ABSTRACT
Background In human immunodeficiency virus 1 (HIV-1)–infected individuals, exposure to a protease inhibitor (PI)-based highly active antiretroviral therapy (HAART) regimen increases cardiovascular disease and endothelial dysfunction. However, the mechanisms of PI-induced effects on endothelial cells (ECs) are not known. Furthermore, strategies to suppress these deleterious outcomes of PIs need to be developed. Insulin-induced PI3K/Akt signaling and endothelial nitric oxide (NO)-synthase (eNOS) phosphorylation regulates NO production by ECs that maintain vascular homeostasis. We evaluated whether nelfinavir (NEL), a potent HIV-1 PI that suppresses Akt phosphorylation, can alter insulin-induced NO production in human aortic endothelial cells (HAECs) and whether insulin sensitization of HAECs via the peroxisome proliferator-activated receptor-gamma agonists, thiazolidinediones, can ameliorate these side effects.
Methods Real-time NO production in HAECs was monitored by fluorimetric dyes DAF-FM DA and DAF-2 DA. Immunodetection studies were used to determine the phosphorylation of Akt, eNOS, insulin receptor-β (IR-β), insulin receptor substrate-1 (IRS-1), and PI3K/p85α. Expression of eNOS messenger RNA was measured by reverse transcription polymerase chain reaction.
Results In vitro exposure (72 hours) of HAECs to NEL (0.25-2 μg/mL) decreased both basal (2.5-fold) and insulin-induced NO production (4- to 5-fold). NEL suppressed insulin-induced phosphorylation of both Akt and eNOS at serine residues 473 and 1177, respectively. NEL decreased tyrosine phosphorylation of IR-β, IRS-1, and PI3K. Coexposure to troglitazone (TRO; 250 nM) ameliorated the suppressive effects of NEL on insulin signaling and NO production. Coexposure to TRO also increased eNOS expression in NEL-treated HAECs.
Conclusion Our findings indicate that treatment with potent insulin sensitizers may protect against PI-mediated endothelial dysfunction during long-term HAART.
Footnotes
↵Funding: This study was supported in part by grants from the Louisiana Cancer Research Consortium (DM) and the National Institutes of Health: R01-HL63128 (KCA, DM), R01-HL073691 (KCA, DM), and R01- HL083480 (JL, DM).
1This work was carried out in collaboration with Prof. Krishna C. Agrawal (past chairman of the Department of Pharmacology at Tulane University) and his postdoctoral fellow Dr Kai Liu. In a tragic accident on December 11, 2009, Dr Agrawal passed away. May God rest his soul. Afterward, Dr Liu returned to China. The work has been recently completed by Dr Debasis Mondal and Dr Milton Hamblin and is dedicated to the memory of Dr Agrawal.
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