Human Immunodeficiency Virus P24 Research Articles

Serologic markers for staging human immunodeficiency virus infections

In the present study, 80 serum specimens were tested for human immunodeficiency virus (HIV) antibody by enzyme immunoassay (EIA) and indirect fluorescent antibody tests, HIV (p24) antigen, and Western Blot analyses. A total of 40 specimens were HIV antigen-positive, 35 were antigen-negative and five were indeterminant. Among the 40 antigen-positive sera, 38 had positive antibodies by EIA with confirmation by Western Blot. Two cases were antigen-positive and were thought to be early stages where antibodies had not yet developed. Among the 38 sera, 30 (79%) had decreased or had no reactions by indirect fluorescent antibody tests. Among the 35 antigen-negative cases, all 35 had positive antibodies by EIA and all 35 had bands at gp41 by Western Blot. Among 84 HIV-infected patients, 30 had >400 CD4+ cells per cubic millimeter, 21 patients had 200–400 CD4+ cells and 33 had <200 CD4+ cells. A total of 28 (93%) of the HIV antigen-negative cases with full banding patterns by Western Blot had >400 CD4+ cells. In contrast 18 (55%) of the patients with antigen positivity and incomplete banding on Western Blot had <200 CD4+ cells.

Clinical experience with zidovudine for patients with acquired immune deficiency syndrome and acquired immune deficiency syndrome-related complex

We have treated 113 patients with zidovudine since its licensure, 80 with acquired immunodeficiency syndrome and 33 with acquired immunodeficiency syndrome-related complex. This paper reports on the efficacy and toxicity observed in these patients. Improved well-being, reduced frequency and severity of opportunist infections were notable in the first year of follow-up. More rapid improvement in pulmonary physiological tests during recovery from Pneumocystis carinii pneumonia was also observed in treated patients. Patients with lower initial platelet counts showed early increases in platelet counts. There was a consistent fall in human immunodeficiency virus (HIV) p24 antigen during treatment, although not always to undetectable levels. CD4 cell counts showed a rise in the first months of treatment but these were not sustained, despite continuing clinical benefit. Neuropsychological and clinical evidence of benefit in HIV encephalopathy are described. We have analysed the factors influencing marrow toxicity and have found that low CD4 count and the intercurrent use of ganciclovir and dapsone increase myelotoxicity. We describe the clinical and biochemical features of the myopathy associated with long-term use of zidovudine and summarise our findings on dose-reduction associated meningo-encephalitis.

Mononuclear phagocytes of blood and bone marrow: comparative roles as viral reservoirs in human immunodeficiency virus type 1 infections.

We examined human immunodeficiency virus (HIV) production in cultured mononuclear cells from 36 seropositive homosexual males. Production was detected by using an HIV p24 antigen ELISA assay in blood mononuclear cells in 54% of asymptomatic, 71% of acquired immunodeficiency syndrome (AIDS)-related complex, and 100% of AIDS patients. When the peripheral blood mononuclear cells were separated into monocytes and CD4+ T cells, we found that the CD4+ T-cell fraction was preferentially infected in the three clinical stages. The ability to isolate HIV from blood monocyte-derived macrophages was similar in the three stages (24-33%) and required coculture with phytohemagglutinin-stimulated lymphoblasts. Bone marrow and blood mononuclear cells cultured simultaneously yielded virus from both sources in 13 individuals. Again the prime source of virus was the nonadherent bone marrow mononuclear cells, which contained CD4+ T cells, and not the adherent monocyte-enriched fraction. We conclude that blood mononuclear cells yield productive virus more readily as disease progresses and that infection is detected in association with CD4+ T-cell-enriched fractions. In our large sample of patients, monocyte infection was detected in only a small fraction, suggesting that this cell type is neither a primary nor an exclusive reservoir of HIV infection.

ANTI-RETROVIRAL EFFECTS OF INTERFERON-α IN AIDS-ASSOCIATED KAPOSI'S SARCOMA

21 patients with AIDS and Kaposi's sarcoma were enrolled in an open therapeutic trial to determine the in vivo anti-retroviral activity of recombinant interferon-α (IFN-α). 8 (38%) showed a complete or partial anti-tumour response. The mean pretreatment CD4 count for the responders was 399 cells/μl vs 154 cells/μl for the non-responders. All 5 of the patients with more than 400 CD4 cells/μl pretreatment showed a significant reduction in tumour, whereas none of the 7 patients with under 150 CD4 cells/μl had any response. 5 of the 6 complete or partial responders with greater than 50 pg/ml of human immunodeficiency virus (HIV) p24 before IFN therapy showed a 75% or greater reduction by 12 weeks of therapy, with 3 patients having persistently negative HIV cultures. The anti-viral effects were also most pronounced in the patients with the highest CD4 counts. These data demonstrate the potential benefits, both anti-tumour and anti-retroviral, of treatment with IFN-α in the early stages of HIV infection and Kaposi's sarcoma.

Human immunodeficiency virus gp120 and p24 oligoclonal antibody in acquired immunodeficiency syndrome cerebrospinal fluid and sera.

Human immunodeficiency virus gp120 and p24 oligoclonal antibody in acquired immunodeficiency syndrome cerebrospinal fluid and sera.

The influence of human immunodeficiency virus (HIV) infection on antibody responses to influenza vaccines.

To ascertain whether subjects infected with human immunodeficiency virus (HIV) generally develop protective hemagglutination inhibition antibody responses to inactivated influenza vaccines. Prospective study of 104 persons before and after immunization. Outpatient clinic and hospital ward. Persons with the acquired immunodeficiency syndrome (AIDS) (n = 25), AIDS-related complex (n = 14), and HIV-seropositive men with only lymphadenopathy or no symptoms (n = 27). Controls were HIV-seronegative homosexual men (n = 22) and HIV-seronegative heterosexuals (n = 16). Subjects were immunized with inactivated vaccines containing 15 micrograms of each of the following influenza virus hemagglutinins: A/Taiwan/1/86 (HINI), A/Mississippi/1/85 (H3N2), A/Chile/1/83 (HINI), and B/Ann Arbor/1/86. Fourfold or greater antibody responses occurred less frequently in subjects with HIV infections than in HIV-seronegative controls. Protective levels (1:64 or greater) of hemagglutination inhibition antibodies were attained by 94% to 100% of HIV-seronegative controls, 52% to 89% of HIV-seropositive asymptomatic subjects, and 13% to 50% of subjects with AIDS or AIDS-related complex. No increase in the prevalence or level of serum HIV p24 antigen or clinical deterioration was detected among HIV-infected persons after influenza immunization. Because of the poor antibody responses to influenza vaccines among HIV-infected subjects, even in many with no or minimal symptoms, alternative strategies for preventing influenza, such as booster doses of influenza vaccine, prophylaxis with amantidine, or both should be considered.

Entamoeba histolytica antigen-specific induction of human immunodeficiency virus replication

Replication of human immunodeficiency virus (HIV) may be initiated in infected lymphocytes by antigenic or mitogenic stimulation. A soluble protein derived from an invasive strain of Entamoeba histolytica (amoebic antigen [AA]) was used to study the lymphoblastic responses of T lymphocytes derived from 8 HIV-seronegative homosexual men (controls) and 15 HIV-seropositive homosexual men (patients). The soluble protein was also used in long-term cultures as a stimulus for HIV replication. No control or patient produced detectable lymphoblastic responses to AA in a 6-day tritiated-thymidine incorporation assay. Of 15 patients, 5 (33%) produced HIV p24 (ranging from 31 pg/ml to 151 ng/ml) in response to AA in 30-day cell cultures. HIV p24 was expressed in three of seven patients in response to AA but not to the T-lymphocyte mitogen phytohemagglutinin. Implications for managing HIV-infected patients are discussed.

Detection of human immunodeficiency virus by antigen capture ELISA in culture fluids and blood specimens

A newly developed HIV p24 antigen capture ELISA was compared with the reverse transcriptase assay (RT) for the detection of human immunodeficiency virus (HIV) in culture fluids of virus-infected lymphocytes. The comparison was made by testing 350 culture fluids derived from lymphocyte cultures of 88 patients and collected 7, 14, 21 and 28 days after co-cultivation with lymphocytes from healthy donors. For 87 out of 88 patients there was agreement in the results of the two methods (98·9%) but the antigen capture assay gave positive results 7–14 days earlier than the RT assay with culture fluid specimens from five patients; specimens from only one patient gave discrepant results with a constantly negative RT activity but positive HIV p24 antigen detection at the second and third passage in culture. We also investigated the ability of the antigen capture assay to detect the antigen directly in 246 clinical specimens (including serum, plasma, lymphocytes and cerebrospinal fluid) from 100 seropositive patients, and found that 16% of the patients were positive. These and the former results compared well with those obtained using commercial antigen capture assays. A similar percentage of positivity (16·2%) was observed in a panel of 37 serum specimens from seropositive patients whose antibody response pattern was evaluated by Western blot analysis. Besides, a higher percentage of positivity (66·6%) was observed in a panel of 12 plasma specimens from hemophiliac children for whom virus isolation was also performed.

Treatment of the acquired immunodeficiency syndrome (AIDS) and AIDS-related complex with a regimen of 3'-azido-2',3'-dideoxythymidine (azidothymidine or zidovudine) and acyclovir. A pilot study.

On the basis of observation that acyclovir potentiates the in-vitro antiviral activity of 3-azido-2',3'-dideoxythymidine (also known as azidothymidine or zidovudine) against human immunodeficiency virus (HIV), we administered a regimen of azidothymidine and acyclovir to eight patients with the acquired immunodeficiency syndrome (AIDS) or AIDS-related complex. An oral regimen of 100 mg of azidothymidine and 800 mg of acyclovir every 4 hours was in general well tolerated, with the principal toxicity being megaloblastic erythroid changes. The pharmacokinetics of the two drugs were independent of each other. Six patients received the drug combination for at least 10 weeks; all had increased numbers of T4+ lymphocytes (P = 0.028), and two of three assessable patients had reversal of anergy. Two patients tested positive for serum HIV p24 antigen at entry, but became negative with treatment. Data for this small group suggest that this drug combination can be tolerated in patients with severe HIV infections; this study can be used as a basis for larger studies of this drug combination.

Seropositivity for HIV and the development of AIDS or AIDS related condition: three year follow up of the San Francisco General Hospital cohort.

The three year actuarial progression rate to the acquired immune deficiency syndrome (AIDS) in a cohort of men in San Francisco who were seropositive for the human immunodeficiency virus (HIV) was 22%. An additional 26 (19%) developed AIDS related conditions. Beta 2 Microglobulin concentration, packed cell volume, HIV p24 antigenaemia, and the proportion and number of T4 lymphocytes each independently predicted progression to AIDS. Beta 2 Microglobulin was the most powerful predictor. The 111 subjects tested who were normal by all predictors (40%) had a three year progression of 7%, and the 68 subjects who were abnormal by two or more predictors (24%) had a progression rate of 57%. Two thirds of all men who progressed to AIDS were in the last group. The median T4 lymphocyte count in subjects who did not progress to AIDS fell from 626 x 10(6) to 327 x 10(6)/l. HIV p24 antigenaemia developed in 7% of the subjects per year. The proportion who were abnormal by two or more predictive variables rose to 41%. At three years an estimated two thirds of the seropositive subjects showed clinical AIDS, an AIDS related condition, or laboratory results that were highly predictive of AIDS. It is concluded from the observed rates and the distribution of predictive variables at three years that half of the men who were seropositive for HIV will develop AIDS by six years after the start of the study, and three quarters will develop AIDS or an AIDS related condition.

Biochemical and immunological analysis of human immunodeficiency virus gag gene products p17 and p24.

Human immunodeficiency virus (HIV) p24 was purified to homogeneity and subjected to NH2-terminal sequencing. The sequence determined perfectly corresponded to the amino acid sequence predicted from the nucleotide sequence of a middle portion of the HIV first open frame: the gag gene. Edman degradation of purified HIV p17 revealed instead a blocked NH2 terminus. Hybridomas secreting monoclonal antibodies to p24 and p17 were developed and used to immunologically characterize these two HIV gag gene products. They identified two gag precursor polyproteins in the cytoplasm of HIV-infected cells: Pr53gag, which corresponds to the primary translational product, and Pr39gag, which corresponds to an intermediate product of cleavage of Pr53gag. These monoclonal antibodies allowed us also to study posttranslational modification of HIV p24 and p17. p24 was found to be phosphorylated, which is a very unusual feature for a major retroviral core protein. p17 was found to be myristylated, as are all NH2-terminal gag proteins of the known human retroviruses.

Detection of HIV p17 antigen in lymphocytes but not epithelial cells from cervicovaginal secretions of women seropositive for HIV: implications for heterosexual transmission of the virus.

Human immunodeficiency virus (HIV) has been isolated from cervicovaginal secretions from infected women and is thought to be cell associated. To identify which cells harbour viral antigen, we used monoclonal antibodies to OKT4 and a monoclonal antibody directed against HIV p17 core antigen to perform indirect immunofluorescence assays of genital secretions from 17 HIV seropositive and 17 HIV seronegative women with leucorrhoea. OKT4 positive lymphocytes were detected in all tested samples. HIV p17 antigen was detected in the genital fluid lymphocytes in nine out of 14 seropositive subjects from whom lymphocytes were available. No viral antigen was detected in genital fluid lymphocytes of seronegative subjects, nor in any cervicovaginal epithelial cells. This study shows that lymphocytes are the major source of HIV in cervicovaginal secretions of infected women. Conditions that increase the lymphocyte population in the female genital tract, such as sexually transmitted disease (STD), chronic inflammation of the cervix, and menstruation, may facilitate the transmission of HIV during sexual intercourse.

PHASE I STUDIES OF 2',3'-DIDEOXYCYTIDINE IN SEVERE HUMAN IMMUNODEFICIENCY VIRUS INFECTION AS A SINGLE AGENT AND ALTERNATING WITH ZIDOVUDINE (AZT)

Five dose regimens of 2',3'-dideoxycytidine (ddC) were administered, intravenously for 2 weeks then orally for 4 or more weeks, to 20 patients with acquired immunodeficiency syndrome (AIDS) or AIDS-related complex (ARC). ddC was well absorbed from the gut and crossed the blood-brain barrier. 10 of the 15 patients who received 0·03-0·09 mg/kg every 4 h had increases in their absolute number of T4 + T cells at week 2 (p < 0·05), though in many these rises were not sustained. 11 of 13 evaluable patients had a fall in their serum human immunodeficiency virus (HIV) p24 antigen by week 2 of therapy (p < 0·01); in 4 patients the p24 antigen subsequently rose to baseline while in others the decline was sustained. Dose-related toxic effects included cutaneous eruptions, fever, mouth sores, thrombocytopenia, and neutropenia. A reversible painful peripheral neuropathy developed in 10 patients after 6-14 weeks' treatment. These results suggest that ddC has activity against HIV in vivo and has a different toxicity profile from that of zidovudine (AZT). 6 patients with AIDS or ARC were given an alternating regimen of oral AZT (200 mg every 4 h for 7 days) and oral ddC (0·03 mg/kg every 4 h for 7 days). The regimen was well tolerated, and the 5 patients who completed 9 or more weeks of treatment had sustained rises in their T4 + T cells and/or falls in p24 antigen.

Localization of human immunodeficiency virus antigens in infected cells by scanning/transmission-immunogold techniques.

An application of high resolution scanning/transmission electron microscopy (STEM) and gold-labelling techniques for the rapid detection of human immunodeficiency virus (HIV) in infected cells has been developed. Experimental in vitro studies for detecting two HIV structural proteins, gp41 and p17, were performed following an indirect labeling procedure that uses monoclonal anti-p17 and anti-gp41 antibodies as primary antibodies and 40 nm gold-linked goat antimouse IgG as secondary antibodies. The cells were then studied by STEM in the scanning mode. Unambiguous localization of the viral antigens was possible by combining the three-dimensional image provided by the secondary electron image and the atomic number-dependent backscattered electron image for the identification of the gold marker. This technique combines both the morphological information and the rapid procedures of scanning electron microscopy with the precise and sensitive antigen detection provided by the use of STEM and immunological methods. The preliminary results of its application to the study of peripheral blood mononuclear cells from four anti-HIV-seropositive patients showing the presence of specific labeling in all of them suggest that it might prove useful for early detection of HIV infection before seroconversion, as well as for quantitative studies.

Microinjection and expression of an infectious proviral clone and subgenomic envelope construct of a human immunodeficiency virus.

An infectious proviral clone of the human immunodeficiency virus (HIV) was microinjected into the cell nucleus in six cell lines derived from caprine, ovine, bovine, or human solid tissue to study the utility of this method in effecting viral gene expression in nonlymphoid cells. Immunofluorescence assays for HIV demonstrated viral gene expression in only 5% of cells (100-200 cells per line) 24-48 h after microinjection; however, no reverse transcriptase activity was detectable, presumably due to a low level of virus release in this limited number of cells. Therefore, to indirectly assess infectious virus release, microinjected cells were cocultured with human T4 antigen-positive lymphocytes (H9) sensitive to HIV infection. Syncytia formation, electron microscopy, reverse transcriptase activity, and radioimmunoassay for HIV p24 were used to monitor viral gene expression in cocultures. HIV was efficiently recovered by cocultivating H9 with microinjected cells 48 h after microinjection, regardless of the tissue type or species of origin. H9 syncytia were visualized in some cocultures as early as day 5 but were readily apparent in all experiments on days 7-10. Syncytia induction in H9 was the earliest and most reliable indicator of infectious virus release. A recombinant construct containing a subgenomic envelope gene derived from the proviral clone of HIV was microinjected into human glioblastoma cells. Twenty-four to 48 h after manipulation, 5-20% of microinjected cells were found by immunofluorescence assay to express low levels of a putative gp120. These results suggest a possible approach to producing virus-free HIV envelope antigens in mammalian cells and may be relevant to subunit vaccine development.

Cerebrospinal fluid abnormalities in homosexual men with and without neuropsychiatric findings.

We have studied cerebrospinal fluid obtained from 38 homosexual or bisexual men participating in a prospective study of the neuropsychological disorders associated with human immunodeficiency virus (HIV) infection. Twenty-two subjects had neuropsychiatric findings and seropositivity, 11 asymptomatic subjects had seroconverted within 6 to 24 months, and 5 subjects were seronegative controls. Only 1 had acquired immunodeficiency syndrome-related complex, and none had the acquired immunodeficiency syndrome when initially studied. There was a high rate of cerebrospinal fluid abnormalities in men with neuropsychiatric findings, including pleocytosis (41%), elevated IgG and IgG index (47%), and oligoclonal bands (18%). Even in the absence of neuropsychiatric findings, the asymptomatic HIV-seropositive subjects frequently had spinal fluid abnormalities. Of those with neuropsychiatric findings, HIV p24 antigen was detectable in cerebrospinal fluid in only 1 individual, yet HIV was isolated in 11 of 16 (69%) compared to 2 of 7 (29%) of those without neuropsychiatric findings. The identification of cerebrospinal fluid abnormalities in individuals known to have become infected within 6 to 24 months supports the hypothesis that the nervous system is an early target for HIV.

Human Immunodeficiency Virus Antigenemia

The article by Kessler et al 1 in this issue ofThe Journaldescribes the acute illness associated with human immunodeficiency virus (HIV) infection in four individuals. Of interest was the detection of HIV antigenemia during the course of this self-limited illness before enzyme-linked immunosorbent and Western blot assays for antibody to HIV became positive. The presence of circulating antigen thus helped to establish the diagnosis. Other investigators 2 have reported that HIV antigenemia precedes seroconversion in an unknown proportion of acutely infected individuals. The evidence available to date suggests that the circulating antigen contains the core protein of the HIV (p24) and possibly other epitopes of the virus, and that when antibody to the p24 component of the virus increases to measurable concentrations in serum or plasma, the antigen can no longer be detected. 3 In some individuals, months to years after infection, anti-p24 concentrations fall in association with

Human immunodeficiency virus (HIV) gp120 and p24 oligoclonal antibody (ab) in acquired immunodeficiency syndrome (AIDS) cerebrospinal fluid (CSF) and sera

Human immunodeficiency virus (HIV) gp120 and p24 oligoclonal antibody (ab) in acquired immunodeficiency syndrome (AIDS) cerebrospinal fluid (CSF) and sera

Pathogenesis of HIV and its implications for serodiagnosis and monitoring of antiviral therapy

Human immunodeficiency virus (HIV) is lymphotropic and neurotropic. In vivo clinical and immunological abnormalities develop in a large proportion of long-term HIV antibody seropositive persons. Different stages of HIV infection are marked by expression of HIV genes, production of HIV antibodies, formation of antigen/antibody complexes and clearance of such complexes. Transient HIV antigenemia appearing generally 6–8 wk prior to HIV antibody (HIV-Ab) seroconversion and lasting 3–4 mth is generally seen in acute infection. IgM antibodies predominantly to core proteins may occasionally be detectable when, or just before, IgG antibodies appear. If IgG antibodies to both envelope and core proteins persist in the absence of HIV-Ag the short-term prognosis is relatively good. However, HIV-Ag seroconversion may appear at any time after HIV-Ab seroconversion. Progression to AIDS is strongly associated with declining or absent levels of IgG antibodies to p24. IgG2 and IgG4 antibodies to HIV, which are mainly directed to p24, disappear most dramatically. Titers of antibodies to HIV p24 below 64 are strongly associated with the presence of HIV antigen and a poor clinical outcome.