HIV-1 viral envelope glycoprotein gp120 triggers an inflammatory response in cultured rat astrocytes and regulates the functional expression of P-glycoprotein.
In this work, we examined the ability of gp120, a human immunodeficiency virus-1 (HIV-1) viral envelope glycoprotein, to trigger the innate immune response in astrocytes, an HIV-1 brain cellular target, and we investigated the functional expression of the ATP-binding cassette membrane transporter P-glycoprotein (P-gp) in primary cultures of rat astrocytes treated with gp120 or cytokines [tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6]. Standard 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium and d-mannitol uptake assays confirmed that HIV-1(96ZM651) gp120 treatment did not alter cell viability or membrane permeability. Semiquantitative reverse-transcriptase polymerase chain reaction analysis and enzyme-linked immunosorbent assay demonstrated increased TNF-alpha, IL-1beta, and IL-6 mRNA and protein expression in cultures treated with HIV-1(96ZM651) gp120, suggesting in vitro activation of immune responses. Cytokine secretion was detected when CXCR4 but not CCR5 was inhibited with a specific antibody, implying that cytokine secretion is primarily mediated via CCR5 in astrocytes triggered with HIV-1(96ZM651) gp120. P-gp protein expression was increased in astrocyte cultures exposed to TNF-alpha (2.9-fold) or IL-1beta (1.6-fold) but was decreased profoundly in the presence of IL-6 (8.9-fold), suggesting that IL-6 is primarily involved in modulating P-gp expression. In parallel, after HIV-1(96ZM651) gp120 treatment, immunoblotting analysis showed a significant decrease in P-gp expression (4.7-fold). Furthermore, the accumulation of two P-gp substrates, digoxin and saquinavir (an HIV-1 protease inhibitor), was enhanced (1.5- to 1.8-fold) in HIV-1(96ZM651) gp120-treated astrocyte monolayers but was not altered by P-gp inhibitors [e.g., valspodar (PSC833) and elacridar (GF120918)], suggesting a loss of transport activity. Taken together, these data imply that HIV-1(96ZM651) gp120 or cytokine treatment modulate P-gp functional expression in astrocytes, which may lead to complex drug-transporter interactions during HIV-1 encephalitis-associated immune responses.
Ronaldson
Patrick T
PT
Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, 144 College Street, Toronto, ON, Canada, M5S 3M2.
Bendayan
Reina
R
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D000818:Animals; D001253:Astrocytes / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D004285:Dogs; D005786:Gene Expression Regulation / Q000187:drug effects*; D015699:HIV Envelope Protein gp120 / Q000276:immunology* / Q000494:pharmacology*; D006801:Humans; D007249:Inflammation / Q000276:immunology*; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D019713:Receptors, CCR5 / Q000235:genetics; D019718:Receptors, CXCR4 / Q000235:genetics
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D016207:Cytokines; D015699:HIV Envelope Protein gp120; D012333:RNA, Messenger; D019713:Receptors, CCR5; D019718:Receptors, CXCR4