiency may cause endothelial dysfunction and to investigate the involved mechanisms. Vascular rings were made from the aortic vessels of wild type (WT) or APN knockout (APN −/−) mice. Endothelial function, total NO production, eNOS expression/phosphorylation, superoxide production, and peroxynitrite formation were determined. ACh and acidified NaNO 2 (endothelial dependent and independent vasodilators, respectively) caused similar concentration-dependent vasorelaxation in WT vascular rings. APN −/− rings had a normal response to acidified NaNO 2, but a markedly reduced response to ACh (> 50% reduction vs. WT, P < 0.01). Both superoxide and peroxynitrite production were increased in APN −/− vessels ( P < 0.01 vs. WT). Pretreatment with superoxide scavenger Tiron significantly, but incompletely restored vascular vasodilatory response to ACh. In APN −/− vessels, eNOS expression was normal, but NO production and eNOS phosphorylation was significantly reduced ( P < 0.01 vs. WT). Treatment of APN −/− mice in vivo with the globular domain of adiponectin reduced aortic superoxide production, increased eNOS phosphorylation, and normalized vasodilatory response to ACh. Increased NO inactivation combined with decreased basal NO production contributes to endothelial dysfunction development when there is a paucity of APN production. Interventions directed towards increasing plasma APN levels may improve endothelial function, and reduce cardiovascular complications suffered by diabetic patients.

Adiponectin receptors are expressed in hypothalamus and colocalized with proopiomelanocortin and neuropeptide Y in rodent arcuate neurons

Adiponectin is involved in the control of energy homeostasis in peripheral tissues through Adipor1 and Adipor2 receptors. An increasing amount of evidence suggests that this adipocyte-secreted hormone may also act at the hypothalamic level to control energy homeostasis. In the present study, we observed the gene and protein expressions of Adipor1 and Adipor2 in rat hypothalamus using different approaches. By immunohistochemistry, Adipor1 expression was ubiquitous in the rat brain. By contrast, Adipor2 expression was more limited to specific brain areas such as hypothalamus, cortex, and hippocampus. In arcuate and paraventricular hypothalamic nuclei, Adipor1, and Adipor2 were expressed by neurons and astrocytes. Furthermore, using transgenic green fluorescent protein mice, we showed that Adipor1 and Adipor2 were present in pro-opiomelanocortin (POMC) and neuropeptide Y (NPY) neurons in the arcuate nucleus. Finally, adiponectin treatment by intracerebroventricular injection induced AMP-activated protein kinase (AMPK) phosphorylation in the rat hypothalamus. This was confirmed by in vitro studies using hypothalamic membrane fractions. In conclusion, Adipor1 and Adipor2 are both expressed by neurons (including POMC and NPY neurons) and astrocytes in the rat hypothalamic nuclei. Adiponectin is able to increase AMPK phosphorylation in the rat hypothalamus. These data reinforced a potential role of adiponectin and its hypothalamic receptors in the control of energy homeostasis.

Adiponectin upregulates ferritin heavy chain in skeletal muscle cells.

OBJECTIVE—Adiponectin is an adipocyte-derived protein that acts to reduce insulin resistance in the liver and muscle and also inhibits atherosclerosis. Although adiponectin reportedly enhances AMP-activated protein kinase and inhibits tumor necrosis factor-α action downstream from the adiponectin signal, the precise physiological mechanisms by which adiponectin acts on skeletal muscles remain unknown.RESEARCH DESIGN AND METHODS—We treated murine primary skeletal muscle cells with recombinant full-length human adiponectin for 12 h and searched, using two-dimensional electrophoresis, for proteins upregulated more than threefold by adiponectin compared with untreated cells.RESULTS—We found one protein that was increased 6.3-fold with adiponectin incubation. MALDI-TOF (matrix-assisted laser desorption/ionization−top of flight) mass spectrometric analysis identified this protein as ferritin heavy chain (FHC). When murine primary skeletal muscle cells were treated with adiponectin, IκB-α phosphorylation was observed, suggesting that adiponectin stimulates nuclear factor (NF)-κB activity. In addition, FHC upregulation by adiponectin was inhibited by NF-κB inhibitors. These results suggest NF-κB activation to be involved in FHC upregulation by adiponectin. Other NF-κB target genes, manganese superoxide dismutase (MnSOD) and inducible nitric oxide synthase (iNOS), were also increased by adiponectin treatment. We performed a reactive oxygen species (ROS) assay using CM-H2DCFDA fluorescence and found that ROS-reducing effects of adiponectin were abrogated by FHC or MnSOD small-interfering RNA induction.CONCLUSIONS—We have demonstrated that adiponectin upregulates FHC in murine skeletal muscle tissues, suggesting that FHC elevation might partially explain how adiponectin protects against oxidative stress in skeletal muscles.

Adiponectin stimulates Wnt inhibitory factor-1 expression through epigenetic regulations involving the transcription factor specificity protein 1

Adiponectin (ADN) is an adipokine possessing growth inhibitory activities against various types of cancer cells. Our previous results demonstrated that ADN could impede Wnt/beta-catenin-signaling pathways in MDA-MB-231 human breast carcinoma cells [Wang,Y. et al. (2006) Adiponectin modulates the glycogen synthase kinase-3 beta/beta-catenin signaling pathway and attenuates mammary tumorigenesis of MDA-MB-231 cells in nude mice. Cancer Res., 66, 11462-11470]. Here, we extended our studies to elucidate the effects of ADN on regulating the expressions of Wnt inhibitory factor-1 (WIF1), a Wnt antagonist frequently silenced in human breast tumors. Our results showed that ADN time dependently stimulated WIF1 gene and protein expressions in MDA-MB-231 cells. Overexpression of WIF1 exerted similar inhibitory effects to those of ADN on cell proliferations, nuclear beta-catenin activities, cyclin D1 expressions and serum-induced phosphorylations of Akt and glycogen synthase kinase-3 beta. Blockage of WIF1 activities significantly attenuated the suppressive effects of ADN on MDA-MB-231 cell growth. Furthermore, our in vivo studies showed that both supplementation of recombinant ADN and adenovirus-mediated overexpression of this adipokine substantially enhanced WIF1 expressions in MDA-MB-231 tumors implanted in nude mice. More interestingly, we found that ADN could alleviate methylation of CpG islands located within the proximal promoter region of WIF1, possibly involving the specificity protein 1 (Sp1) transcription factor and its downstream target DNA methyltransferase 1 (DNMT1). Upon ADN treatment, the protein levels of both Sp1 and DNMT1 were significantly decreased. Using silencing RNA approaches, we confirmed that downregulation of Sp1 resulted in an increased expression of WIF1 and decreased methylation of WIF1 promoter. Taken together, these data suggest that ADN might elicit its antitumor activities at least partially through promoting WIF1 expressions.

Adiponectin prevents atherosclerosis by increasing cholesterol efflux from macrophages

Plasma high density lipoprotein (HDL)-cholesterol levels are inversely correlated to the risk of atherosclerotic cardiovascular diseases. Reverse cholesterol transport (RCT) is one of the major protective systems against atherosclerosis, in which HDL particles play a crucial role to carry cholesterol derived from peripheral tissues to the liver. Recently, ATP-binding cassette transporters (ABCA1, ABCG1) and scavenger receptor (SR-BI) have been identified as important membrane receptors to generate HDL by removing cholesterol from foam cells. Adiponectin (APN) secreted from adipocytes is one of the important molecules to inhibit the development of atherosclerosis. Epidemiological studies have revealed a positive correlation between plasma HDL-cholesterol and APN concentrations in humans, although its mechanism has not been clarified. Therefore, in the present study, we investigated the role of APN on RCT, in particular, cellular cholesterol efflux from human monocyte-derived and APN-knockout (APN-KO) mice macrophages. APN up-regulated the expression of ABCA1 in human macrophages, respectively. ApoA-1-mediated cholesterol efflux from macrophages was also increased by APN treatment. Furthermore, the mRNA expression of LXRα and PPARγ was increased by APN. In APN-KO mice, the expression of ABCA1, LXRα, PPARγ, and apoA-I-mediated cholesterol efflux was decreased compared with wild-type mice. In summary, APN might protect against atherosclerosis by increasing apoA-I-mediated cholesterol efflux from macrophages through ABCA1-dependent pathway by the activation of LXRα and PPARγ.