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ABSTRACT: The adipocyte-derived cytokine leptin acts as a metabolic switch, connecting the body's metabolism to high-energy consuming processes such as reproduction and immune responses. Accumulating evidence suggests that leptin plays a role in human pathologies, such as autoimmune diseases and cancer, thus providing a rationale for the development of leptin antagonists. In the present study, we generated and evaluated a panel of neutralizing nanobodies targeting the LR (leptin receptor). A nanobody comprises the variable domain of the naturally occurring single-chain antibodies found in members of the Camelidae family. We identified three classes of neutralizing nanobodies targeting different LR subdomains: i.e. the CRH2 (cytokine receptor homology 2), Ig-like and FNIII (fibronectin type III) domains. Only nanobodies directed against the CRH2 domain inhibited leptin binding. We could show that a nanobody that targets the Ig-like domain potently interfered with leptin-dependent regulation of hypothalamic NPY (neuropeptide Y) expression. As a consequence, daily intraperitoneal injection increased body weight, body fat content, food intake, liver size and serum insulin levels. All of these characteristics resemble the phenotype of leptin and LR-deficient animals. The results of the present study support proposed models of the activated LR complex, and demonstrate that it is possible to block LR signalling without affecting ligand binding. These nanobodies form new tools to study the mechanisms of BBB (blood-brain barrier) leptin transport and the effect of LR inhibition in disease models.
Biochemical Journal 08/2011; 441(1):425-34. · 4.90 Impact Factor
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Sandrine Aspeslagh,
Yali Li,
Esther Dawen Yu,
Nora Pauwels,
Matthias Trappeniers,
Enrico Girardi, Tine Decruy,
Katrien Van Beneden,
Koen Venken,
Michael Drennan,
Luc Leybaert,
Jing Wang,
Richard W Franck,
Serge Van Calenbergh,
Dirk M Zajonc,
Dirk Elewaut
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ABSTRACT: Invariant natural killer T (iNKT) cells are known to have marked immunomodulatory capacity due to their ability to produce copious amounts of effector cytokines. Here, we report the structure and function of a novel class of aromatic α-galactosylceramide structurally related glycolipids with marked Th1 bias in both mice and men, leading to superior tumour protection in vivo. The strength of the Th1 response correlates well with enhanced lipid binding to CD1d as a result of an induced fit mechanism that binds the aromatic substitution as a third anchor, in addition to the two lipid chains. This induced fit is in contrast to another Th1-biasing glycolipid, α-C-GalCer, whose CD1d binding follows a conventional key-lock principle. These findings highlight the previously unexploited flexibility of CD1d in accommodating galactose-modified glycolipids and broaden the range of glycolipids that can stimulate iNKT cells. We speculate that glycolipids can be designed that induce a similar fit, thereby leading to superior and more sustained iNKT cell responses in vivo.
The EMBO Journal 06/2011; 30(11):2294-305. · 9.20 Impact Factor
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Sylvie Seeuws,
Peggy Jacques,
Jens Van Praet,
Michael Drennan,
Julie Coudenys, Tine Decruy,
Ellen Deschepper,
Lien Lepescheux,
Philippe Pujuguet,
Line Oste,
Nick Vandeghinste,
Reginald Brys,
Gust Verbruggen,
Dirk Elewaut
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ABSTRACT: Disease severity in collagen-induced arthritis (CIA) is commonly assessed by clinical scoring of paw swelling and histological examination of joints. Although this is an accurate approach, it is also labour-intensive and the application of less invasive and less time-consuming methods is of great interest. However, it is still unclear which of these methods represents the most discriminating measure of disease activity.
We undertook a comparative analysis in which different measurements of inflammation and tissue damage in CIA were studied on an individual mouse level. We compared the current gold standard methods - clinical scoring and histological examination - with alternative methods based on scoring of X-ray or micro-computed tomography (CT) images and investigated the significance of systemically expressed proteins, involved in CIA pathogenesis, that have potential as biomarkers.
Linear regression analysis revealed a marked association of serum matrix metalloproteinase (MMP)-3 levels with all features of CIA including inflammation, cartilage destruction and bone erosions. This association was improved by combined detection of MMP-3 and anti-collagen IgG2a antibody concentrations. In addition, combined analysis of both X-ray and micro-CT images was found to be predictive for cartilage and bone damage. Most remarkably, validation analysis using an independent data set proved that variations in disease severity, induced by different therapies, could be accurately represented by predicted values based on the proposed parameters.
Our analyses revealed that clinical scoring, combined with serum MMP-3, anti-collagen IgG2a measurement and scoring of X-ray and micro-CT images, yields a comprehensive insight into the different aspects of disease activity in CIA.
Arthritis research & therapy 01/2010; 12(4):R160. · 4.27 Impact Factor
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Leo Leung,
Cyrille Tomassi,
Katrien Van Beneden, Tine Decruy,
Matthias Trappeniers,
Dirk Elewaut,
Yifang Gao,
Tim Elliott,
Aymen Al-Shamkhani,
Christian Ottensmeier,
Jörn M Werner,
Anthony Williams,
Serge Van Calenbergh,
Bruno Linclau
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ABSTRACT: The synthesis of 2',2'-difluoro KRN7000 is described. In vivo evaluation demonstrates that this fluorinated glycolipid induces CD1d-dependent TCR activation of NKT cells, with a bias towards Th2 cytokine production.
ChemMedChem 02/2009; 4(3):329-34. · 3.15 Impact Factor
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Journal of the American Chemical Society 01/2009; 130(49):16468-9. · 9.91 Impact Factor
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Leo Leung,
Cyrille Tomassi,
Katrien Van Beneden, Tine Decruy,
Dirk Elewaut,
Tim Elliott,
Aymen Al-Shamkhani,
Christian Ottensmeier,
Serge Van Calenbergh,
Joern Werner,
Tony Williams,
Bruno Linclau
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ABSTRACT: The synthesis of 4-deoxy-4,4-difluoro-KRN7000 starting from phytosphingosine is described. Key steps include a regioselective benzylation of azidophytosphingosine and a deoxofluor-mediated fluorination of the corresponding 4-ketone. This fluorination failed completely when the adjacent 3-OH was protected as benzyl ether but proceeded well when a benzoyl group was used. The biological evaluation reveals a bias toward Th1 cytokine induction upon Natural Killer T cell activation.
Organic Letters 10/2008; 10(20):4433-6. · 5.86 Impact Factor
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Matthias Trappeniers,
Stijn Goormans,
Katrien Van Beneden, Tine Decruy,
Bruno Linclau,
Aymen Al-Shamkhani,
Tim Elliott,
Christian Ottensmeier,
Joern M Werner,
Dirk Elewaut,
Serge Van Calenbergh
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ABSTRACT: alpha-GalCer (also known as KRN7000) is an immunomodulatory glycolipid that is known to potently activate invariant natural killer T (NKT) cells upon CD1d-mediated stimulation. Because Th1 and Th2 cytokines, which are released after alpha-GalCer presentation, antagonize each other's effects, alpha-GalCer analogues that induce a biased Th1/Th2 response are highly awaited. In this context, we report the synthesis and in vitro evaluation of alpha-Gal-D-xylo-Cer and two alpha-Gal-L-lyxo-Cer analogues, one with the natural acyl chain, the other with a truncated chain.
ChemMedChem 08/2008; 3(7):1061-70. · 3.15 Impact Factor
