Uncoupling protein 1 gene expression implicates brown adipocytes in highly active antiretroviral therapy-associated lipomatosis

Highly active antiretroviral therapy (HAART) in AIDS patients often results in partial lipodystrophy. A subset of patients under HAART also develop lipomatosis, often characterized by enlarged dorsocervical fat pads referred to as 'buffalo hump' [¹]. A recent study estimated a 6% prevalence of 'buffalo hump' lipomas among HAART-treated patients [²]. The pathophysiological mechanisms underlying lipodystrophy and lipomatosis are poorly understood. The anatomical distribution of HAART-associated lipomas is reminiscent of that in multiple symmetric lipomatosis. This unusual syndrome has diverse origins including mitochondrial dysfunction [³]. The mitochondrial toxicity associated with HAART and the similar anatomical distribution has led to the hypothesis that HAART-associated 'buffalo hump' syndrome and multiple symmetric lipomatosis could have a common aetiopathology [⁴]. Lipomas in patients with mitochondrial DNA mutations contain a mixture of univacuolated and multivacuolated fat cells that resemble brown fat rather than white fat [⁵]. However, the phenotype of the adipose cells developing in HAART-associated lipomas is unknown.

Brown adipose tissue is specialized in thermogenesis and is formed by multivacuolated adipocytes. It is almost absent in adults, although some brown adipocytes are interspread within the white fat depots [⁶]. The unequivocal cell identity of brown adipocytes can be monitored only by determining uncoupling protein 1 (UCP-1) gene expression, a unique molecular marker that distinguishes brown from white adipocytes [⁷]. Understanding the cellular basis of lipomatosis may contribute to the knowledge of the aetiopathology of the overall adipose alterations associated with HAART in AIDS patients. We therefore analysed UCP-1 gene expression in HAART-associated lipomas to identify the potential implication of brown adipocytes in HAART-associated lipomatosis.

We studied three HIV-positive patients (L1-L3) undergoing liposuction for the removal of the dorsocervical fat-pad and a fourth HIV-positive patient undertaking the surgical removal of a lipoma in the forearm (L4). They had been on HAART composed of three antiretroviral drugs: stavudine plus two other transcriptase inhibitors (L2-L4), or plus a transcriptase inhibitor and a protease inhibitor (L1). The controls (C1-C5) were HIV-positive patients on HAART with a pattern of drug composition and duration similar to the patients showing lipomatosis. They had clinical characteristics consistent with HAART-associated lipodystrophy (fat wasting from the face, buttocks, limbs and central adiposity), but they did not show lipomatosis. The controls were similar to patients with lipomas with respect to their mean age (41.0 versus 41.8 years, P = 0.86), serum CD4 cells/μl (697 versus 438, P = 0.35), serum cholesterol (5.9 versus 4.8 + 0.8 mmol/l, P = 0.86) and triglyceride levels (1.7 versus 1.8 mmol/l, P = 0.90). Controls and patients had achieved virological control of HIV infection (1.43 versus 1.77 log₁₀ for HIV RNA, P = 0.38). Biopsy samples of subcutaneous adipose tissue from controls were taken from the abdominal area. A sample of perirenal fat from an unrelated patient already used as a positive control for UCP-1 gene expression [⁸] was analysed in parallel. UCP-1 messenger RNA was determined by a reverse transcriptase-polymerase chain reaction plus Southern blot assay specifically designed for highly sensitive analysis of UCP-1 gene expression in humans, whereas cyclophilin-A mRNA was assayed as an internal control using reverse transcriptase-polymerase chain reaction, as already reported [⁸].

The results indicated that the lipomas showed a clear signal for UCP-1 mRNA, somewhat weaker than that in the positive control (Fig. 1). No signal or faint signals were detected in the non-lipoma subcutaneous adipose tissue of lipodystrophic patients. These results demonstrate that HAART-associated lipomas express the UCP-1 gene, and therefore cells of the brown adipocyte lineage are implicated in HAART-associated lipomatosis.

Fig. 1

Residual brown adipocytes or preadipocytes have already been implicated in lipomatosis of unknown aetiology and in patients bearing mtDNA mutations [^4,8]. The fact that lipomas with aetiopathology as diverse as mtDNA mutations or HAART share UCP-1 gene expression reinforces the hypothesis that mitochondrial disturbances, of either genetic or pharmacological origin, cause a dysregulation of adipocyte biology leading to the appearance of highly proliferative brown adipocyte-like cells and lipomas. Brown adipocytes could be preferential targets of HAART-induced mitochondrial toxicity as these cells have a high mitochondrial content, in contrast with white adipocytes. Impaired mitochondrial function may thus elicit a compensatory response based on brown adipocyte proliferation. Brown adipocytes have a high capacity to proliferate and differentiate under the effects of pathophysiological stimuli such as, for instance, phaeochromocytoma [⁷]. Moreover, a dual 'white and brown' fat morphology appears in the subcutaneous adipose tissue from lipoatrophic areas of HAART-treated patients [⁹]. A comparison of UCP-1 mRNA levels in the adipose depots of lipodystrophic HAART patients will be required to establish whether a dysregulation of brown versus white adipocyte biology is also implicated in the overall development of lipodystrophy, apart from the specific involvement in lipomatosis reported here.