RESUMO
Cholesterol is required for chondrocyte differentiation and bone formation. Apolipoprotein A1 (apoA-1) plays a major role in lipoprotein clearance and cholesterol redistribution. We report here that apoA-1 is expressed during chondrocyte differentiation in vitro and in vivo. In differentiating chondrocytes, the expression of the liver X receptor (LXR) is modulated and its expression correlates to the expression of apoA-1. The expression of other LXR target genes related to cholesterol homeostasis such as ABCA1 cholesterol transporter and sterol regulatory element-binding protein 1 (SREBP1) is similarly regulated. Small molecule ligands activating either LXR or retinoid X receptor (RXR) lead to a dramatic increase in apoA-1 mRNA and protein expression in cultured chondrocytes. These ligands strongly induce ABCA1 cholesterol transporter expression and effectively mediate cholesterol efflux from hypertrophic chondrocytes. In addition, we report that, in the same cells, the ligands down modulate Serum Amyloid A expression induced by bacterial lipopolysaccharide. Our studies provide evidence that LXR/RXR mediate a fine regulation of cholesterol homeostasis in differentiating chondrocytes.
Assuntos
Apolipoproteína A-I/química , Colesterol/metabolismo , Condrócitos/metabolismo , Regulação da Expressão Gênica , Receptores X de Retinoides/fisiologia , Fatores de Transcrição/fisiologia , Animais , Western Blotting , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Cartilagem/metabolismo , Adesão Celular , Técnicas de Cultura de Células , Diferenciação Celular , Embrião de Galinha , Condrócitos/citologia , Colágeno Tipo X/metabolismo , Meios de Cultura Livres de Soro/farmacologia , Proteínas de Ligação a DNA/metabolismo , Dimerização , Eletroforese em Gel de Poliacrilamida , Homeostase , Imunoprecipitação , Ligantes , Lipopolissacarídeos/metabolismo , Lipoproteínas/química , Receptores X do Fígado , Receptores Nucleares Órfãos , Reação em Cadeia da Polimerase , RNA/metabolismo , RNA Mensageiro/metabolismo , Receptores Citoplasmáticos e Nucleares , Receptores X de Retinoides/química , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína Amiloide A Sérica/biossíntese , Proteína de Ligação a Elemento Regulador de Esterol 1 , Temperatura , Fatores de Tempo , Fatores de Transcrição/química , Fatores de Transcrição/metabolismoRESUMO
Ex-FABP is an extracellular fatty acid binding protein, expressed during chicken embryo development in cartilage, muscle fibers, and blood granulocytes. Transfection of chondrocytes and myoblasts with anti-sense Ex-FABP cDNA results in inhibition of cell proliferation and apoptosis induction. Ex-FABP expression is dramatically enhanced by inflammatory stimuli and in pathological conditions. In this paper, by in situ whole mount and immunohistochemistry analysis we show that, at early developmental stage, Ex-FABP is diffuse in all tissues of chick embryos. Particularly high level of transcript and protein are expressed in the heart. During acute phase response (APR) induced by endotoxin LPS injection, a marked increase of Ex-FABP mRNA was observed in embryos, highest Ex-FABP expression being in heart and liver. To investigate in vivo the biological role of Ex-FABP, we have directly microinjected chicken embryos with antibody against Ex-FABP. Almost 70% of chicken embryos died and the target tissue was the heart. We detected in heart of the treated embryos a significant increase of apoptotic cells and high level of fatty acids. We propose that the accumulation of fatty acid, specific ligand of Ex-FABP, in the cell microenvironment is responsible of heart cell death, and we suggest that Ex-FABP may act as a survival protein by playing a role as scavenger for fatty acids.
