CD36 mRNA and Protein Expression Levels Are Significantly Increased in the Heart and Testis of apoE Deficient Mice in Comparison to Wild Type (C57BL/6)

CD36, an 88kd-adhesion molecule, plays a major role as a scavenging receptor implicated in cellular lipid metabolism. Secretory mammary epithelium, microvasculature endothelium, adipocytes, smooth muscle cells, and platelets express CD36. In addition, CD36 expression is significantly enhanced in macrophages differentiating into foam cells. The effect of pathological levels of cholesterol, as observed in apoE(−/−), on vascular CD36 expression is, at this stage, not known. In this study, a quantitative analysis of CD36 transcription and protein expression levels, present in tissues of male C57BL/6 and apolipoprotein-E (apoE) deficient mice was carried out by Northern and Western blots. Four-week-old animals were fed a chow diet over different periods of time (0, 6, 16, or 20 weeks). Immunohistochemistry was used to localize CD36 protein expression in the heart and testis. Results indicate that CD36 transcription is increased in hearts of apoE deficient animals (100% higher at 6 weeks, and 30% higher at 16 and 20 weeks) in comparison to wild type. This was confirmed at the protein level, which showed an increase of at least 100% at 6 weeks, and between 40% to 50% increase at 16 and 20 weeks of apoE(−/−) mice compared to controls. In addition, CD36 transcription levels were significantly increased in testis of apoE animals (at least 100% at 6, 16, and 20 weeks) compared to C57BL/6 wild type. Such an increase was also confirmed at the protein level (65% increase at 16 weeks in apoE mice compared to control). Finally, localization of CD36 protein expression by immunohistochemistry showed that it was expressed in the capillaries of heart and testis endothelial cells and also at the head of spermatozoid during spermatogenesis. These results indicate that high circulating cholesterol levels, in apoE deficient mice, significantly enhance the expression of CD36 in the heart and testis. Such enhanced CD36 expression might lead to organ remodeling and/or dysfunction

CD36 Inhibitors Reduce Postprandial Hypertriglyceridemia and Protect against Diabetic Dyslipidemia and Atherosclerosis

CD36 is recognized as a lipid and fatty acid receptor and plays an important role in the metabolic syndrome and associated cardiac events. The pleiotropic activity and the multiple molecular associations of this scavenger receptor with membrane associated molecules in different cells and tissues have however questioned its potential as a therapeutic target. The present study shows that it is possible to identify low molecular weight chemicals that can block the CD36 binding and uptake functions. These inhibitors were able to reduce arterial lipid deposition, fatty acid intestinal transit, plasma concentration of triglycerides and glucose, to improve insulin sensitivity, glucose tolerance and to reduce the plasma concentration of HbAc1 in different and independent rodent models. Correlation between the anti-CD36 activity of these inhibitors and the known pathophysiological activity of this scavenger receptor in the development of atherosclerosis and diabetes were observed at pharmacological doses. Thus, CD36 might represent an attractive therapeutic target

ETUDE STRUCTURALE ET FONCTIONNELLE DE DEUX MOLECULES D'ADHESION (CD62P ET CD36) DANS L'INITIATION ET LE DEVELOPPEMENT DES LESIONS ATHEROMATEUSES (DOCTORAT)

LYON1-BU Santé (693882101) / SudocPARIS-BIUM (751062103) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

The terminal six amino-acids of the carboxy cytoplasmic tail of CD36 contain a functional domain implicated in the binding and capture of oxidized low-density lipoprotein.

CD36, a major adhesion molecule expressed by monocytes/macrophages, plays a key role in the binding and internalization of oxidized low-density lipoprotein (OxLDL). This adhesion molecule, a member of an important scavenger receptor family, contains a very short C-terminal cytoplasmic tail that is known to induce intracellular signalling events. However, the domains on the cytoplasmic tail involved in such signal transduction are unknown. In this study, we have investigated the functional components of the cytoplasmic tail by site-directed mutagenesis coupled with functional OxLDL and monoclonal antibody (mAb) binding studies. Seven truncated or punctual CD36 constructs, localized in the cytoplasmic tail, were produced by site-directed mutagenesis. Each construct was stably expressed in HEK293 cells. We used a quantitative and a qualitative method, labelling OxLDL with either iodine or rhodamine, to determine the functional importance of the cytoplasmic domains in OxLDL internalization. Results indicate that: (1) a deletion of the last amino-acid (construct K472STOP) significantly reduces, compared with wild-type, the binding, internalization and degradation of OxLDL; (2) truncation of the last six amino-acids (construct R467STOP) significantly reduces OxLDL binding; (3) the above two constructs (K472STOP and R467STOP) showed a reduced rate of OxLDL internalization compared with wild-type; (4) the binding and rate of internalization of an anti-CD36 monoclonal antibody (10/5) was not affected by the above mentioned mutants (K472STOP and R467STOP), compared with wild-type. This study shows, for the first time, a specific site on the CD36 cytoplasmic tail that is critical for the binding, endocytosis and targeting of OxLDL

Inhibition of ppTG in the plasma at four hours following the olive oil gavage following administration of 50 mg/kg of active (AP5055, AP5258) or inactive (AP5156) analogues.

<p>Inhibition of ppTG in the plasma at four hours following the olive oil gavage following administration of 50 mg/kg of active (AP5055, AP5258) or inactive (AP5156) analogues.</p

Reduction of plasma triglycerides.

<p>Comparative effect of CD36 inhibitors on the plasma concentrations of TG in different rat models, A: Dose dependent reduction in a fructose fed rat, AP5055 was administrated at different doses for 3 w (n = 12), B: AP5258 was administrated to diabetic ZDF rats (C = Control, T = Treated) for a period of 2w at 10 mg/kg (n = 8).</p

Chemical structures and activities of the CD36 inhibitors AP 5055, AP 5258 and the negative analog AP5156.

<p>Dose dependent inhibition of ox-LDL uptake and accumulation by THP1 cells at 37°C. Uptake was measured as the cyanin3 labeled ox-LDL uptake at constant cell number.</p

Anti-CD36 activity.

<p>Effect of AP5055 on the molecular interaction between CD36 and ox-LDL using CD36-HEK and wild type (wt) cells at 4°C. A: effect on the electrophoretic mobility of ox-LDL, B: Affinity crosslinking of ox-LDL to membrane expressed CD36, biotinylated ox-LDL were cross linked at 4°C, the ox-LDL complex was immunoprecipitated with an anti ox-LDL antibody and complex-associated CD36 was detected on immunoblot, using an anti CD36 antibody.</p

Anti-CD36 activity of AP5055 (dark) and AP5258 (grey) on CD36-HEK cells.

<p>Non transfected cells (wt), were used as control: A: ox-LDL uptake at 37°C, B: Palmitate uptake at 37°C, C: typical inhibition of ox-LDL binding at 4°C, D: dose dependent inhibition of AP 5055 and AP5258 on ox-LDL binding, AP5156 used as negative control had no effect (hatched bar), E: Comparative inhibition of ox-LDL binding by AP5055 and AP5258 at different ox-LDL concentrations.</p