Objective: HIV lipodystrophy is characterised by abnormal adipose tissue distribution and
metabolism, as a result of altered adipocyte function and gene expression. The protease inhibitor
ritonavir is associated with the development of lipodystrophy. Quantifying changes in adipogenic gene
expression in the presence of ritonavir may help to identify therapeutic targets for HIV lipodystrophy.
Methods: Affymetrix Mouse Genome 430 2.0 oligonucleotide microarray was used to investigate gene
expression in 3T3-L1 adipocytes treated with 20 µmol/l ritonavir or vehicle control (ethanol). Pparg,
Adipoq, Retn and Il6 expression were validated by real time RT-PCR. Transcriptional signalling
through PPAR-γ was investigated using a DNA-binding ELISA. Changes in adipocyte function were
investigated through secreted adiponectin quantification using ELISA and Oil Red O staining for triglyceride storage.
Results: Expression of 389 genes was altered by more than 5-fold in the presence of ritonavir (all P < 0.001). Gene
ontology analysis revealed down-regulation of genes responsible for adipocyte triglyceride accumulation including
complement factor D (Cfd; 238.42-fold), Cidec (73.75-fold) and Pparg (5.63-fold). Glucose transport genes were also
down-regulated including Adipoq (24.42-fold) and Glut4 (13.36-fold), while Il6 was up-regulated (10.39-fold). PPAR-γ
regulatory genes Cebpa (11.33-fold) and liver-X-receptor α (Nr1h3) were down-regulated. Changes in Pparg, Adipoq and
Il6 were confirmed by RT-PCR. PPAR-γ binding to its nuclear consensus site, adiponectin secretion and triglyceride
accumulation were all reduced by ritonavir.
Conclusion: Ritonavir had a significant effect on expression of genes involved in adipocyte differentiation, lipid
accumulation and glucose metabolism. Down-regulation of Pparg may be mediated by changes in Cebpa, Lcn2 and