Endocrine Abstracts (2006) 11 P884

Gene expression profile in orbital fibroblasts from a tao patient before and after adipocytic differentiation

P Agretti, G de Marco, E Ferrarini, A Dimida, D Sansone, M Banco, M de Servi, S Lisi, C Marcocci, G de Marinò, P Vitti, A Pinchera & M Tonacchera

Dipartimento di Endocrinologia, Centro Eccellenza AmbiSEN, Pisa, Italy.

Thyroid associated ophthalmopathy (TAO) is a chronic autoimmune disorder characterized by an increased volume of the adipose/connective tissue in the human orbit. Fibroblasts from the connective orbital tissue are able to differentiate into mature adipocytes if grown in particular culture conditions. Aim of the present study was to determine the gene expression profile of orbital fibroblasts after adipocytic differentiation. Fibroblasts in primary culture were obtained from orbital connective tissue from a TAO patient. For adipocytic differentiation, fibroblasts were grown in the presence of biotin, panthotenic acid, transferrin, T3, insulin, cPGI2, dexamethasone and IBMX. Total RNA was extracted from orbital fibroblasts and double strand cDNA and biotinylated cRNA were synthesized. cRNA was hybridized on U133 set arrays following the Affymetrix protocol. The acquired images were analyzed to obtain the expression levels of different transcripts. The GeneMAPP software was used to insert each gene in the contest of metabolic pathways. In differentiated fibroblasts 409 modulated genes were identified, the majority being down-regulated (97%) with respect to control fibroblasts. Among the down-regulated genes numerous factors involved in the inflammatory process and many chemotactic cytokines like macrophage inflammatory protein 2-alpha, interleukin-8, small inducible cytokine B6, macrophage inflammatory protein 2-beta were identified. Among the genes which were up-regulated, genes codifing for nuclear protein involved in the control of DNA replication (DNA replication licensing factor MCM4) and in RNA synthesis (RNA polymerase II elongation factor ELL3) were identified. The gene that mainly increased its expression in differentiated fibroblasts was 60S acidic ribosomal protein P2 codifing for a protein involved in protein synthesis. In conclusion, 97% of genes from fibroblasts subjected to adipocytic differentiation were down-regulated and they were represented mainly by genes involved in the inflammatory process. An increased expression of genes implicated in DNA and RNA synthesis was observed.