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Adipocytes in obesity have inappropriately low cholesterol while adiponectin release is reduced. Cholesterol shortage may contribute to low adiponectin and 3T3-L1 cells treated with lovastatin have diminished adiponectin in cell supernatants. LDL and HDL deliver cholesterol to adipocytes. LDL but not HDL increases adiponectin in cell supernatants of primary human adipocytes. The effect of LDL is not blocked by receptor associated protein suggesting that members of the LDL-receptor family are not involved. To evaluate whether these in vitro observations translate into changes in systemic adiponectin, adiponectin was measured in serum of three patients before, immediately after and 3d after LDL-apheresis. Whereas circulating lipoproteins are reduced immediately after apheresis adiponectin is not changed. Therefore, acute lowering of lipoproteins does not affect systemic adiponectin also excluding that plenty of adiponectin is bound to lipoprotein particles. Accordingly, levels of adiponectin in purified lipoproteins are quite low. Familial hypobetalipoproteinemia (FHBL) is a rare disorder associated with low plasma LDL. Serum adiponectin is, however, similar compared to healthy controls. Thus, neither LDL nor HDL directly contributes to circulating adiponectin concentrations.

Lipid storage and adipokine secretion are critical features of adipocytes. Annexin A6 (AnxA6) is a lipid-binding protein regulating secretory pathways and its role in adiponectin release was examined. The siRNA-mediated AnxA6 knock-down in 3T3-L1 preadipocytes impaired proliferation, and differentiation of AnxA6-depleted cells to mature adipocytes was associated with higher soluble adiponectin and increased triglyceride storage. The latter was partly attributed to reduced lipolysis. Accordingly, AnxA6 overexpression in 3T3-L1 adipocytes lowered cellular triglycerides and adiponectin secretion. Indeed, serum adiponectin was increased in AnxA6 deficient mice. Expression analysis identified AnxA6 protein to be more abundant in intra-abdominal compared to subcutaneous adipose tissues of mice and men. AnxA6 protein levels increased in white adipose tissues of obese mice and here, levels were highest in subcutaneous fat. AnxA6 protein in adipocytes was upregulated by oxidative stress which might trigger AnxA6 induction in adipose tissues and contribute to impaired fat storage and adiponectin release.

The adipokine adiponectin is well known to affect the function of immune cells and upregulation of CCL2 by adiponectin in monocytes/macrophages has already been reported. In the current study the effect of adiponectin on CCL2, -3, -4, and -5 and their corresponding receptors CCR1, CCR2, and CCR5 has been analyzed. Adiponectin elevates mRNA and protein of the CC chemokines in primary human monocytes. Simultaneously the surface abundance of CCR2 and CCR5 is reduced while CCR1 is not affected. Downregulation of CCR2 by adiponectin is blocked by a CCR2 antagonist although expression of the CCL2 regulated genes CCR2 and TGF-beta 1 is not altered in the adiponectin-incubated monocytes. CCL2, -3, and -5 concentrations measured in supernatants of monocytes of normal-weight (NW), overweight (OW), and type 2 diabetic (T2D) patients positively correlate with BMI and are increased in obesity and T2D. In contrast CCL4 is similarly abundant in the supernatants of all of these monocytes. The degree of adiponectin-mediated induction of the chemokines CCL3, -4, and -5 negatively correlates with their basal levels and upregulation of CCL3 and CCL5 is significantly impaired in OW and T2D cells. Serum concentrations of these chemokines are almost equal in the three groups and do not correlate with the levels in monocyte supernatants. In conclusion these data demonstrate that adiponectin stimulates release of CCL2 to CCL5 in primary human monocytes, and induction in cells of overweight probands is partly impaired. Adiponectin also lowers surface abundance of CCR2 and CCR5 and downregulation of CCR2 seems to depend on autocrine/paracrine effects of CCL2.

Connective tissue growth factor (CTGF) is induced in liver fibrosis and enhances the activity of transforming growth factor β (TGFβ). Recently we have shown that the hepatoprotective adipokine adiponectin downregulates CTGF in primary human hepatocytes (PHH). In the current study, the mechanisms mediating suppression of CTGF by adiponectin and the well described downstream effector of adiponectin receptor 2 (AdipoR2), peroxisome proliferator activated receptor α (PPARα), were analyzed in more detail. Adiponectin downregulated CTGF mRNA and protein in primary human hepatocytes (PHH) and suppression was blocked by a PPARα antagonist indicating that AdipoR2 is involved. The PPARα agonists fenofibrate and WY14643 also reduced CTGF protein in these cells. Adiponectin further impaired TGFβ-mediated upregulation of CTGF. Phosphorylation of the TGFβ downstream effectors SMAD2 and –3 was reduced in PHH incubated with adiponectin or PPARα agonists suggesting that early steps in TGFβ signal transduction are impaired. CTGF and TGFβ mRNA levels were increased in human non-fibrotic non-alcoholic steatohepatitis (NASH), and here AdipoR2 expression was significantly reduced. Current data show that CTGF and TGFβ are already induced in non-fibrotic NASH and this may be partly explained by low adiponectin bioactivity which interferes with TGFβ signaling by reducing phosphorylation of SMAD2/3 and by downregulating CTGF.

Fatty liver is commonly detected in obesity and has been identified as a risk factor for the progression of hepatic fibrosis in a wide range of liver diseases. Transforming growth factor beta (TGFβ) and activin A, both members of the TGFβ superfamiliy, are central regulators in liver fibrosis and regeneration, and the effect of hepatocyte lipid accumulation on the release of these proteins was studied. Primary human hepatocytes (PHH) were incubated with palmitic acid or oleic acid to increase lipid storage. Whereas activin A and its natural inhibitor follistatin were not affected, TGFβ was 2-fold increased. The hepatoprotective adipokine adiponectin dose-dependently induced activin A while lowering follistatin but did not alter TGFβ. Activin A was markedly reduced in hepatocyte cell lines compared to PHH and was not induced upon adiponectin incubation demonstrating significant differences of primary and transformed cells. In free fatty acid (FFA)-incubated PHH adiponectin-mediated induction of activin A was impaired. Inhibition of TGFβ receptors ALK4/5 and blockage of SMAD3 phosphorylation rescued activin A synthesis in FFA and in TGFβ incubated cells suggesting that FFA inhibit adiponectin activity by inducing TGFβ. To evaluate whether serum levels of activin A and its antagonist are altered in patients with hepatic steatosis, both proteins were measured in the serum of patients with sonographically diagnosed fatty liver and age- and BMI-matched controls. Systemic adiponectin was significantly reduced in patients with fatty liver but activin A and follistatin were not altered. In summary the current data demonstrate that lipid accumulation in hepatocytes induces TGFβ which impairs adiponectin bioactivity, and thereby may contribute to liver injury.

Transforming growth factor (TGF) β is the central cytokine in fibrotic liver diseases. We analyzed whether hepatoprotective adiponectin directly interferes with TGFβ1 signaling in primary human hepatocytes (PHH). Adiponectin induces the TGFβ decoy receptor BMP-and activin-membrane-bound inhibitor (BAMBI) in PHH. Overexpression of BAMBI in hepatoma cells impairs TGFβ-mediated phosphorylation of SMAD2 and induction of connective tissue growth factor. BAMBI is lower in human fatty liver with a higher susceptibility to liver fibrosis and negatively correlates with BMI of the donors. Hepatic BAMBI is reduced in rodent models of liver inflammation and fibrosis. In summary, the current data show that hepatoprotective effects of adiponectin include induction of BAMBI which is reduced in human fatty liver and rodent models of metabolic liver injury.

Type 2 diabetes (T2D) is characterized by increased oxidative stress contributing to the development of cardiovascular disease (CVD). Monocytes are critically important in the pathogenesis of CVD and antioxidant enzymes like superoxide dismutase (SOD2) protect these cells from excessive reactive oxygen species (ROS). Adiponectin is an adipocyte-derived protein with atheroprotective function and the effect of adiponectin on monocyte SOD2 was analyzed herein. Adiponectin upregulated SOD2 mRNA and dose- and time-dependently induced SOD2 protein in primary human monocytes. Elevated systemic free fatty acids (FFA) are commonly found in T2D patients and palmitic acid as well as oleic acid reduced monocyte SOD2 protein. Adiponectin mediated upregulation of SOD2, however, was not affected by FFA incubation. SOD2 protein was reduced in T2D monocytes compared to monocytes of age- and body mass index-matched healthy controls. Adiponectin still induced SOD2 in T2D monocytes but efficiency tended to be reduced. In summary this study indicates that elevated systemic free fatty acids and impaired adiponectin activity contribute to reduced SOD2 and most likely increased oxidative stress in T2D monocytes.

Galectin-3 plays a role in atherosclerotic diseases, and the effect of adiponectin that protects from atherosclerotic diseases on monocytic galectin-3 was analysed. Adiponectin reduced galectin-3 mRNA, its cellular and soluble form, and this effect was impaired in T2D cells. Cellular galectin-3 was higher in monocytes of overweight than normal-weight donors and was highest in T2D cells. Cellular galectin-3 positively correlated with the BMI of the donors and negatively with soluble monocyte galectin-3. Circulating levels of total adiponectin did not correlate with cellular or soluble galectin-3 indicating that additional factors contribute to higher cellular monocytic galectin-3 in obesity and T2D.

Adiponectin circulates as trimer (LMW), hexamer (MMW) and high molecular weight multimer (HMW) but the distribution and effects of these isoforms have not been studied in detail. Monocytes were isolated from normal weight and overweight controls and patients with type 2 diabetes mellitus (T2D) and monocytic release of IL-6 positively correlated with the body mass index (BMI). HMW-adiponectin further enhanced and LMW-adiponectin reduced IL-6 release in monocytes. Systemic total adiponectin, and the HMW isoform were not different in these groups but MMW-adiponectin was lower in T2D, and LMW-adiponectin was reduced in the obese and T2D. Circulating LMW-adiponectin negatively correlated to monocytic IL-6 release. Systemic IL-6 was higher in the obese control group and T2D, respectively, but did not correlate with monocytic IL-6 secretion. Therefore, the current study indicates that HMW-adiponectin exerts pro- and LMW-adiponectin antiinflammatory effects and reduced LMW-adiponectin in obesity may partly contribute to elevated monocytic IL-6 release.

Low circulating levels of high molecular weight adiponectin (HMW-Apm) have been linked to dyslipidaemia and systemic HMW-Apm negatively correlates with very low density lipoprotein (VLDL), apolipoprotein B (ApoB), and ApoE and is positively associated with ApoA-I. Therefore, it was investigated whether HMW-Apm alters the hepatic synthesis of ApoB, ApoE, and ApoA-I or the activity of the hepatic ATP-binding cassette transporter A1 (ABCA1), as the main determinant of plasma HDL. HMW-Apm reduces hepatic ApoB and ApoE release whereas ABCA1 protein, activity and ApoA-I were not altered. Global gene expression analysis revealed that hepatic nuclear factor 4-alpha (HNF4-alpha) and HNF4-alpha regulated genes like ApoB are downregulated by HMW-Apm and this was confirmed at the mRNA and protein level. Therefore it is concluded that HMW-adiponectin may ameliorate dyslipidaemia by reducing the hepatic release of ApoB and ApoE, whereas ABCA1 function and ApoA-I secretion are not influenced.

The adiponectin receptors AdipoR1 and AdipoR2 have been identified to mediate the insulin-sensitizing effects of adiponectin. Although AdipoR2 was suggested to be the main receptor for this adipokine in hepatocytes, AdipoR1 protein is highly abundant in primary human hepatocytes and hepatocytic cell lines. Nuclear receptors are main regulators of lipid metabolism and activation of peroxisome proliferator-activated receptor α and γ, retinoid X receptor (RXR), and liver X receptor (LXR) by specific ligands may influence AdipoR1 abundance. AdipoR1 protein is neither altered by RXR or LXR agonists nor by pioglitazone. In contrast, fenofibric acid reduces AdipoR1 whereas hepatotoxic troglitazone upregulates AdipoR1 protein in HepG2 cells. Taken together this work shows for the first time that AdipoR1 protein is expressed in human hepatocytes but that it is not a direct target gene of nuclear receptors. Elevated AdipoR1 induced by hepatotoxic troglitazone may indicate a role of this receptor in adiponectin-mediated beneficial effects in liver damage.

Non-alcoholic fatty liver disease (NAFLD) comprising hepatic steatosis, non-alcoholic steatohepatitis (NASH), and progressive liver fibrosis is considered the most common liver disease in western countries. Fatty liver is more prevalent in overweight than normal-weight people and liver fat positively correlates with hepatic insulin resistance. Hepatic steatosis is regarded as a benign stage of NAFLD but may progress to NASH in a subgroup of patients. Besides liver biopsy no diagnostic tools to identify patients with NASH are available, and no effective treatment has been established. Visceral obesity is a main risk factor for NAFLD and inappropriate storage of triglycerides in adipocytes and higher concentrations of free fatty acids may add to increased hepatic lipid storage, insulin resistance, and progressive liver damage. Most of the adipose tissue-derived proteins are elevated in obesity and may contribute to systemic inflammation and liver damage. Adiponectin is highly abundant in human serum but its levels are reduced in obesity and are even lower in patients with hepatic steatosis or NASH. Adiponectin antagonizes excess lipid storage in the liver and protects from inflammation and fibrosis. This review aims to give a short survey on NAFLD and the hepatoprotective effects of adiponectin.

Adiponectin whose systemic levels are reduced in obesity-related diseases ameliorates insulin sensitivity and regulates biological processes like apoptosis, proliferation, migration and inflammation. Adiponectin binds to adiponectin receptors, AdipoR1 and AdipoR2, which are ubiquitously expressed. Clathrin-dependent endocytosis of AdipoR1 and adiponectin has been demonstrated to modulate adiponectin bioactivity. Recently, APPL1 has been identified as an AdipoR1 and AdipoR2 binding protein. Furthermore, activated protein kinase C1, endoplasmic reticulum protein 46 and protein kinase CK2β subunit form a complex with AdipoR1. This review summarizes recent studies exploiting heterologous expression of adiponectin receptors in yeast, and the type and function of the recently described adiponectin receptor associated proteins.

Adiponectin is believed to exert hepatoprotective effects and induces CXCL8, a chemokine that functions as a survival factor, in vascular cells. In the current study, it is demonstrated that adiponectin also induces CXCL8 expression in primary human hepatocytes but not in hepatocellular carcinoma cell lines. Knock down of the adiponectin receptor (AdipoR) 1 or AdipoR2 by small-interfering RNA indicates that AdipoR1 is involved in adiponectin-stimulated CXCL8 release. Adiponectin activates nuclear factor (NF)-κB in primary hepatocytes and pharmacological inhibition of NF-κB, the p38 mitogen-activated protein kinase, and extracellular signal-regulated kinase (ERK) 1/ERK2 reduces adiponectin-mediated CXCL8 secretion. Furthermore, adiponectin also activates STAT3 involved in interleukin (IL)-6 and leptin-mediated CXCL8 induction in primary hepatocytes. Inhibition of JAK2 by AG-490 does not abolish adiponectin-stimulated CXCL8, indicating that this kinase is not involved. Pretreatment of primary cells with “STAT3 Inhibitor VI,” however, elevates hepatocytic CXCL8 secretion, demonstrating that STAT3 is a negative regulator of CXCL8 in these cells. In accordance with this assumption, IL-6, a well-characterized activator of STAT3, reduces hepatocytic CXCL8. Therefore, adiponectin-stimulated induction of CXCL8 seems to be tightly controlled in primary human hepatocytes, whereas neither NF-κB, STAT3, nor CXCL8 are influenced in hepatocytic cell lines. CXCL8 is a survival factor, and its upregulation by adiponectin may contribute to the hepatoprotective effects of this adipokine.

It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D).Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments.Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 microg/ml) significantly (p0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 microg/ml) significantly (p=0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p=0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients.CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.

Background: Adiponectin is a circulating protein with hepatoprotective effects. Aims: To study the relationship of excessive alcohol consumption and serum adiponectin levels (SAL). Patients and Methods: The SAL were determined in (i) heavy drinkers without advanced liver damage during the course of alcohol withdrawal, (ii) patients with chronic hepatitis C virus (HCV) infection, (iii) patients with alcohol-associated cirrhosis, and (iv) healthy volunteers that consumed excessive amounts of alcohol for only a short period of time. Further, primary human hepatocytes (PHH) and adipocytes were incubated in vitro with alcohol or serum of patients. Results: Patients with chronic alcohol consumption had significantly higher SAL than HCV-patients with comparable degrees of liver damage. In alcoholics, but not in HCV patients, SAL positively correlated with serum levels of aminotransferases. Further, SAL correlated with the amount of alcohol consumption but declined during the course of alcohol abstinence. After short-term excessive alcohol consumption SAL were not elevated in healthy individuals. Adiponectin mRNA was detectable in adipocytes but not in hepatocytes, and alcohol failed to induce adiponectin in both cell types. In contrast, serum of active drinkers induced adiponectin expression in adipocytes while serum from the same individuals collected after alcohol withdrawal had no effect. Conclusions: Alcohol exhibits a specific effect on SAL that is dose and time dependent, and correlates with the degree of hepatic damage. Alcohol does not seem to affect adiponectin expression directly in adipocytes but potentially via mediators systemically released as a result of the chronic alcohol intake.

Adiponectin is an adipocyte-derived protein with atheroprotective and immunoregulatory function. Adiponectin and activin A reduce foam cell formation and adiponectin activates the p38 MAPK pathway that is well described to induce activin A. Therefore, it was analyzed whether adiponectin alters activin A in primary human monocytes. Adiponectin dose- and time-dependently induced activin A in the supernatant, and the maximal amount was observed after 12 h of incubation. Adiponectin-stimulated release of activin A was blocked by a p38 MAPK inhibitor. Metformin and pioglitazone are drugs frequently used to treat diabetic patients and metformin slightly reduced monocytic activin A release whereas pioglitazone had no effect. Type 2 diabetes is associated with elevated inflammatory systemic cytokines but activin A serum levels were similar in slim probands, overweight controls and type 2 diabetic patients. Furthermore, activin A did not correlate to systemic adiponectin, body mass index, waist to hip ratio or C-reactive protein. These findings indicate that adiponectin upregulates monocytic activin A release via the p38 MAPK pathway, and this may in part explain the immunoregulatory and antiatherosclerotic effects of this adipokine.

Background: Cartonectin (collagenous repeat-containing sequence of 26-kDa protein; CORS-26) was described as a new adipokine of the C1q/TNF molecular superfamily C1q/TNF-related protein-3 (CTRP-3), secreted by the adipocytes of mice and humans. The receptor and function of cartonectin are unknown and the recombinant protein is not commercially available. Objective: To investigate the effects of recombinant cartonectin on the secretion of adipokines such as adiponectin, leptin, and resistin from adipocytes of human and murine origin. The effect of the BMI of the adipocyte donor was also investigated. Methods and Procedures: Human adipocytes from pooled lean and preobese healthy individuals and murine 3T3-L1 adipocytes were used for stimulation experiments. Recombinant cartonectin was expressed in insect H5 cells. Adipokine secretion was measured using enzyme-linked immunosorbent assay. In addition, western blot analysis and luciferase reporter gene assays were employed. Results: Cartonectin (1, 10, 50, and 250 ng/ml) in higher doses stimulates the secretion of adiponectin and resistin from murine adipocytes. This effect is not caused by an induction of peroxisome proliferator-activated receptor-γ (PPAR-γ) protein expression, as confirmed by western blot analysis. Also, luciferase reporter gene assay revealed that cartonectin failed to induce luciferase activity at the peroxisome proliferator-activated receptor responsive element site containing the adiponectin/luciferase promoter fragment. Human adipocytes from lean individuals secrete higher amounts of adiponectin and leptin when compared with adipocytes of individuals with a preobesity BMI (25–30 kg/m2). Cartonectin failed to stimulate adiponectin or leptin secretion from human adipocytes, irrespective of the BMI value. Discussion: Cartonectin is a new adipokine that differentially regulates the secretion of classical adipokines, with marked differences between the human and the murine systems. These effects are species-dependent, while basal adipokine secretion is influenced by the BMI.

Adiponectin, an adipocyte-derived serum protein, is known to positively affect the glucose and lipid metabolism and these effects are mediated by its receptors, AdipoR1 and R2. Serum adiponectin levels are inversely associated with breast cancer risk, but the molecular mechanisms underlying this association are not fully elucidated. Thus, the purpose of this study was to investigate the influence of adiponectin on breast cancer cells in vitro. We were able to demonstrate the expression of AdipoR1 and R2 in MCF-7, MDA-MB-231 and SK-BR-3 breast cancer cells on the mRNA level. Furthermore, the AdipoR1 protein could be detected by immunoblot analysis. In MCF-7 breast cancer cells, the expression of AdipoR1 significantly declined after stimulation with 17-beta estradiol, whereas the cyclin A2 expression significantly increased. Both effects were inhibited by the addition of adiponectin. Treatment with different concentrations of adiponectin in steroid-hormone-free medium did not affect cell proliferation or apoptosis. In contrast, after the addition of 17-beta estradiol, adiponectin slightly decreased the growth of the MDA-MB-231 and SK-BR3 cells but increased proliferation of the hormone-dependent MCF-7 breast cancer cells. Adiponectin also triggered cellular apoptosis in MDA-MB-231 breast cancer cells in the presence of 17-beta estradiol. These findings suggest that a cross-talk between adiponectin and estrogen receptor signaling exists in breast cancer cells and that adiponectin effects on the growth and apoptosis of breast cancer cells in vitro are dependent on the presence of 17-beta estradiol.

Adiponectin, an adipocyte-derived serum protein, is known to positively affect the glucose and lipid metabolism and these effects are mediated by its receptors, AdipoR1 and R2. Serum adiponectin levels are inversely associated with breast cancer risk, but the molecular mechanisms underlying this association are not fully elucidated. Thus, the purpose of this study was to investigate the influence of adiponectin on breast cancer cells in vitro. We were able to demonstrate the expression of AdipoR1 and R2 in MCF-7, MDA-MB-231 and SK-BR-3 breast cancer cells on the mRNA level. Furthermore, the AdipoR1 protein could be detected by immunoblot analysis. In MCF-7 breast cancer cells, the expression of AdipoR1 significantly declined after stimulation with 17-beta estradiol, whereas the cyclin A2 expression significantly increased. Both effects were inhibited by the addition of adiponectin. Treatment with different concentrations of adiponectin in steroid-hormone-free medium did not affect cell proliferation or apoptosis. In contrast, after the addition of 17-beta estradiol, adiponectin slightly decreased the growth of the MDA-MB-231 and SK-BR3 cells but increased proliferation of the hormone-dependent MCF-7 breast cancer cells. Adiponectin also triggered cellular apoptosis in MDA-MB-231 breast cancer cells in the presence of 17-beta estradiol. These findings suggest that a cross-talk between adiponectin and estrogen receptor signaling exists in breast cancer cells and that adiponectin effects on the growth and apoptosis of breast cancer cells in vitro are dependent on the presence of 17-beta estradiol.

Adiponectin circulates in the body in high concentrations, and 100-fold lower amounts were described in the cerebrospinal fluid (CSF) of mice, whereas in humans, contradictory results have been published. To clarify whether adiponectin is present in human CSF and is derived from the circulation, it was determined in human CSF and plasma of 52 nonselected patients. Adiponectin was detected by immunoblot in CSF and was quantified in CSF and serum by ELISA. CSF adiponectin was positively correlated to systemic levels, and the CSF/serum adiponectin ratio was correlated to the CSF/serum albumin ratio. Furthermore, disturbed function of the blood-brain barrier (BBB) was associated with an elevated CSF/serum adiponectin ratio. Adiponectin mRNA was not found in the brain, indicating that adiponectin crosses the BBB and/or the blood-cerebrospinal fluid barrier (BCB). Rat adiponectin with a COOH-terminal tag was injected into the tail vein of rats and was detected 3 h later in CSF. However, CSF adiponectin in humans and rats was ∼0.1% of the serum concentration and therefore was below the 0.5% expected in the CSF because of the residual leakage of an undisturbed BBB/BCB. Taken together, data from the present study show that adiponectin in human CSF is far below the level expected by the baseline BBB/BCB permeability, indicating that adiponectin enters the brain much less efficiently than albumin, thus supporting recent data that exclude adiponectin transport to the CSF. Additional studies are needed to reveal whether these low levels of adiponectin in CSF have a physiological function.

AIM: To determine circulating and hepatic adiponectin in rodents with fatty liver disease or liver cirrhosis and investigate expression of the adiponectin receptors AdipoR1 on the mRNA and protein level and AdipoR2 on the mRNA level. METHODS: Fat fed rats were used as a model for fatty liver disease and bile duct ligation in mice to investigate cirrhotic liver. Expression of AdipoR1 and AdipoR2 mRNA was determined by real time RT-PCR. AdipoR1 protein was analysed by immunoblot. Adiponectin was measured by ELISA. RESULTS: Systemic adiponectin is reduced in fat-fed rats but is elevated in mice after bile duct ligation (BDL). Hepatic adiponectin protein is lower in steatotic liver but not in the liver of BDL-mice when compared to controls. Adiponectin mRNA was not detected in human liver samples or primary human hepatocytes nor in rat liver but recombinant adiponectin is taken up by isolated hepatocytes in-vitro. AdipoR1 mRNA and AdipoR1 protein levels are similar in the liver tissue of control and fat fed animals whereas AdipoR2 mRNA is induced. AdipoR2 mRNA and AdipoR1 mRNA and protein is suppressed in the liver of BDL-mice. CONCLUSION: Our studies show reduced circulating adiponectin in a rat model of fatty liver disease whereas circulating adiponectin is elevated in a mouse model of cirrhosis and similar findings have been described in humans. Diminished hepatic expression of adiponectin receptors was only found in liver cirrhosis.

Background Adipose tissue is recognized as a compartment secreting highly active molecules. Creeping fat represents a characteristic feature of Crohn's disease (CD). Proinflammatory or anti-inflammatory adipose-derived secretory products, now generally called adipocytokines, may play a role in the pathogenesis of CD. Materials and methods Adipose tissue specimens were obtained from creeping fat contiguous to the involved intestine of 10 patients with CD. Mesenteric adipose tissue specimens resected from 13 patients with colon cancer (CC) and from 7 patients with diverticulitis served as controls. Three fat tissue specimen per well and 6 to 8 wells per patient were incubated ex vivo for 24 h. The release of adiponectin, resistin, leptin, interleukin-6, macrophage colony-stimulating factor, monocyte chemotactic protein-1, and migration inhibitory factor was measured by ELISA. The expression of AdipoR1 and AdipoR2 receptors was investigated by real-time quantitative polymerase chain reaction in a subset of adipose tissues. Results The secretion of adiponectin and macrophage colony-stimulating factor, as well as leptin and migration inhibitory factor, was significantly upregulated in CD compared with CC and diverticulitis or CC only, respectively. Resistin, interleukin-6, and monocyte chemotactic protein-1 were not specifically induced in CD but were associated with unspecific inflammation. Adiponectin receptor expression was not different in CC, CD, or diverticulitis. Steroid treatment in CD patients had differential effects on the ex vivo secretion of adipocytokines. Conclusions A specific secretion pattern of proinflammatory and anti-inflammatory adipocytokines indicates the significance of adipose tissue in intestinal inflammation in CD. Future investigations of this intestinal compartment may provide novel insights into the pathophysiological role of creeping fat and CD.

Adiponectin (APM) is an adipocyte-derived adipokine with immunosuppressive, antidiabetic, and antiatherosclerotic properties. Low molecular weight (LMW)- and higher molecular weight (HMW)-APM circulate in the serum and activate different signaling pathways. We were interested to see whether LMW-APM exerts different effects on monocytic cells compared with the HMW isoform. Therefore, the effects of recombinant LMW-APM produced in insect cells and the APM from higher eukaryotic cells containing HMW forms on monocytic cells were investigated with respect to apoptosis and inflammation. LMW- and HMW-APM induce apoptosis in nondifferentiated THP-1 cells, reduce macrophage scavenger receptor (MSR) A mRNA expression, and stimulate phosphorylation of adenosine monophosphate-activated protein kinase (AMPK). However, HMW-APM induces the secretion of interleukin (IL)-6 in human monocytes and THP-1 cells but does not suppress lipopolysaccharide (LPS)-induced IL-6 secretion. In contrast, LMW-APM reduces LPS-mediated IL-6 release and furthermore, stimulates IL-10 secretion, most likely by reducing the abundance of inhibitor of nuclear factor (NF)-κB kinase β, leading to a diminished nuclear translocation of NF-κB p65. Our data indicate that the different APM isoforms do share common effects on monocytic cells but also induce isoform-specific responses. Although apoptosis, the activation of AMPK, and the reduction of MSR are mediated by all APM isoforms, only LMW-APM displays anti-inflammatory properties.

Proteins secreted by adipocytes (adipokines) play an important role in the pathophysiology of type 2 diabetes mellitus and the associated chronic and low-grade state of inflammation. It was the aim to characterize the antiinflammatory potential of the new adipocytokine, C1q/TNF-related protein-3 (CTRP-3), which shows structural homologies to the pleiotropic adipocytokine adiponectin. mRNA and protein expression of CTRP-3 was analyzed by RT-PCR and Western blot. Recombinant CTRP-3 and small interfering RNA-based strategies were used to investigate the effect of CTRP-3 on toll-like receptor (TLR) ligand, lipopolysaccharide (LPS)-, and lauric acid-induced chemokine release of monocytes and adipocytes. Together with complex ELISA-based techniques, a designed TLR4/myeloid differentiation protein-2 fusion molecule shown to bind LPS was used to prove the ability of CTRP-3 to act as endogenous LPS antagonist. CTRP-3 is synthesized in monocytes and adipocytes. The recombinant protein dose-dependently inhibits the release of chemokines in monocytes and adipocytes that were induced by lauric acid, LPS, and other TLR ligands in vitro and ex vivo. CTRP-3 inhibits monocyte chemoattractant protein-1 release in adipocytes, whereas small interfering RNA-mediated knockdown of CTRP-3 up-regulates monocyte chemoattractant protein-1 release, reduces lipid droplet size, and decreases intracellular triglyceride concentration in adipocytes, causing a dedifferentiation into a more proinflammatory and immature phenotype. By using a designed TLR4/MD-2 fusion molecule, it is shown by different techniques that CTRP-3 specifically and effectively inhibits the binding of LPS to its receptor, TLR4/MD-2. CTRP-3 inhibits three basic and common proinflammatory pathways involved in obesity and type 2 diabetes mellitus (adipo-inflammation) by acting as an endogenous LPS antagonist of the adipose tissue.

Background Systemic adiponectin is reduced in patients with cardiovascular disease (CVD) and low adiponectin may contribute to the pathogenesis of atherosclerosis. However, circulating adiponectin is elevated in type 1 diabetes (T1D) patients, who have also a higher incidence to develop CVD. Because monocytes play an important role in atherosclerosis, we analysed the influence of adiponectin on cytokine and chemokine release in monocytes from T1D patients and controls. Methods Systemic adiponectin was determined in the plasma and the high-molecular weight (HMW) form of adiponectin was analysed by immunoblot. Monocytes were isolated from T1D patients and controls and the adiponectin-stimulated release of interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1, CCL2) and interleukin-8 (IL-8, CXCL8) was analysed. Results Systemic adiponectin was higher in T1D patients. Immunoblot analysis of the plasma indicate abundance of HMW adiponectin in T1D patients and controls. IL-6, CCL2 and CXCL8 secretion in response to adiponectin were found induced in monocytes from controls whereas only IL-6 was upregulated in T1D cells. The induction of IL-6 by adiponectin was abrogated by an inhibitor of the NFκB pathway. Conclusion These data indicate that adiponectin-mediated induction of IL-6, CCL2 and CXCL8 is disturbed in monocytes from T1D patients and therefore elevated systemic adiponectin in T1D patients may be less protective when compared to controls.

The adiponectin paralog CORS-26 (collagenous repeat-containing sequence of 26kDa protein) is a member of the C1q/TNF-alpha molecular superfamily. CORS-26 is a secreted protein and baculovirus-produced CORS-26 released in the supernatant of insect cells forms stable trimers. Adiponectin exerts anti-inflammatory effects in LPS-treated monocytic cells and CORS-26 also reduces IL-6 and TNF-alpha secretion but does not increase IL-10. Suppression of NFkappaB signalling may explain the anti-inflammatory actions of CORS-26. Furthermore CORS-26 protein was detected in human monocytic and dendritic cells. The present data demonstrate for the first time that CORS-26 forms trimers, exerts anti-inflammatory properties and that it is expressed in monocytic cells. Therefore CORS-26 may provide a new target for pharmacological drugs in inflammatory diseases like the metabolic syndrome.

Aldehyde oxidase 1 (AOX1) is highly abundant in the liver and oxidizes aldehydes thereby generating reactive oxygen species. Enzymes involved in detoxification of aldehydes are expressed in adipocytes and alter adipogenesis, therefore the functional role of AOX1 in adipocytes was analyzed. AOX1 mRNA was higher in visceral compared to subcutaneous human adipose tissue but AOX1 protein was detected in both fat depots. AOX1 expression in adipocytes was confirmed by immunohistochemistry and immunoblot. AOX1 was induced during adipocytic differentiation and was downregulated by fenofibrate in differentiated cells. Knock-down of AOX1 in preadipocytes led to impaired lipid storage and adiponectin release in the differentiated cells. These data indicate that AOX1 is essential for adipogenesis and may link energy and drug metabolism.