10th International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV 6-8 November 2008, London, UK

OPPOSITE EFFECTS OF NEVIRAPINE AND EFAVIRENZ ON DIFFERENTIATION AND GENE EXPRESSION OF HUMAN ADIPOCYTES IN CULTURE

Antiviral Therapy 2008; 13(Suppl. 4):A24 (abstract no. P-04)

JM Gallego-Escuredo¹, J Diaz-Delfin¹, JC Domingo¹, MM Gutierrez², MG Mateo², P Domingo², M Giralt¹ and F Villarroya^1
1Deptatamento de Bioquimica i Biologia Molecular, Universitat de Barcelona, Barcelona, Spain; ²Hospital de la Santa Creu I Sant Pau, Barcelona, Spain

AIM: Efavirenz has been reported to have negative effects on mouse 3T3L1 adipocytes in culture and similar effects have been suggested for human adipocytes. Nevirapine has been less studied and slight positive effects have been reported for mouse brown adipocyte differentiation. Recent data indicating some extent of lipodystrophic disturbances associated with the presence of efavirenz in antiretroviral treatment regimes highlight the importance of the direct assessment of the action of these two non-nucleoside reverse transcriptase inhibitors on human adipose cells. The objective of this study was to compare the action of these two drugs on human adipocyte differentiation and gene expression.

METHODS: Human adipocytes were differentiated in primary culture from precursor cells obtained from liposuction from healthy individuals. Firstly, human pre-adipocytes were exposed to distinct concentrations of the drugs throughout the process of differentiation of pre-adipocytes into adipocytes (10 days). In a second experimental setting, already differentiated adipocytes were exposed to the drugs during 24 h. The extent of morphological adipocyte differentiation was measured by optical microscopy and by counting the cells accumulating lipid droplets. Molecular analysis was performed by RNA isolation (RNAeasy; Qiagen) and quantitative determination of transcripts using real-time PCR (TaqMan; Applied Biosystems).

RESULTS: Efavirenz caused pre-adipocyte cell death at concentrations higher than 10 µM, whereas no signs of toxicity of nevirapine were observed at this range of concentration. Lower doses of efavirenz (0.5, 2 and 4 µM) caused a dose-dependent reduction in adipocyte differentiation evidenced by the impairment in the acquisition of adipocyte morphology and the reduction in the mRNA expression of master genes of adipogenesis (PPARγ) and adipocyte phenotype (that is, adiponectin and lipoprotein lipase). No changes in mitochondrial DNA abundance or in the expression of mitochondrial DNA-encoded (COII) or nuclear DNA-encoded (COIV) transcripts for mitochondrial proteins were observed. Nevirapine treatment did not cause any impairment in adipogenic differentiation on the basis of adipocyte morphology or expression of marker genes. In fact, 20 µM nevirapine resulted in a mild positive induction of adipogenesis (upregulation of PPARγ, adiponectin and lipoprotein lipase mRNAs). Treatment of already differentiated adipocytes with efavirenz (0.5, 4 and 10 µM) also caused a reduction in the mRNAs of PPARγ and adiponectin, whereas no changes occurred for these transcripts in adipocytes treated with the same concentrations of nevirapine. Again, 20 µM nevirapine induced significantly PPARγ and adiponectin mRNA levels.

CONCLUSIONS: Efavirenz impairs morphological adipogenic differentiation and adipogenic gene expression without causing mitochondrial toxicity. In contrast, nevirapine did not impair and even induced slight adipogenesis. Considering the evidence that alterations in the adipogenic processes take place in lipoatrophic adipose tissue from patients, inclusion of non-nucleoside reverse transcriptase inhibitors in antiretroviral regimes design should take into account the anti-adipogenic properties of efavirenz with respect to nevirapine.

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2008-11-06
P-04

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