Inhaled Steroids and Active Smoking Drive Chronic Obstructive Pulmonary Disease Symptoms and Biomarkers to a Greater Degree Than Airflow Limitation.

RATIONALE: Chronic obstructive pulmonary disease (COPD) is a heterogeneous disease, and development of novel therapeutics requires an understanding of pathophysiologic phenotypes. OBJECTIVES: The purpose of the Airways Disease Endotyping for Personalized Therapeutics (ADEPT) study was to correlate clinical features and biomarkers with molecular characteristics in a well-profiled COPD cohort. METHODS: A total of 67 COPD subjects (forced expiratory volume in the first second of expiration [FEV1]: 45%-80% predicted) and 63 healthy smoking and nonsmoking controls underwent multiple assessments including patient questionnaires, lung function, and clinical biomarkers including fractional exhaled nitric oxide (FENO), induced sputum, and blood. MEASUREMENTS AND MAIN RESULTS: The impact of inhaled corticosteroids (ICSs), and to a lesser extent current smoking, was more associated with symptom control, exacerbation rates, and clinical biomarkers, than severity by FEV1. The ICS-treated smoking subjects were most symptomatic, with significantly elevated scores on patient-reported outcomes and more annual exacerbations (P < .05). Inhaled corticosteroid users had greater airflow obstruction and air trapping compared with non-ICS users, regardless of smoking status. Smoking, regardless of ICS use, was associated with significantly lower FENO (P < .05). Smoking, in non-ICS users, was associated with an elevated proportion of sputum neutrophils and reduced sputum macrophages. Increased serum C-reactive protein was observed in smokers but not in ICS and nonsmoking ICS users (P < .05). In contrast, only air trapping and neutrophilic inflammation increased with severity, defined by postbronchodilator FEV1. CONCLUSIONS: Compared with COPD severity by FEV1, ICS use and current smoking were better determinants of clinical characteristics and biomarkers. Use of the ADEPT COPD data promises to prove useful in defining biological phenotypes to facilitate personalized therapeutic approaches.

Lack of high BMI-related features in adipocytes and inflammatory cells in the infrapatellar fat pad (IFP).

BACKGROUND: Obesity is associated with the development and progression of osteoarthritis (OA). Although the infrapatellar fat pad (IFP) could be involved in this association, due to its intracapsular localization in the knee joint, there is currently little known about the effect of obesity on the IFP. Therefore, we investigated cellular and molecular body mass index (BMI)-related features in the IFP of OA patients. METHODS: Patients with knee OA (N = 155, 68% women, mean age 65 years, mean (SD) BMI 29.9 kg/m2 (5.7)) were recruited: IFP volume was determined by magnetic resonance imaging in 79 patients with knee OA, while IFPs and subcutaneous adipose tissue (SCAT) were obtained from 106 patients undergoing arthroplasty. Crown-like structures (CLS) were determined using immunohistochemical analysis. Adipocyte size was determined by light microscopy and histological analysis. Stromal vascular fraction (SVF) cells were characterized by flow cytometry. RESULTS: IFP volume (mean (SD) 23.6 (5.4) mm3) was associated with height, but not with BMI or other obesity-related features. Likewise, volume and size of IFP adipocytes (mean 271 pl, mean 1933 µm) was not correlated with BMI. Few CLS were observed in the IFP, with no differences between overweight/obese and lean individuals. Moreover, high BMI was not associated with higher SVF immune cell numbers in the IFP, nor with changes in their phenotype. No BMI-associated molecular differences were observed, besides an increase in TNFα expression with high BMI. Macrophages in the IFP were mostly pro-inflammatory, producing IL-6 and TNFα, but little IL-10. Interestingly, however, CD206 and CD163 were associated with an anti-inflammatory phenotype, were the most abundantly expressed surface markers on macrophages (81% and 41%, respectively) and CD163+ macrophages had a more activated and pro-inflammatory phenotype than their CD163- counterparts. CONCLUSIONS: BMI-related features usually observed in SCAT and visceral adipose tissue could not be detected in the IFP of OA patients, a fat depot implicated in OA pathogenesis.

Identification of airway mucosal type 2 inflammation by using clinical biomarkers in asthmatic patients.

BACKGROUND: The Airways Disease Endotyping for Personalized Therapeutics (ADEPT) study profiled patients with mild, moderate, and severe asthma and nonatopic healthy control subjects. OBJECTIVE: We explored this data set to define type 2 inflammation based on airway mucosal IL-13-driven gene expression and how this related to clinically accessible biomarkers. METHODS: IL-13-driven gene expression was evaluated in several human cell lines. We then defined type 2 status in 25 healthy subjects, 28 patients with mild asthma, 29 patients with moderate asthma, and 26 patients with severe asthma based on airway mucosal expression of (1) CCL26 (the most differentially expressed gene), (2) periostin, or (3) a multigene IL-13 in vitro signature (IVS). Clinically accessible biomarkers included fraction of exhaled nitric oxide (Feno) values, blood eosinophil (bEOS) counts, serum CCL26 expression, and serum CCL17 expression. RESULTS: Expression of airway mucosal CCL26, periostin, and IL-13-IVS all facilitated segregation of subjects into type 2-high and type 2-low asthmatic groups, but in the ADEPT study population CCL26 expression was optimal. All subjects with high airway mucosal CCL26 expression and moderate-to-severe asthma had Feno values (≥35 ppb) and/or high bEOS counts (≥300 cells/mm3) compared with a minority (36%) of subjects with low airway mucosal CCL26 expression. A combination of Feno values, bEOS counts, and serum CCL17 and CCL26 expression had 100% positive predictive value and 87% negative predictive value for airway mucosal CCL26-high status. Clinical variables did not differ between subjects with type 2-high and type 2-low status. Eosinophilic inflammation was associated with but not limited to airway mucosal type 2 gene expression. CONCLUSION: A panel of clinical biomarkers accurately classified type 2 status based on airway mucosal CCL26, periostin, or IL-13-IVS gene expression. Use of Feno values, bEOS counts, and serum marker levels (eg, CCL26 and CCL17) in combination might allow patient selection for novel type 2 therapeutics.

Validated and longitudinally stable asthma phenotypes based on cluster analysis of the ADEPT study.

BACKGROUND: Asthma is a disease of varying severity and differing disease mechanisms. To date, studies aimed at stratifying asthma into clinically useful phenotypes have produced a number of phenotypes that have yet to be assessed for stability and to be validated in independent cohorts. The aim of this study was to define and validate, for the first time ever, clinically driven asthma phenotypes using two independent, severe asthma cohorts: ADEPT and U-BIOPRED. METHODS: Fuzzy partition-around-medoid clustering was performed on pre-specified data from the ADEPT participants (n = 156) and independently on data from a subset of U-BIOPRED asthma participants (n = 82) for whom the same variables were available. Models for cluster classification probabilities were derived and applied to the 12-month longitudinal ADEPT data and to a larger subset of the U-BIOPRED asthma dataset (n = 397). High and low type-2 inflammation phenotypes were defined as high or low Th2 activity, indicated by endobronchial biopsies gene expression changes downstream of IL-4 or IL-13. RESULTS: Four phenotypes were identified in the ADEPT (training) cohort, with distinct clinical and biomarker profiles. Phenotype 1 was "mild, good lung function, early onset", with a low-inflammatory, predominantly Type-2, phenotype. Phenotype 2 had a "moderate, hyper-responsive, eosinophilic" phenotype, with moderate asthma control, mild airflow obstruction and predominant Type-2 inflammation. Phenotype 3 had a "mixed severity, predominantly fixed obstructive, non-eosinophilic and neutrophilic" phenotype, with moderate asthma control and low Type-2 inflammation. Phenotype 4 had a "severe uncontrolled, severe reversible obstruction, mixed granulocytic" phenotype, with moderate Type-2 inflammation. These phenotypes had good longitudinal stability in the ADEPT cohort. They were reproduced and demonstrated high classification probability in two subsets of the U-BIOPRED asthma cohort. CONCLUSIONS: Focusing on the biology of the four clinical independently-validated easy-to-assess ADEPT asthma phenotypes will help understanding the unmet need and will aid in developing tailored therapies. TRIAL REGISTRATION: NCT01274507 (ADEPT), registered October 28, 2010 and NCT01982162 (U-BIOPRED), registered October 30, 2013.

Inflammatory Cells in Patients with Endstage Knee Osteoarthritis: A Comparison between the Synovium and the Infrapatellar Fat Pad.

OBJECTIVE: To get a better understanding of inflammatory pathways active in the osteoarthritic (OA) joint, we characterized and compared inflammatory cells in the synovium and the infrapatellar fat pad (IFP) of patients with knee OA. METHODS: Infiltrating immune cells were characterized by flow cytometry in 76 patients with knee OA (mean age 63.3, 52% women, median body mass index 28.9) from whom synovial tissue (n = 40) and IFP (n = 68) samples were obtained. Pain was assessed by the visual analog scale (VAS; 0-100 mm). Spearman rank correlations and linear regression analyses adjusted for sex and age were performed. RESULTS: Macrophages and T cells, followed by mast cells, were the most predominant immune cells in the synovium and IFP, and were equally abundant in these tissues. Macrophages and T cells secreted mostly proinflammatory cytokines even without additional stimulation, indicating their activated state. Accordingly, most CD4+ T cells had a memory phenotype and contained a significant population of cells expressing activation markers (CD25+, CD69+). Interestingly, the percent of CD69+ T cells was higher in synovial than IFP CD4+ T cells. Preliminary analyses indicated that the number of synovial CD4+ T cells were associated with VAS pain (ß 0.51, 95% CI 0.09-1.02, p = 0.02). CONCLUSION: Our data suggest that the immune cell composition of the synovium and the IFP is similar, and includes activated cells that could contribute to inflammation through secretion of proinflammatory cytokines. Moreover, preliminary analyses indicate that synovial CD4+ T cells might associate with pain in patients with endstage OA of the knee.

Longitudinally Stable, Clinically Defined Clusters of Patients with Asthma Independently Identified in the ADEPT and U-BIOPRED Asthma Studies.

BACKGROUND: ADEPT (Airways Disease Endotyping for Personalized Therapeutics) and U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcome Consortium) are independent asthma biomarker studies that aim to enable personalization of therapies. METHODS: Patients in both studies were identified by similar criteria, and similar clinical parameters and biomarkers were assessed in blood, sputum, and airway samples. Fuzzy partition-around-medoid clustering was performed on the ADEPT dataset (n = 154) and independently on the U-BIOPRED asthma dataset (n = 82), filtered to match ADEPT inclusion criteria. For both studies, the same eight easily measurable clinical variables were used, and ADEPT also included methacholine airway hyperresponsiveness. Models for cluster classification probabilities were derived and applied to the 12-month longitudinal ADEPT data and the full U-BIOPRED adult asthma dataset (n = 397) as independent external validation. MEASUREMENTS AND MAIN RESULTS: Four clusters were identified in the ADEPT-asthma study population with distinct clinical and biomarker profiles. In general, Cluster 1 consists of patients with mild asthma not treated with steroids and well controlled with preserved lung function and a low-inflammatory phenotype; Cluster 2 is partially controlled, with mild airflow obstruction but severe airway hyperresponsiveness and a Th2 phenotype (brittle phenotype); Cluster 3 is partially controlled with mild airflow obstruction but reduced vital capacity, less bronchodilator reversibility, and a non-Th2 phenotype with neutrophilic inflammation (chronic obstructive pulmonary disease-like); and Cluster 4 is poorly controlled, with marked airflow obstruction, marked bronchodilator reversibility, and a mixed inflammatory phenotype. Overall, the ADEPT clusters were stable over 12 months and reproduced by identifying four analogous clusters in the U-BIOPRED asthma dataset, with distributions for most clustering and nonclustering variables similar to ADEPT. CONCLUSIONS: We report four clinical clusters in ADEPT and confirmed these by external validation in U-BIOPRED. The ADEPT clusters have distinct clinical and molecular characteristics, are stable over 12 months, and present opportunities for the development of tailored therapeutics for asthma.

Adipocytes modulate the phenotype of human macrophages through secreted lipids.

Previous studies have shown accumulation and an enhanced proinflammatory profile of macrophages in adipose tissue of obese mice, indicating the presence of an interaction between adipocytes and macrophages in this tissue. However, the consequences of this interaction in humans are yet incompletely understood. In this study, we explored the modulating effects of adipocytes on the phenotype of macrophages in humans and studied the possible molecular pathways involved. Adipocyte-conditioned media (ACM) treatment of macrophages for 48 h strongly reduced the LPS-induced IL-12p40 secretion by macrophages, whereas the production of TNF-α and other cytokines remained largely unaffected. This effect was independent of the source of adipocytes. Interestingly, the level of inhibition correlated directly with body mass index (BMI) of the adipocyte donor. Because adipocytes release many different cytokines, adipokines, and lipids, we have separated the protein and lipid fractions of ACM, to obtain insight into the molecular nature of the soluble mediators underlying the observed effect. These experiments revealed that the inhibitory effect resided predominantly in the lipid fraction. Further studies revealed that PGE2 and linoleic and oleic acid were potent inhibitors of IL-12p40 secretion. Interestingly, concentrations of these ACM-derived lipids increased with increase in BMI of the adipocyte donor, suggesting that they could mediate the BMI-dependent effects of ACM. To our knowledge, these results provide first evidence that obesity-related changes in adipose tissue macrophage phenotype could be mediated by adipocyte-derived lipids in humans. Intriguingly, these changes appear to be different from those in murine obesity.

Profiling the secretion of soluble mediators by end stage osteoarthritis synovial tissue explants reveals a reduced responsiveness to an inflammatory trigger.

OBJECTIVE: Evidence is accumulating that synovial tissue plays an active role in osteoarthritis (OA), however, exact understanding of its contribution is lacking. In order to further elucidate its role in the OA process, we aimed to identify the secretion pattern of soluble mediators by synovial tissue and to assess its ability to initiate cartilage degeneration. METHODS: Synovial tissue explants (STEs) obtained from donors without history of OA (n = 8) or from end stage OA patients (n = 16) were cultured alone or together with bovine cartilage explants in the absence or presence of IL-1α. The secretion of 48 soluble mediators was measured and the effect on glycosaminoglycan (GAG) release and matrix metalloproteinase (MMP) activity was determined. RESULTS: Normal and OA STEs secreted comparable levels of almost all measured soluble mediators. However, in the presence of IL-1α these mediators were less secreted by OA than by normal STEs of which 15 differed significantly (p<0.01). No effect of normal or OA STEs on GAG release from the cartilage explants was observed, and no differences in MMP activity between OA and normal STEs were detected. CONCLUSIONS: Unexpectedly, a comparable secretion profile of soluble mediators was found for OA and normal STEs while the reduced responsiveness of OA STEs to an inflammatory trigger indicates a different state of this tissue in OA patients. The effects could be the result of prolonged exposure to an inflammatory environment in OA development. Further understanding of the pro-inflammatory and inflammation resolving mechanisms during disease progression in synovial tissue may provide valuable targets for therapy in the future.

GLP-1 receptor activation inhibits VLDL production and reverses hepatic steatosis by decreasing hepatic lipogenesis in high-fat-fed APOE*3-Leiden mice.

OBJECTIVE: In addition to improve glucose intolerance, recent studies suggest that glucagon-like peptide-1 (GLP-1) receptor agonism also decreases triglyceride (TG) levels. The aim of this study was to evaluate the effect of GLP-1 receptor agonism on very-low-density lipoprotein (VLDL)-TG production and liver TG metabolism. EXPERIMENTAL APPROACH: The GLP-1 peptide analogues CNTO3649 and exendin-4 were continuously administered subcutaneously to high fat diet-fed APOE*3-Leiden transgenic mice. After 4 weeks, hepatic VLDL production, lipid content, and expression profiles of selected genes involved in lipid metabolism were determined. RESULTS: CNTO3649 and exendin-4 reduced fasting plasma glucose (up to -30% and -28% respectively) and insulin (-43% and -65% respectively). In addition, these agents reduced VLDL-TG production (-36% and -54% respectively) and VLDL-apoB production (-36% and -43% respectively), indicating reduced production of VLDL particles rather than reduced lipidation of apoB. Moreover, they markedly decreased hepatic content of TG (-39% and -55% respectively), cholesterol (-30% and -55% respectively), and phospholipids (-23% and -36% respectively), accompanied by down-regulation of expression of genes involved in hepatic lipogenesis (Srebp-1c, Fasn, Dgat1) and apoB synthesis (Apob). CONCLUSION: GLP-1 receptor agonism reduces VLDL production and hepatic steatosis in addition to an improvement of glycemic control. These data suggest that GLP-receptor agonists could reduce hepatic steatosis and ameliorate dyslipidemia in patients with type 2 diabetes mellitus.

Secreted frizzled-related protein 5 suppresses adipocyte mitochondrial metabolism through WNT inhibition.

Preadipocytes secrete several WNT family proteins that act through autocrine/paracrine mechanisms to inhibit adipogenesis. The activity of WNT ligands is often decreased by secreted frizzled-related proteins (SFRPs). Sfrp5 is strongly induced during adipocyte differentiation and increases in adipocytes during obesity, presumably to counteract WNT signaling. We tested the hypothesis that obesity-induced Sfrp5 expression promotes the development of new adipocytes by inhibiting endogenous suppressors of adipogenesis. As predicted, mice that lack functional SFRP5 were resistant to diet-induced obesity. However, counter to our hypothesis, we found that adipose tissue of SFRP5-deficient mice had similar numbers of adipocytes, but a reduction in large adipocytes. Transplantation of adipose tissue from SFRP5-deficient mice into leptin receptor-deficient mice indicated that the effects of SFRP5 deficiency are tissue-autonomous. Mitochondrial gene expression was increased in adipose tissue and cultured adipocytes from SFRP5-deficient mice. In adipocytes, lack of SFRP5 stimulated oxidative capacity through increased mitochondrial activity, which was mediated in part by PGC1α and mitochondrial transcription factor A. WNT3a also increased oxygen consumption and the expression of mitochondrial genes. Thus, our findings support a model of adipogenesis in which SFRP5 inhibits WNT signaling to suppress oxidative metabolism and stimulate adipocyte growth during obesity.

Loss of toll-like receptor 3 function improves glucose tolerance and reduces liver steatosis in obese mice.

OBJECTIVE: Emerging evidence suggests a link between innate immunity and development of type 2 diabetes mellitus (T2D); however, the molecular mechanisms linking them are not fully understood. Toll-like Receptor 3 (TLR3) is a pathogen pattern recognition receptor that recognizes the double-stranded RNA of microbial or mammalian origin and contributes to immune responses in the context of infections and chronic inflammation. The objective of this study was to determine whether TLR3 activity impacts insulin sensitivity and lipid metabolism. MATERIALS AND METHODS: Wild type (WT) and TLR3 knock-out (TLR3(-/-)) mice were fed a high fat diet (HFD) and submitted to glucose tolerance tests (GTTs) over a period of 33 weeks. In another study, the same group of mice was treated with a neutralizing monoclonal antibody (mAb) against mouse TLR3. RESULTS: TLR3(-/-) mice fed an HFD developed obesity, although they exhibited improved glucose tolerance and lipid profiles compared with WT obese mice. In addition, the increase in liver weight and lipid content normally observed in WT mice on an HFD was significantly ameliorated in TLR3(-/-) mice. These changes were accompanied by up-regulation of genes involved in cholesterol efflux such as PPARδ, LXRα, and LXRα-targeting genes and down-regulation of pro-inflammatory cytokine and chemokine genes in obese TLR3(-/-) mice. Furthermore, global gene expression profiling in liver demonstrated TLR3-specific changes in both lipid biosynthesis and innate immune response pathways. CONCLUSIONS: TLR3 affects glucose and lipid metabolism as well as inflammatory mediators, and findings in this study reveal a new role for TLR3 in metabolic homeostasis. This suggests antagonizing TLR3 may be a beneficial therapeutic approach for the treatment of metabolic diseases.

Cytokine production by infrapatellar fat pad can be stimulated by interleukin 1ß and inhibited by peroxisome proliferator activated receptor α agonist.

BACKGROUND: Infrapatellar fat pad (IPFP) might be involved in osteoarthritis (OA) by production of cytokines. It was hypothesised that production of cytokines is sensitive to environmental conditions. OBJECTIVES: To evaluate cytokine production by IPFP in response to interleukin (IL)1ß and investigate the ability to modulate this response with an agonist for peroxisome proliferator activated receptor α (PPARα), which is also activated by lipid-lowering drugs such as fibrates. METHODS: Cytokine secretion of IPFP was analysed in the medium of explant cultures of 29 osteoarthritic patients. IPFP (five donors) and synovium (six donors) were cultured with IL-1ß and PPARα agonist Wy14643. Gene expression of IL-1ß, monocyte chemoattractant protein (MCP1), (IL-6, tumour necrosis factor (TNF)α, leptin, vascular endothelial growth factor (VEGF), IL-10, prostaglandin-endoperoxide synthase (PTGS)2 and release of TNFα, MCP1 and prostaglandin E(2) were compared with unstimulated IPFP and synovium explants. RESULTS: IPFP released large amounts of inflammatory cytokines, adipokines and growth factors. IL-1ß increased gene expression of PTGS2, TNFα, IL-1ß, IL-6 and VEGF and increased TNFα release in IPFP. MCP1, leptin, IL-10 gene expression and MCP1, leptin and PGE(2) release did not increase significantly. Synovium responded to IL-1ß similarly to IPFP, except for VEGF gene expression. Wy14643 decreased gene expression of PTGS2, IL-1ß, TNFα, MCP1, VEGF and leptin in IPFP explants and IL-1ß, TNFα, IL-6, IL-10 and VEGF in synovium that responded to IL-1ß. CONCLUSION: IPFP is an active tissue within the joint. IPFP cytokine production is increased by IL-1ß and decreased by a PPARα agonist. The effects were similar to effects seen in synovium. Fibrates may represent a potential disease-modifying drug for OA by modulating inflammatory properties of IPFP and synovium.

Infrapatellar fat pad of patients with end-stage osteoarthritis inhibits catabolic mediators in cartilage.

OBJECTIVE: Adipose tissue is known to release inflammatory cytokines and growth factors. In this exploratory study, the authors examined whether the infrapatellar fat pad (IPFP) closely located to cartilage in the knee joint can affect cartilage metabolism. In addition, the authors analysed whether the macrophage types present in IPFP could explain the effect on cartilage. METHODS: IPFP explants obtained during total knee replacement of 29 patients with osteoarthritis (OA) were used to make fat-conditioned medium (FCM). Explants of bovine cartilage were cultured with or without FCM. Nitric oxide (NO) and glycosaminoglycan release and gene expression of matrix-degrading enzymes in cartilage were analysed. To stimulate catabolic processes in the cartilage, the authors added interleukin 1ß, and the effect of six FCMs was evaluated. The presence of different types of macrophages (CD68+, CD86+ and CD206+) in OA IPFPs was compared with subcutaneous adipose tissue samples and IPFP samples from patients with an anterior cruciate ligament rupture. RESULTS: FCM alone reduced NO and glycosaminoglycan release and matrix metalloproteinase (MMP)1 gene expression by the cartilage. Moreover, when catabolic conditions were enhanced with interleukin 1ß, FCM inhibited NO production as well as MMP1 and MMP3 gene expression and increased collagen type II gene expression. Significantly more CD206+ cells were present in OA IPFP samples than in subcutaneous fat or anterior cruciate ligament IPFP samples. CONCLUSION: In contrast to the authors' expectations, medium conditioned by end-stage OA IPFP inhibited catabolic processes in cartilage. CD206+ cells present in the IPFPs used for making the FCM might have contributed to the inhibition of catabolic processes in the cartilage.

Association between weight or body mass index and hand osteoarthritis: a systematic review.

OBJECTIVE: To investigate the association between weight or body mass index (BMI) and the development of hand osteoarthritis. METHODS: Systematic review of observational studies. Medical databases were searched up to April 2008. Articles that presented data on the association between weight and hand osteoarthritis were selected. The qualities of these studies were then assessed by two independent reviewers using a 19 criteria scoring system. Using the mean scores of all studies as a cut-off value, the studies were deemed as high or low quality. Study quality and study designs were combined to determine the level of evidence using best-evidence synthesis, which consisted of five levels of evidence. RESULTS: From the 25 studies included, two had cohort, three case-control and 20 cross-sectional study designs. Fifteen studies were considered high-quality studies. Of these high-quality studies, one cohort, two case-control and seven cross-sectional studies showed a positive association between weight or BMI and hand osteoarthritis. Based on three high-quality studies with preferred study designs (one cohort and two case-control) with a positive association, the level of evidence of the association between overweight and developing hand osteoarthritis is moderate. The approximate risk ratio of this association is 1.9. CONCLUSION: Weight or BMI is associated with the development of hand osteoarthritis. The level of evidence of published studies is moderate according to best-evidence synthesis. Further high-quality cohort or case-control studies are needed to elucidate the role of weight in hand osteoarthritis.

CNTO736, a novel glucagon-like peptide-1 receptor agonist, ameliorates insulin resistance and inhibits very low-density lipoprotein production in high-fat-fed mice.

CNTO736 is a glucagon-like peptide (GLP) 1 receptor agonist that incorporates a GLP-1 peptide analog linked to the Mimetibody platform. We evaluate the potential of acute and chronic CNTO736 treatment on insulin sensitivity and very low-density lipoprotein (VLDL) metabolism. For acute studies, diet-induced insulin-resistant C57BL/6 mice received a single intraperitoneal injection of CNTO736 or vehicle. Chronic effects were studied after 4 weeks of daily intraperitoneal administration. A hyperinsulinemic-euglycemic clamp monitored insulin sensitivity. A single dose of CNTO736 reduced fasting plasma glucose levels (CNTO736, 4.4 +/- 1.0; control, 6.3 +/- 2.4 mM) and endogenous glucose production (EGP) (CNTO736, 39 +/- 11; control, 53 +/- 13 micromol/min/kg) and increased insulin-mediated glucose uptake (CNTO736, 76 +/- 25; control, 54 +/- 13 micromol/min/kg). Chronic administration of CNTO736 reduced fasting glucose levels (CNTO736, 4.1 +/- 0.8; control 6.0 +/- 1.0 mM), improved insulin-dependent glucose uptake (CNTO736, 84 +/- 19; control, 61 +/- 15 micromol/min/kg), and enhanced inhibition of EGP (CNTO736, 91 +/- 18; control, 80 +/- 10% inhibition). In addition, chronic dosing with CNTO736 reduced fasting EGP (CNTO736, 39 +/- 9; control, 50 +/- 8 micromol/min/kg) and VLDL production (CNTO736, 157 +/- 23; control, 216 +/- 36 micromol/h/kg). These results indicate that CNTO736 reinforces insulin's action on glucose disposal and production in diet-induced insulin-resistant mice. In addition, CNTO736 reduces basal hepatic glucose and VLDL output in these animals. The data suggest that CNTO736 may be a useful tool in the treatment of type 2 diabetes.

Protein engineering strategies for sustained glucagon-like peptide-1 receptor-dependent control of glucose homeostasis.

OBJECTIVE: We have developed a novel platform for display and delivery of bioactive peptides that links the biological properties of the peptide to the pharmacokinetic properties of an antibody. Peptides engineered in the MIMETIBODY platform have improved biochemical and biophysical properties that are quite distinct from those of Fc-fusion proteins. CNTO736 is a glucagon-like peptide 1 (GLP-1) receptor agonist engineered in our MIMETIBODY platform. It retains many activities of native GLP-1 yet has a significantly enhanced pharmacokinetic profile. Our goal was to develop a long-acting GLP-1 receptor agonist with sustained efficacy. RESEARCH DESIGN AND METHODS: In vitro and in vivo activity of CNTO736 was evaluated using a variety of rodent cell lines and diabetic animal models. RESULTS: Acute pharmacodynamic studies in diabetic rodents demonstrate that CNTO736 reduces fasting and postprandial glucose, decreases gastric emptying, and inhibits food intake in a GLP-1 receptor-specific manner. Reduction of food intake following CNTO736 dosing is coincident with detection of the molecule in the circumventricular organs of the brain and activation of c-fos in regions protected by the blood-brain barrier. Diabetic rodents dosed chronically with CNTO736 have lower fasting and postprandial glucose and reduced body weight. CONCLUSIONS: Taken together, our data demonstrate that CNTO736 produces a spectrum of GLP-1 receptor-dependent actions while exhibiting significantly improved pharmacokinetics relative to the native GLP-1 peptide.

Wnt10b inhibits obesity in ob/ob and agouti mice.

The Wnt family of secreted signaling molecules has profound effects on diverse developmental processes, including the fate of mesenchymal progenitors. While activation of Wnt signaling blocks adipogenesis, inhibition of endogenous Wnt/beta-catenin signaling by Wnt10b promotes spontaneous preadipocyte differentiation. Transgenic mice with expression of Wnt10b from the FABP4 promoter (FABP4-Wnt10b) have less adipose tissue when maintained on a normal chow diet and are resistant to diet-induced obesity. Here we demonstrate that FABP4-Wnt10b mice largely avert weight gain and metabolic abnormalities associated with genetic obesity. FABP4-Wnt10b mice do not gain significant body weight on the ob/ob background, and at 8 weeks of age, they have an approximately 70% reduction in visceral and subcutaneous adipose tissues compared with ob/ob mice. Similarly, on the lethal yellow agouti (A(y)) background, FABP4-Wnt10b mice have 50-70% less adipose tissue weight and circulating leptin at 5 months of age. Wnt10b-Ay mice are more glucose tolerant and insulin sensitive than A(y) controls, perhaps due to reduced expression and circulation of resistin. Reduced expression of inflammatory cytokines may also contribute to improved glucose homeostasis.

Identification of a PTH regulated gene selectively induced in vivo during PTH-mediated bone formation.

The biological activities of parathyroid hormone (PTH) on bone are quite complex as demonstrated by its catabolic and anabolic activities on the skeleton. Although there have been many reports describing genes that are regulated by PTH in osteoblast cells, the goal of this study was to utilize a well-established in vivo PTH anabolic treatment regimen to identify genes that mediate bone anabolic effects of PTH. We identified a gene we named PTH anabolic induced gene in bone (PAIGB) that has been reported as brain and acute leukemia cytoplasmic (BAALC). Therefore, using the latter nomenclature, we have discovered that BAALC is a PTH-regulated gene whose mRNA expression was selectively induced in rat tibiae nearly 100-fold (maximal) by a PTH 1-34 anabolic treatment regimen in a time-dependent manner. Although BAALC is broadly expressed, PTH did not regulate BAALC expression in other PTH receptor expressing tissues and we find that the regulation of BAALC protein by PTH in vivo is confined to mature osteoblasts. Further in vitro studies using rat UMR-106 osteoblastic cells show that PTH 1-34 rapidly induces BAALC mRNA expression maximally by 4 h while the protein was induced by 8 h. In addition to being regulated by PTH 1-34, BAALC expression can also be induced by other bone forming factors including PGE(2) and 1,25 dihydroxy vitamin D(3). We determined that BAALC is regulated by PTH predominantly through the cAMP/PKA pathway. Finally, we demonstrate in MC3T3-E1 osteoblastic cells that BAALC overexpression regulates markers of osteoblast differentiation, including downregulating alkaline phosphatase and osteocalcin expression while inducing osteopontin expression. We also demonstrate that these transcriptional responses mediated by BAALC are similar to the responses elicited by PTH 1-34. These data, showing BAALC overexpression can mimic the effect of PTH on markers of osteoblast differentiation, along with the observations that BAALC is induced selectively with a bone anabolic treatment regimen of PTH (not a catabolic treatment regimen), suggest that BAALC may be an important mediator of the PTH anabolic action on bone cell function.