Role of potassium in human immunodeficiency virus production and cytopathic effects.

Acute infection of CD4+ lymphoid cells by human immunodeficiency virus type 1 (HIV-1) induces an increase in the intracellular concentration of potassium (K+). Media containing reduced or elevated concentrations of K+ were used to investigate the role of this ion in HIV-1 production and cytopathology. Incubation of CD4+ lymphoblastoid cells acutely infected by HIV-1 (strain LAI) in low K+ medium resulted in an approximately 50% decrease in HIV-1 production and markedly diminished HIV-1 induced cytopathic effects (CPE) relative to cells incubated in medium containing a normal K+ concentration (approximately 5 mM). Incubation of HIV-1 infected cells in media containing elevated concentrations of K+ medium. Cells mM) increased HIV-1 production by two- to fivefold over the amount produced in cells incubated in normal K+ medium. Cells incubated in high K+ media also displayed enhanced HIV-1-induced cytopathology. The decrease in HIV-1 production by low K+ medium and increase by high K+ media could be a accounted for by effects on HIV-1 reverse transcription. However, low K+ medium inhibited HIV-1 protein synthesis and high K+ media increased HIV-1 protein synthesis. These results suggest that the HIV-1-induced increase in intracellular is required for efficient viral replication and to induce cytopathology.

Inhibition of HIV type 1 production by hygromycin B.

HIV infection alters the cellular uptake of ions and other small molecules. This study was designed to determine whether hygromycin B, a low molecular weight (MW 527) aminoglycoside protein synthesis inhibitor that is normally impermeable to mammalian cells at micromolar concentrations, can selectively inhibit HIV expression and cytopathology. CD4+ T lymphoblastoid cells (H9) and peripheral blood mononuclear cells (PBMCs) were infected with HIV-1, then incubated in medium containing various concentrations of hygromycin B. HIV-1-induced formation of multinucleated giant cells and single cell killing were dramatically reduced in the presence of micromolar concentrations of hygromycin B. Hygromycin B also inhibited HIV-1 production in a dose-dependent manner during acute infection. G418, a larger and more hydrophobic aminoglycoside (MW 692), did not display the same selective inhibition of HIV-1 production as hygromycin B. Relative to mock-infected cells, protein synthesis in acutely infected H9 cells was selectively inhibited by hygromycin B. Hygromycin B also reduced HIV production in PBMCs and in H9 cells persistently infected with HIV. PCR analysis demonstrated that hygromycin B did not inhibit HIV-1 reverse transcription. These results demonstrate that HIV-1 infection renders cells more sensitive to hygromycin B than uninfected cells, and provides support for the hypothesis that HIV-1 induces an alteration of plasma membrane permeability. The HIV-modified cell membrane may be a potential target for antiviral intervention and chemotherapy.

A synthetic peptide corresponding to the carboxy terminus of human immunodeficiency virus type 1 transmembrane glycoprotein induces alterations in the ionic permeability of Xenopus laevis oocytes.

The carboxy-terminal 29 amino acids of the human immunodeficiency virus type 1 transmembrane glycoprotein (HIV-1 TM) are referred to as lentivirus lytic peptide 1 (LLP-1). Synthetic peptides corresponding to LLP-1 have been shown to induce cytolysis and to alter the permeability of cultured cells to various small molecules. To address the mechanisms by which LLP-1 induces cytolysis and membrane permeability changes, various concentrations of LLP-1 were incubated with Xenopus laevis oocytes, and two-electrode, voltage-clamp recording measurements were performed. LLP-1 at concentrations of 75 nM and above induced dramatic alterations in the resting membrane potential and ionic permeability of Xenopus oocytes. These concentrations of LLP-1 appeared to induce a major disruption of plasma membrane electrophysiological integrity. In contrast, concentrations of LLP-1 of 20-50 nM induced changes in membrane ionic permeability that mimic changes induced by compounds, such as the bee venom peptide melittin, that are known to form channel-like structures in biological membranes at sublytic concentrations. An analog of LLP-1 with greatly reduced cytolytic activity failed to alter the electrophysiological properties of Xenopus oocytes. Thus, by altering plasma membrane ionic permeability, the carboxy terminus of TM may contribute to cytolysis of HIV-1-infected CD4+ cells.

Use of antipolymer antibody assay in recipients of silicone breast implants.

BACKGROUND: Local complications (encapsulation, rashes, rupture, and leakage) can occur after placement of silicone gel-containing breast implants (SBI). Whether SBI exposure results in systemic manifestations in some recipients is controversial. We have carried out a blinded study to assess whether there is any difference between SBI recipients and non-exposed controls in the proportions positive for serum antibodies directed against polymeric substances. METHODS: We recruited female SBI recipients (including those without symptoms) who presented to a single rheumatology clinic. A physician global assessment was used to classify SBI recipients who did not meet criteria for specific autoimmune diseases according to the severity of local and systemic signs and symptoms. Controls were recruited from among clinic staff and their acquaintances. Results of the antipolymer antibody (APA) assay were compared with those of an assay for antinuclear antibodies (ANA) and with the severity of the signs and symptoms. FINDINGS: Positive APA results were found in one (3%) of 34 SBI recipients with limited symptoms, two (8%) of 26 with mild symptoms, seven (44%) of 16 with moderate symptoms, and 13 (68%) of 19 with advanced symptoms. Four (17%) of 23 healthy non-SBI-exposed controls and two (10%) of 20 non-exposed women with classic autoimmune diseases were positive for APA. Thus, women with moderate or advanced symptoms were significantly more likely than those with limited or mild symptoms, or non-exposed controls to have APA (p < 0.001). The proportion with positive ANA results was higher for women with classic autoimmune diseases 14 (70%) of 20 than for any SBI-exposed subgroup (0-33%). INTERPRETATION: The APA assay can objectively contribute to distinguishing between SBI recipients with limited or mild signs and symptoms. SBI recipients with more severe manifestations, and patients with specific autoimmune diseases. Further studies will be needed to define the signs and symptoms associated with exposure to SBI.

Incompletely reverse-transcribed human immunodeficiency virus type 1 genomes in quiescent cells can function as intermediates in the retroviral life cycle.

Using a quantitative polymerase chain reaction (PCR) method, we have previously shown that a molecularly cloned isolate of human immunodeficiency virus type 1 (HIV-1) can efficiently enter quiescent primary lymphocytes; however, the reverse transcription process is not completed in these cells. In this study, we further characterized the reverse transcription of HIV-1 in quiescent cells, and our results indicate that while initiation of reverse transcription occurs simultaneously in both activated and quiescent lymphocytes, it not only ends prematurely but also proceeds more slowly in quiescent cells. We also performed experiments to address the role of partial reverse transcripts as intermediates in the viral life cycle. We used azidothymidine either before or after infection with HIV-1 to prevent formation of and further DNA synthesis by partial reverse transcripts, respectively. Decreases in virus production from these cells following mitogenic stimulation indicated that partial reverse transcripts can contribute significantly to virus rescue from infected quiescent cells stimulated subsequent to infection. Furthermore, we established that mitogenic stimulation of infected quiescent cells induces reinitiation of DNA synthesis from partial reverse transcripts. However, the virus rescue is inefficient relative to the initial multiplicity of infection, and this is explained by inefficient completion of DNA synthesis from the partial reverse transcript. Thus, the arrest of reverse transcription in quiescent cells may play an important role in HIV-1 pathogenesis by contributing to the inefficient infection of potential target cells in the peripheral blood of HIV-1-infected individuals.

Failure to detect human T-cell leukemia virus-related sequences in multiple sclerosis blood.

We tested 11 patients with multiple sclerosis for the presence of human T-cell leukemia virus type I (HTLV-I)- or type II (HTLV-II)-related sequences. DNA from blood mononuclear cells was analyzed by the polymerase chain reaction utilizing three different oligonucleotide primer pairs. Two of these primer pairs detect sequences shared between HTLV-I and HTLV-II in either p24, gag protein, or in p21, env transmembrane protein. The third primer pair was synthesized based on regions in the pol gene where amino acid sequences are conserved between HTLV-I, HTLV-II, and the related bovine leukemia virus. The multiple sclerosis samples were consistently negative while appropriate control samples were positive. We conclude that viruses related to HTLV-I, HTLV-II, or bovine leukemia virus are not present in the blood of patients with multiple sclerosis and, therefore, that HTLV-bovine leukemia virus-related viruses are not likely to be involved in the pathogenesis of multiple sclerosis.