Human Immunodeficiency Virus Proteins Gp120 Research Articles

HIV-associated disruption of mucosal epithelium facilitates paracellular penetration by human papillomavirus

The incidence of human papillomavirus (HPV)-associated epithelial lesions is substantially higher in human immunodeficiency virus (HIV)-infected individuals than in HIV-uninfected individuals. The molecular mechanisms underlying the increased risk of HPV infection in HIV-infected individuals are poorly understood. We found that HIV proteins tat and gp120 were expressed within the oral and anal mucosal epithelial microenvironment of HIV-infected individuals. Expression of HIV proteins in the mucosal epithelium was correlated with the disruption of epithelial tight junctions (TJ). Treatment of polarized oral, cervical and anal epithelial cells, and oral tissue explants with tat and gp120 led to disruption of epithelial TJ and increased HPV pseudovirion (PsV) paracellular penetration in to the epithelium. PsV entry was observed in the basal/parabasal cells, the cells in which the HPV life cycle is initiated. Our data suggest that HIV-associated TJ disruption of mucosal epithelia may potentiate HPV infection and subsequent development of HPV-associated neoplasia.

Increased CDK5 Expression in HIV Encephalitis Contributes to Neurodegeneration via Tau Phosphorylation and Is Reversed with Roscovitine

Recent treatments with highly active antiretroviral therapy (HAART) regimens have been shown to improve general clinical status in patients with human immunodeficiency virus (HIV) infection; however, the prevalence of cognitive alterations and neurodegeneration has remained the same or has increased. These deficits are more pronounced in the subset of HIV patients with the inflammatory condition known as HIV encephalitis (HIVE). Activation of signaling pathways such as GSK3β and CDK5 has been implicated in the mechanisms of HIV neurotoxicity; however, the downstream mediators of these effects are unclear. The present study investigated the involvement of CDK5 and tau phosphorylation in the mechanisms of neurodegeneration in HIVE. In the frontal cortex of patients with HIVE, increased levels of CDK5 and p35 expression were associated with abnormal tau phosphorylation. Similarly, transgenic mice engineered to express the HIV protein gp120 exhibited increased brain levels of CDK5 and p35, alterations in tau phosphorylation, and dendritic degeneration. In contrast, genetic knockdown of CDK5 or treatment with the CDK5 inhibitor roscovitine improved behavioral performance in the water maze test and reduced neurodegeneration, abnormal tau phosphorylation, and astrogliosis in gp120 transgenic mice. These findings indicate that abnormal CDK5 activation contributes to the neurodegenerative process in HIVE via abnormal tau phosphorylation; thus, reducing CDK5 might ameliorate the cognitive impairments associated with HIVE.

Retrograde and anterograde transport of HIV protein gp120 in the nervous system

Neurodegeneration and gliosis are prominent pathological features of subjects with human immunodeficiency virus (HIV) dementia complex (HAD). In these patients, neurodegeneration occurs in uninfected neurons. In addition, these patients develop sensory neuropathy despite the antiretroviral therapy. The HIV protein gp120, which mimics some of the pathological alterations seen in HAD, is retrogradely transported in rodent neurons. However, it is still unclear whether gp120 can also be transported anterogradely and whether axonal transport can occur in the peripheral nervous system (PNS). To determine whether gp120 is transported retrogradely and/or anterogradely, we injected gp120IIIB together with the retrograde tracer fluoro-ruby (FR) or the anterograde tracer 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyamine perchlorate (DiI) into the rat superior colliculi. We discovered that gp120 is retrogradely transported with FR along a direct pathway from the superior colliculus to the retina and anterogradely transported with DiI to several areas of the occipital cortex. To determine whether gp120 is also axonally transported in the peripheral nerves, gp120 and FR were injected into the sciatic nerve. No gp120 immunoreactivity was found in the sciatic nerve or dorsal root ganglia, suggesting that gp120 axonal transport does not occur in the PNS. Gp120 axonal transport may play a role in neuronal injury. Therefore, we examined apoptosis at various time points after gp120 injection. Activated caspase-3 was evident within neurons transporting gp120. These results indicate that axonal transport of gp120 might exacerbate the pathogenesis of HIV-1.

Strategy for purifying maltose binding protein fusion proteins by affinity precipitation

The maltose binding protein (MBP) affinity tag has been extensively used for protein purification. A commercial grade cationic starch could precipitate MBP or an MBP-tagged protein quantitatively by simultaneous addition of 10% (w/v) polyethylene glycol (PEG) and 50 mM calcium chloride. The precipitated MBP or MBP-tagged protein could be selectively dissociated by suspending the precipitate in 1 M NaCl. In the case of a soluble MBP fusion with a fragment of human immunodeficiency virus protein gp120, 38% of the contaminating proteins could be removed by precipitation with PEG/CaCl 2 and 100% of the fusion protein was recovered. In all cases, the purified proteins showed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the expected changes in fluorescence emission spectra upon binding to maltose.

Human immunodeficiency virus (HIV)-1 proteins and cytoskeleton: partners in viral life and host cell death

Cytoskeletal components play a major role in the human immunodeficiency virus-1 (HIV-1) infection. A wide variety of molecules belonging to the microfilament system, including actin filaments and actin binding proteins, as well as microtubules have a key role in regulating both cell life and death. Cell shape maintenance, cell polarity and cell movements as well as cytoplasmic trafficking of molecules determining cell fate, including apoptosis, are in fact instructed by the cytoskeleton components. HIV infection and viral particle production seem to be controlled by cytoskeleton as well. Furthermore, HIV-associated apoptosis failure can also be regulated by the actin network function. In fact, HIV protein gp120 is able to induce cytoskeleton-driven polarization, thus sensitizing T cells to CD95/Fas-mediated apoptosis. The microfilament system seems thus to be a sort of cytoplasmic supervisor of the viral particle, the host cell and the bystander cell's very fate.

Protection of human cerebral neurons from neurodegenerative insults by gene delivery of soluble tumor necrosis factor p75 receptor

Apoptosis plays an important role in neuronal cell death in both chronic and acute human neurodegenerative diseases, including amyotrophic lateral sclerosis, Huntington's disease, cerebral ischemia, and human immunodeficiency virus (HIV) encephalopathy. We evaluated the ability of the extracellular binding domain of a dimeric tumor necrosis factor receptor (p75TNFR) to prevent neurotoxicity and death of human fetal cerebral neurons that were exposed in vitro to toxic agents known to be implicated in human neurological disorders, including tumor necrosis factor (TNFalpha) and the HIV proteins Tat and gp120. The extracellular domain of p75TNFR is capable of binding and neutralizing both soluble and transmembrane-anchored TNFalpha. We efficiently transduced human neurons using adenoviral vectors expressing p75TNFR (Ad.p75TNFR) or a control gene (lacZ). Treatment of control cultures with the toxic agents TNFalpha, TNFalpha plus actinomycin D, or Tat and gp120, induced neurotoxic alterations and apoptotic death of neurons. By contrast, transduction of neurons with Ad.p75TNFR prevented apoptosis and cell death due to these agents. We conclude that viral vector transfer of the p75TNFR gene efficiently protects human neurons from TNFalpha-, Tat- or gp120-induced apoptosis and cell death. These results suggest that p75TNFR transduction of neurons by viral vectors could be therapeutically useful in the treatment of many human neurodegenerative diseases.

Gonadal steroids differentially modulate neurotoxicity of HIV and cocaine: testosterone and ICI 182,780 sensitive mechanism

BackgroundHIV Associated Dementia (HAD) is a common complication of human immunodeficiency virus (HIV) infection that erodes the quality of life for patients and burdens health care providers. Intravenous drug use is a major route of HIV transmission, and drug use is associated with increased HAD. Specific proteins released as a consequence of HIV infection (e.g., gp120, the HIV envelope protein and Tat, the nuclear transactivating protein) have been implicated in the pathogenesis of HAD. In primary cultures of human fetal brain tissue, subtoxic doses of gp120 and Tat are capable of interacting with a physiologically relevant dose of cocaine, to produce a significant synergistic neurotoxicity. Using this model system, the neuroprotective potential of gonadal steroids was investigated.Results17β-Estradiol (17β-E2), but not 17α-estradiol (17α-E2), was protective against this combined neurotoxicity. Progesterone (PROG) afforded limited neuroprotection, as did dihydrotestosterone (DHT). The efficacy of 5α-testosterone (T)-mediated neuroprotection was robust, similar to that provided by 17β-E2. In the presence of the specific estrogen receptor (ER) antagonist, ICI-182,780, T's neuroprotection was completely blocked. Thus, T acts through the ER to provide neuroprotection against HIV proteins and cocaine. Interestingly, cholesterol also demonstrated concentration-dependent neuroprotection, possibly attributable to cholesterol's serving as a steroid hormone precursor in neurons.ConclusionCollectively, the present data indicate that cocaine has a robust interaction with the HIV proteins gp120 and Tat that produces severe neurotoxicity, and this toxicity can be blocked through pretreatment with ER agonists.

Human immunodeficiency virus protein gp120 interferes with beta-adrenergic receptor-mediated protein phosphorylation in cultured rat cortical astrocytes.

1. We have previously shown that acute exposure to the HIV coat protein gp120 interferes with the beta-adrenergic regulation of astroglial and microglial cells (Levi et al., 1993). In particular, exposure to 100 pM gp120 for 30 min depressed the phosphorylation of vimentin and glial fibrillary acidic protein (GFAP) induced by isoproterenol in rat cortical astrocyte cultures. In the present study we have extended our analysis on the effects of gp120 on astroglial protein phosphorylation. 2. We found that chronic (3-day) treatment of the cells with 100 pM gp120 before exposure to isoproterenol was substantially more effective than acute treatment in depressing the stimulatory effect of the beta-adrenergic agonist on vimentin and GFAP phosphorylation. 3. Even after chronic treatment with gp120, no differences were found in the levels and solubility of these proteins. 4. Besides stimulating the phosphorylation of intermediate filament proteins, isoproterenol inhibited the incorporation of 32P into a soluble acidic protein of 80,000 M(r), which was only minimally present in Triton X-100-insoluble extracts. 5. Treatment of astrocytes with a phorbol ester or exposure to 3H-myristic acid indicated that the acidic 80,000 M(r) protein is a substrate for protein kinase C (PKC) and is myristoylated, thus suggesting that it is related to the MARCKS family of PKC substrates. 6. Acute (30-min) treatment with 100 pM gp120 totally prevented the inhibitory effect of isoproterenol on the phosphorylation of the 80,000 M(r) MARCKS-like protein. 7. Our studies corroborate the hypothesis that viral components may contribute to the neuropathological changes observed in AIDS through the alteration of signal transduction systems in glial cells.

Impairment of neutrophil antifungal activity against hyphae of Aspergillus fumigatus in children infected with human immunodeficiency virus.

Human immunodeficiency virus (HIV)-infected patients may acquire invasive aspergillosis without previously recognized risk factors, such as neutropenia or corticosteroid therapy. Because neutrophils (PMNL) are an important component of host defense in aspergillosis, the antifungal activity of PMNL against hyphae of Aspergillus fumigatus in 31 HIV-infected children was assessed. Hyphal damage was unaffected in 15 HIV-infected children with age-adjusted CD4 cell counts > or = 25% of the normal median value; it was decreased in 16 with CD4 cell counts < 25% (both vs. 20 healthy controls, P = .001. Incubation with sera from 12 of 14 HIV-infected children but not with the recombinant HIV proteins gp120, gp41, and p24 suppressed antifungal activity of normal PMNL compared with normal serum (P = .002). Pretreatment of defective PMNL from 5 patients with granulocyte colony-stimulating factor (G-CSF) partially corrected the defect (P = .002). These findings suggest that impaired serum-mediated antifungal activity against Aspergillus hyphae exists in PMNL of HIV-infected patients with low CD4 cell counts; G-CSF may improve this activity.

HIV antibodies in whole saliva detected by ELISA and western blot assays

Paired serum and saliva samples were tested by enzyme-linked immunosorbent assay (ELISA) and Western blot (WB) for the presence of human immunodeficiency virus (HIV) antibodies. The study group included 36 individuals known to be HIV seropositive and 14 healthy, seronegative controls. HIV antibodies were detected in all but one of the saliva samples of the seropositive subjects. In this particular patient, seroconversion was documented 1 week earlier by sequential testings. A further saliva sample obtained 2 months later was ELISA positive for salivary HIV antibodies. Antibodies against HIV proteins gp120 and gp160 were detected by Western blot assay in all saliva specimens taken from HIV seropositive subjects (including the ELISA-negative patient who seroconverted. Antibodies against other viral proteins (p65, p55, p51, gp41, p35, p24 p18) were found in saliva haphazardly without any clear-cut correlation with the clinical stage of the disease. Pretreatment of the saliva with protease inhibitor was essential for the diagnostic use of saliva for the detection of HIV antibodies by Western blot assay. Calculation of the ratio of titres in serum to those in saliva showed the highest ratios in symptomless subjects (mean +/- SD; 1844 +/- 1412) and the lowest in patients with acquired immune deficiency syndrome (AIDS) (mean +/- SD; 811 +/- 445). The ratio of serum to saliva by ELISA showed a positive correlation with salivary flow rate, indicating a dilution of salivary HIV antibodies with increasing salivary flow rate. The gingival bleeding index was negatively correlated with the ratio, supporting the concept that salivary HIV antibodies transudate from blood to saliva via gingival fluid.

Antigenic specificity of antibody-dependent cell-mediated cytotoxicity directed against human immunodeficiency virus in antibody-positive sera

Antibody-dependent cell-mediated cytotoxicity (ADCC) specific for human immunodeficiency virus (HIV) has been described for HIV-infected individuals. To determine the antigenic specificity of this immune response and to define its relationship to the disease state, an ADCC assay was developed using Epstein-Barr virus-transformed lymphoblastoid cell line targets infected with vaccinia virus vectors expressing HIV proteins. The vaccinia virus vectors induced appropriate HIV proteins (envelope glycoproteins gp160, gp120, and gp41 or gag proteins p55, p40, p24, and p17) in infected lymphoblastoid cell lines as demonstrated by radioimmunoprecipitation and syncytia formation with c8166 cells. Killer cell-mediated, HIV-specific ADCC was found in sera from HIV-seropositive but not HIV-seronegative hemophiliacs. This HIV-specific response was directed against envelope glycoprotein but was completely absent against target cells expressing the HIV gag proteins. The ADCC directed against gp160 was present at serum dilutions up to 1/316,000. There was no correlation between serum ADCC titer and the stage of HIV-related illness as determined by T-helper-cell numbers. These experiments clearly implicated gp160 as the target antigen of HIV-specific ADCC activity following natural infection. Vaccines which stimulate antibodies directed against gp160, which are capable of mediating ADCC against infected cells, could be important for protection against infection by cell-associated virus.