Development and Characterization of a Whole-Cell Radioligand Binding Assay for [<sup>125</sup>I] gp120 OF HIV-1

Tô, Long-Thành; Balasubramanian, V.; Charlton, Maura E.; Francis, Terry; Doyle, Carolyn A.; Sweetnam, Paul M.

doi:10.1080/15321819208019825

Cited by 8 publications

(5 citation statements)

References 32 publications

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“…The data derived from control baseline experiments were consistent with the published literature (38,44). The K d for radioligand binding of HIV gp120 to CD4 ϩ lymphocytes was reported to be 6 nM (39,43). Accordingly, we used 1 g/ml of HIV gp120 as we previously reported in neonatal myocytes (23), which corresponds to 8 nM.…”

Section: Resultssupporting

confidence: 74%

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

Kan

Xie²,

Finkel³

2004

American Journal of Physiology-Cell Physiology

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Kan, Hong, Zirong Xie, and Mitchell S. Finkel. p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes. Am J Physiol Cell Physiol 286: C1-C7, 2004; 10.1152/ ajpcell.00059.2003.-Myocardial dysfunction leading to dilated cardiomyopathy has been documented with surprisingly high frequency in human immunodeficiency virus (HIV)-infected individuals. p38 MAP kinase has been implicated as a mediator of myocardial dysfunction. We previously reported p38 MAP kinase activation by the HIV coat protein gp120 in neonatal rat cardiac myocytes. We now report the direct inotropic effects of HIV gp120 on adult rat ventricular myocytes (ARVM). ARVM were continuously superfused with gp120, and percent fractional shortening (FS) was determined by automated border detection and simultaneous intracellular ionized free Ca 2ϩ concentration ([Ca 2ϩ ]i) measured by fura 2-AM fluorescence: gp120 alone increased FS and increased [Ca 2ϩ ]i within 5 min and then depressed FS without a decrease in [Ca 2ϩ ]i by 20-60 min, which persisted for at least 2 h. Exposure of ARVM to gp120 also resulted in the phosphorylation of the upstream regulator of p38 MAP kinase MKK3/6, p38 MAP kinase itself, and its downstream effector, ATF-2, over a similar time course. ERK (p44/42) and JNK stress signaling pathways were not similarly activated. The effects of the p38 MAP kinase inhibitor were concentration dependent. SB-203580 (10 M) blocked both p38 MAP kinase phosphorylation and the delayed negative inotropic effect of gp120. SB-203580 (5 M) selectively blocked phosphorylation of ATF-2 without blocking the phosphorylation of MKK3/6 or p38 MAP kinase itself. SB-203580 (5 M) administered before, with, or after gp120 blocked the negative inotropic effect of gp120 in ARVM. p38 MAP kinase activation may be a common stress-response mechanism contributing to myocardial dysfunction in HIV and other nonischemic as well as ischemic cardiomyopathies.

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Section: Resultssupporting

confidence: 74%

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

Kan

Xie²,

Finkel³

2004

American Journal of Physiology-Cell Physiology

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“…Immunoprecipitation was performed as described by To et al (1992). Briefly, 50 p1 aliquots of rgpl20 (100 pg/ml) were incubated overnight at 4°C with 10 pl of anti-gpl20 serum or preimmune serum (cat serum 288 or 315, respectively, AIDS Research and Reference Reagent Program).…”

Section: Cell Culture Neurotoxicity and [Ca2+] Fluxmentioning

confidence: 99%

The Envelope Glycoprotein of HIV‐1 Alters NMDA Receptor Function

et al. 1993

Eur J of Neuroscience

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Human immunodeficiency virus (HIV-1) infection often results in central nervous system (CNS) dysfunction, yet the mechanism(s) of action for HIV-1 in the CNS are not fully understood. In the present study gp120, the HIV-1 envelope glycoprotein, was shown to selectively inhibit N-methyl-D-aspartate (NMDA) receptor function. In addition to inhibiting radioligand binding to rat NMDA receptors, gp120 inhibited NMDA-induced currents in Xenopus oocytes, attenuated NMDA-stimulated calcium flux and cytotoxicity in cultured cerebellar granule cells, and provided partial protection against NMDA-induced lethality in vivo. These findings suggest that NMDA receptor complex is a possible site of action of HIV-1 within the CNS.

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“…This in vitro concentration was initially selected based on the published affinity (K d ) for HIV gp120 radioligand binding to CD4? lymphocytes of 6 nM [24]. The application of a highly sensitive catheter-based in vivo hemodynamic technique in awake rats raised the possibility of observing subtle changes at such low doses.…”

Section: Discussionmentioning

confidence: 99%

P38 MAP Kinase Inhibitor Prevents Diastolic Dysfunction in Rats Following HIV gp120 Injection In vivo

2009

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HIV infection in patients is associated with a surprisingly high frequency of diastolic dysfunction followed by the development of a dilated cardiomyopathy. Potential mechanisms include direct effects of HIV proteins, including gp120. We have previously reported direct inotropic and p38 MAP kinase signaling effects of HIV gp120 on isolated cardiac myocytes in vitro. We now report effects of a single injection of HIV gp120 on cardiac hemodynamics in vivo. HIV gp120 (50 microg/kg) was injected intravenously and hemodynamics assessed at 1, 24, 48 and 72 h in freely ambulatory, awake rats. Rats injected with gp120 demonstrated a blunted diastolic response to increasing intravenous (IV) injections of the beta-adrenergic agonist, isoproterenol (ISO), compared with similarly instrumented controls at 48 h (gp120 vs. control, P < 0.01; n = 6, for each). Pre-treatment with the p38 MAP kinase inhibitor, SB 203580, prevented the diastolic dysfunction (gp120 + SB vs. control, P = NS; n = 6, for each). Hemodynamic changes correlated with inhibition of both p38 MAP kinase and troponin I phosphorylation by SB in vivo. There were no differences in ISO dose-response curves between gp120-treated and control rats at 1, 24 or 72 h (P = NS; n = 6, for each). Thus, evidence of diastolic dysfunction is apparent in vivo in rats following a single dose of HIV gp120. To our knowledge, this is the first report of a hemodynamic effect of an HIV protein in vivo. These findings lend further support to the hypothesis that the HIV virus, itself, may contribute to myocardial dysfunction in patients infected with HIV via a p38 MAP kinase mechanism.

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Development and Characterization of a Whole-Cell Radioligand Binding Assay for [¹²⁵I] gp120 OF HIV-1

Cited by 8 publications

References 32 publications

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

The Envelope Glycoprotein of HIV‐1 Alters NMDA Receptor Function

P38 MAP Kinase Inhibitor Prevents Diastolic Dysfunction in Rats Following HIV gp120 Injection In vivo

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Development and Characterization of a Whole-Cell Radioligand Binding Assay for [125I] gp120 OF HIV-1

Cited by 8 publications

References 32 publications

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes

The Envelope Glycoprotein of HIV‐1 Alters NMDA Receptor Function

P38 MAP Kinase Inhibitor Prevents Diastolic Dysfunction in Rats Following HIV gp120 Injection In vivo

Contact Info

Product

Resources

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Development and Characterization of a Whole-Cell Radioligand Binding Assay for [¹²⁵I] gp120 OF HIV-1