3rd International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV 23-26 October 2001, Athens, Greece

SYNERGISTIC ANTI-ADIPOGENIC EFFECTS OF HIV-1 PROTEASE INHIBITORS AND TNF-α IN 3T3-LL CELLS: ROLE OF ECM DEGRADING PRO TEASES

Antiviral Therapy 2001; 6(Suppl. 4):18 (abstract no. 25)

KC Agrawal, VF LaRussa and D Mondal
Tulane Health Sciences Center, New Orleans, La., USA

BACKGROUND: Long term use of HIV-1 protease inhibitors (PI) in HIV patients is associated with lipodystrophy manifested by peripheral wasting and central adipocity.

OBJECTIVE: To delineate the molecular mechanisms associated with lipodystrophy syndrome, we have investigated the effects PIs and TNF-α on adipocyte differentiation in murine 3T3-L1 cells and on modulation of extracellular matrix (ECM) proteins.

METHODS: Adipogenic differentiation in 3T3-Ll cells was monitored by Oil red-O staining. RT-PCR studies were conducted to assess gene expression of pref-1, PAI-1 and PAI-2. Zymgraphic analysis was conducted to monitor activities of matrix metalloproteases (MMPs). Chromogenic assay was utilized to assess plasmin and PAl activity.

RESULTS: A concentration dependent decrease in both lipid (4-59%) and triglyceride (11-49%) levels was observed after 10 days inadipogenic 3T3-Ll cells to indinavir (2-50 μ/ml) or ritonavir (0.4-10 μ/ml). Simultaneous treatment with TNF-(showed a synergistic suppression in lipid levels by 45-95% at 10 U/ml and almost a complete suppression at 100 U/ml. The effect of PIs on insulin induced lipogenesis as monitored by ¹⁴C-glucose incorporation into lipids was suppressed by 21-86% in a concentration dependent manner. Insulin sensitizing agent, troglitazone (80 and 400 nM) effectively blocked the PI mediated adipogenic suppression. Pre-adipocyte factor-1 (pref-1) mRNA levels were down-regulated (four- sixfold) within 48 h post insulin stimulation, however, a smaller decrease (1.2-1.8-fold) was observed in PI exposed cells. During adipogenesis, the decrease in proteolytic activity of MMP-2 and MMP-9 was reversed upon exposure to the PIs. Similarly, the plasminolytic activity was increased and plasminogen activator inhibitor (PAl) activity was decreased in PI treated cells. The insulin mediated induction (three to fourfold) of PAI-1 and PAI-2 message was suppressed upon exposure to PIs, which was reversed by troglitazone treatment.

CONCLUSIONS: These results indicate that the HIV-1 PIs may suppress adipogenesis by disrupting the concerted actions of host proteases that regulate ECM integrity required for initiation of differentiation.

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