Effect of dual-subtype vaccine against feline immunodeficiency virus infection

Dual-subtype feline immunodeficiency virus (FIV) vaccine, consisting of inactivated cells infected with subtypes A (Petaluma strain) and D (Shizuoka strain), was developed and tested for its vaccine efficacy against FIV infection in specific pathogen free (SPF) cats. Animals were monitored for proviral DNA by FIV-specific PCR and for FIV-specific antibody profiles by ELISA and virus-neutralization assays. In addition, blood from challenged cats was inoculated into naive SPF cats to confirm the viral status of the vaccinated cats. All cats immunized with Petaluma vaccine alone were protected against homologous Petaluma challenge, but only one of four cats was protected against heterologous Shizuoka challenge. More importantly, all cats immunized with the dual-subtype vaccine were protected against both Petaluma and Shizuoka challenges. These results suggest that a multi-subtype vaccine approach may provide the broad-spectrum immunity necessary for vaccine protection against strains from different subtypes.     

Cytokine production by cats infected with feline immunodeficiency virus: a longitudinal study

The immune responsiveness of cats naturally or experimentally infected with feline immunodeficiency virus (FIV) was studied. Peripheral blood mononuclear cells (PBMC) from naturally infected, symptomatic animals displayed depressed proliferation and interleukin-2 (IL-2) production in response to mitogens, which was accompanied by a significant increase in IL-1, IL-6 and tumour necrosis factor (TNF) production. Longitudinal studies were performed over a period of 4 years in experimentally infected animals. The responses of cells from these cats to concanavalin A (Con A) were consistently less than those from uninfected cats throughout the period but, owing to variation between cats, were significantly lower on only a few occasions. By contrast, the responses of cells to pokeweed mitogen (PWM) were severely affected and declined progressively throughout the 4-year period. In general, responses to Con A but not PWM could be restored by the addition of exogenous IL-2. The decline in immune responsiveness was concurrent with a decline in feline (f)CD4+ cells and an inversion in the CD4:CD8 ratio. Peak production of IL-1, IL-6 and TNF coincided with periods of depressed immune responses. Additionally, immunodeficient responses and elevated levels of proinflammatory cytokines were concurrent with the presence of clinical signs. We conclude that, like human immunodeficiency virus (HIV), FIV infection results in significant perturbation of the immune response. Responses to PWM appear to correlate with disease progression which suggests that the CD3 pathway is affected in the earlier stages of the disease and that additional activation pathways such as CD2 may not be affected until the animal enters the acquired immune deficient syndrome (AIDS) stage of the disease.     

Apoptosis enhanced by soluble factor produced in feline immunodeficiency virus infection

Feline immunodeficiency virus (FIV)-infected cells have been shown to undergo apoptosis by treatment with tumor necrosis factor alpha (TNF-alpha). This study detected a soluble factor which enhanced the apoptosis induced by TNF-alpha treatment. The sensitivity to TNF-alpha in the induction of apoptosis in a feline fibroblastoid cell line (CRFK) cells was significantly enhanced when the culture supernatant of FIV-infected CRFK cells or plasma samples from FIV-infected cats was added to the culture. These findings suggested that FIV infection induces production of a soluble factor which enhances CRFK cells sensitivity to TNF-alpha induction of apoptosis both in vitro and in vivo. This factor may contribute to the loss of lymphocytes in cats infected with FIV.     

Autologous and heterologous neutralization analyses of primary feline immunodeficiency virus isolates

Feline immunodeficiency virus (FIV) provides a model system with which the significance of neutralizing antibody (NA) in immunosuppressive lentivirus infections may be studied. To date, no detailed analysis of the neutralization properties of primary FIV isolates has been reported. In this study, we have conducted the first comprehensive study of the sensitivity to autologous and heterologous neutralization in a lymphoid cell-based assay of 15 primary FIV isolates and, for comparison, of one tissue culture-adapted strain. Primary isolates in general proved highly NA resistant, although there was considerable individual variation. Variation was also observed in the capacity of immune sera to neutralize heterologous FIV isolates. The ability of sera to neutralize isolates or for isolates to be neutralized by sera did not correlate with epidemiological and genetic relatedness or with the quasispecies complexity of the isolates. From the study of specific-pathogen-free cats experimentally infected with viral isolates associated with NA of different breadths, it appears that the development of FIV vaccines cannot rely on the existence of viral strains inherently capable of inducing especially broad NA responses.     

Biological nature of feline immunodeficiency virus

Genetic factors were recognized as possible predisposing factors in the etiopathogenesis of prepubertal periodontitis (PP). Consequently, the present work was carried out to determine the possible mode of inheritance and if there were any associated chromosomal aberrations in cases with PP. The study included 3 families with probands manifesting PP (the families included 18 individuals). Pedigree analysis was carried out as well as chromosomal analysis. The results pointed out to the possibility of new mutations arising due to various environmental factors (including the use of pesticides and their handling), in addition to a possible autosomal recessive inheritance. Chromosomal analysis showed no association between a certain chromosomal aberration and PP. An interesting finding was that a girl proband proved to be Xo female, i.e. a case of Turner's syndrome. This could be considered the first report of Turner's syndrome with PP.     

Plasma viral RNA load predicts disease progression in accelerated feline immunodeficiency virus infection

Viral RNA load has been shown to indicate disease stage and predict the rapidity of disease progression in human immunodeficiency virus type 1 (HIV-1)-infected individuals. We had previously demonstrated that feline immunodeficiency virus (FIV) RNA levels in plasma correlate with disease stage in infected cats. Here we expand upon those observations by demonstrating that plasma virus load is 1 to 2 logs higher in cats with rapidly progressive FIV disease than in long-term survivors. Differences in plasma FIV RNA levels are evident by 1 to 2 weeks after infection and are consistent throughout infection. We also evaluated humoral immune responses in FIV-infected cats for correlation with survival times. Total anti-FIV antibody titers did not differ between cats with rapidly progressive FIV disease and long-term survivors. These findings indicate that virus replication plays an important role in FIV disease progression, as it does in HIV-1 disease progression. The parallels in virus loads and disease progressions between HIV-1 and FIV support the idea that the accelerated disease model is well suited for the study of therapeutic agents directed at reducing lentiviral replication.     

Antibody-secreting cells specific for simian immunodeficiency virus antigens in lymphoid and mucosal tissues of immunized macaques

OBJECTIVES: To examine whether the route of immunization affects the induction of antibody-secreting cells (ASC) in the circulation of macaques. The distribution of ASC in the rectal mucosa and lymphoid tissues following challenge with simian immunodeficiency virus (SIV) was investigated. DESIGN: Macaques were immunized with recombinant SIV gp120 and p27 antigens by the targeted iliac lymph node (TILN) route of immunization or the nasal and rectal route, augmented by intramuscular immunization [naso-rectal intramuscular (NRI)]. The macaques were challenged with live SIV by the rectal route and ASC were assayed in the circulation before and after SIV challenge, and in the tissues removed at post-mortem. METHODS: ASC were examined in the circulation by Elispot assay. Mononuclear cells were prepared from peripheral blood, iliac and axillary lymph nodes and spleen. Rectal tissue was treated by enzyme digestion to elute mononuclear cells. RESULTS: TILN and NRI immunization induced circulating IgA and IgG ASC to both gp120 and p27. Following rectal challenge with SIV, TILN macaques were protected from infection whereas NRI route-immunized and unimmunized controls became infected. IgA ASC to p27 were increased significantly in the iliac lymph nodes of the TILN immunized macaques compared with unimmunized controls (P < 0.05). Only IgA ASC were found in the rectal mucosa of the immunized protected macaques but both IgA and IgG ASC were detected in the unimmunized infected macaques. Overall the number of IgG ASC specific for p27 was significantly higher in the infected NRI and control macaques than in the protected macaques (P < 0.02). A progressive increase in IgG but not IgA ASC was detected in the peripheral blood mononuclear cells of the unimmunized infected macaques. CONCLUSIONS: The results suggest that cells secreting IgA antibodies to p27 in the iliac lymph nodes of the TILN immunized macaques correlate significantly with protection from infection. The unimmunized infected macaques showed a progressive increase in IgG ASC in the peripheral blood after SIV challenge; this was found in the iliac and axillary lymph nodes and also in the spleen, suggesting that it is an immune response to the SIV infection.     

Comparison of T-cell subpopulations in cats naturally infected with feline leukaemia virus or feline immunodeficiency virus

The antimutagenicity of the Citrus flavonoids naringin, hesperidin, nobiletin, and tangeretin against the mutagens benzo[a]pyrene, 2-aminofluorene, quercetin, and nitroquinoline N-oxide was investigated in the Salmonella/microsome assay. Naringin and hesperidin showed a weak antimutagenic activity against benzo[a]pyrene. Tangeretin was antimutagenic against all indirectly-acting mutagens tested, but in general a large molar excess was necessary. Liquid preincubation increased the antimutagenicity of tangeretin against 2-aminofluorene. Nobiletin acted as an antimutagen against benzo[a]pyrene, but it enhanced the mutagenicity of 2-aminofluorene. However, in a liquid preincubation assay nobiletin also exhibited antimutagenicity against 2-aminofluorene. Both tangeretin and nobiletin inhibited the mutagenicity of quercetin. Quercetin itself acted as an antimutagen against 2-aminofluorene in a Salmonella strain (TA1538) where its mutagenicity was not expressed. Quercetin should not merely be regarded as a genotoxic risk factor in the human diet, since its mutagenicity may be inhibited by accompanying compounds including other flavonoids, and since quercetin itself also exhibits an antimutagenic action. Because of the antimutagenic properties the Citrus flavonoids tested, especially tangeretin and nobiletin, might play a role in the chemoprevention of cancer.     

Neurotoxic effects of feline immunodeficiency virus, FIV-PPR

Apoptotic neuronal death is known to occur in the developing brain and in the mature brain of patients with ischemic and degenerative disorders. Although microglial cells are known to become activated in specific conditions, it has not been elucidated whether they enhance or prevent neuronal apoptosis. The present study was intended to observe how microglial cells are involved in neuronal death. When rat primary cortical neurons were incubated with a nitric oxide (NO) donor sodium nitroprusside (SNP; 300 microM) for 10 min, neuronal death occurred 12-16 hr later. The NO-induced neuronal death was inhibited by cycloheximide, and the SNP-treated neurons were characterized by nuclear fragmentation and intact cell membrane under electron microscopy. Agarose gel electrophoresis demonstrated DNA fragmentation of the SNP-treated neurons. Thus, the NO-induced neuronal death appeared to be apoptosis. When neurons were cocultured with rat primary microglial cells, the SNP treatment failed to induce the neuronal death. Because microglia-conditioned medium also prevented apoptotic neuronal death, microglial cells were considered to secrete antiapoptotic factors. The microglia-conditioned medium rescued neurons even when they were added to neuronal cultures after the SNP treatment, implying that the factors acted on neurons in a manner other than scavenging NO. Interleukin-3, interleukin-6, macrophage colony-stimulating factor, and basic fibroblast growth factor, which are known to be secreted by microglial cells, were not effective in preventing NO-induced neuronal death. Among microglia-derived substances, tumor necrosis factor alpha and plasminogen, which are heat-labile proteins, inhibited neuronal apoptosis. The neuroprotective action of the microglia-conditioned medium, however, still remained, even after it was heated. These findings suggest that microglial cells protect neurons against NO-induced lethal damage by secreting heat-labile and heat-stable neuroprotective factors in vitro.     

Renal involvement in feline immunodeficiency virus infection: p24 antigen detection, virus isolation and PCR analysis

In this work we analyze the following biomaterial: biomaterials used in this study are: reabsorbable Dac Blu, non reabsorbable Dac Blu, non reabsorbable atomized Dac Blu, non reabsorbable thin Dac Blu, reabsorbable Biocoral 450, Calcitite 2040-12, Orthogel, Apagen, BTF 65, Calcitite 4060-2, Osprogel, Bio-Oss, Biostite, Osprovit, Merck Hydroxiapatite. The quantitative XRF analysis was performed by a Philips PW 1480 with Rh tube. This research, besides underlining the possibility of applying the XRF method to the analysis of biomaterials. This study, shows the facility and the rapidity in the preparation of samples and standards in the form of tablets to undergo the analysis: furthermore the study shows the possibility of verify the analysis on the same sample in the future, because the tablet if well conserved, does not deteriorate. We can also verify a good analytic accuracy both for the principal elements (Ca, P) and for trace elements. The analyses show a moderate variability in the Ca/P ratio in the hydroxylapatites, and a greater variability in the secondary and trace elements.     

Cytokine networks with infection: mycobacterial infections, leishmaniasis, human immunodeficiency virus infection, and sepsis

Distinct cytokine profiles are clearly associated with and relate to the severity of several types of infections. Cytokine networks are apparent with selected human infectious diseases, such as mycobacterial infections (leprosy, tuberculosis), the parasitic infection leishmaniasis, human immunodeficiency virus (HIV) infection, and gram-negative sepsis. Cytokine profiles are determined to some extent by two functional subsets of T lymphocytes, Th1 and Th2. The Th1 cytokines (interferon gamma, interleukin-2 [IL-2], IL-12) enhance cell-mediated immunity, inhibit humoral immunity, and result in protective effect for pathogens that are removed primarily through cell-mediated immunity (Mycobacterium tuberculosis, Mycobacterium leprae, Leishmania). The Th2 cytokines (IL-4, IL-5, IL-10, IL-13) enhance humoral immunity and inhibit cell-mediated immunity, and result in protective effect for pathogens removed primarily through humoral mechanisms. Progression of HIV infection is associated with a switch from a Th1 to a Th2 profile. For sepsis, uncontrolled activation of proinflammatory cytokines (IL-1, tumor necrosis factor-alpha, interferon-gamma) may be a fundamental defect that promotes the detrimental aspects of inflammation, whereas Th2 cytokines may be beneficial in controlling inflammation. Knowledge of basic cytokine immunopharmacology, networks, and relationships with infectious processes will aid clinicians in determining treatment approaches that are likely to be effective.     

Human immunodeficiency virus induction of corticotropin in lymphoid cells

Disruption of the linkage among the immune, nervous, and endocrine systems may contribute to the pathology and symptoms of acquired immunodeficiency syndrome (AIDS). We investigated the role of human immunodeficiency virus (HIV) in altering these linkages via induction of corticotropin (ACTH) by lymphocytes. Cultured T lymphocytes (H9 cell line) were infected with HIV-1, after which ACTH production was measured and characterized at various time intervals by immunofluorescence and Western blotting. We report a coordinate expression of ACTH and p24 HIV core protein in H9 cells. Also, the kinetics of HIV-induced ACTH production by H9 T lymphoma cells are demonstrated using three different strains of HIV as well as UV-inactivated HIV. ACTH production corresponded with the appearance of p24 antigen and was maximal 35 days after infection. UV-inactivated HIV and the viral envelope protein, gp120, were also able to induce ACTH production in these cells, indicating that viral replication was not required for the ACTH induction. The HIV-induced ACTH was synthesized de novo and had the size and biological activity of pituitary ACTH. Inhibition of ACTH in HIV-infected lymphocyte cultures by anti-ACTH antiserum enhanced viral p24 expression. The significance of lymphocyte ACTH in AIDS is not clear, but these results suggest that it may restrict HIV replication and possibly infection.     

DNA vaccination affords significant protection against feline immunodeficiency virus infection without inducing detectable antiviral antibodies

To test the potential of a multigene DNA vaccine against lentivirus infection, we generated a defective mutant provirus of feline immunodeficiency virus (FIV) with an in-frame deletion in pol (FIVDeltaRT). In a first experiment, FIVDeltaRT DNA was administered intramuscularly to 10 animals, half of which also received feline gamma interferon (IFN-gamma) DNA. The DNA was administered in four 100-microg doses at 0, 10, and 23 weeks. Immunization with FIVDeltaRT elicited cytotoxic T-cell (CTL) responses to FIV Gag and Env in the absence of a serological response. After challenge with homologous virus at week 26, all 10 of the control animals became seropositive and viremic but 4 of the 10 vaccinates remained seronegative and virus free. Furthermore, quantitative virus isolation and quantitative PCR analysis of viral DNA in peripheral blood mononuclear cells revealed significantly lower virus loads in the FIVDeltaRT vaccinates than in the controls. Immunization with FIVDeltaRT in conjunction with IFN-gamma gave the highest proportion of protected cats, with only two of five vaccinates showing evidence of infection following challenge. In a second experiment involving two groups (FIVDeltaRT plus IFN-gamma and IFN-gamma alone), the immunization schedule was reduced to 0, 4, and 8 weeks. Once again, CTL responses were seen prior to challenge in the absence of detectable antibodies. Two of five cats receiving the proviral DNA vaccine were protected against infection, with an overall reduction in virus load compared to the five infected controls. These findings demonstrate that DNA vaccination can elicit protection against lentivirus infection in the absence of a serological response and suggest the need to reconsider efficacy criteria for lentivirus vaccines.     

Viral dynamics of primary viremia and antiretroviral therapy in simian immunodeficiency virus infection

Mathematical modeling of viral replication dynamics, based on sequential measurements of levels of virion-associated RNA in plasma during antiretroviral treatment, has led to fundamental new insights into human immunodeficiency virus type 1 pathogenesis. We took advantage of the simian immunodeficiency virus (SIV)-infected macaque model to perform detailed measurements and mathematical modeling during primary infection and during treatment of established infection with the antiretroviral drug (R)-9-(2-phosphonylmethoxypropyl)adenine (PMPA). The calculated clearance half-life for productively infected cells during resolution of the peak viremia of primary infection was on the order of 1 day, with slightly shorter clearance half-lives calculated during PMPA treatment. Viral reproduction rates upon discontinuation of PMPA treatment after 2 weeks were approximately twofold greater than those obtained just prior to initiation of treatment in the same animals, likely reflecting accumulation of susceptible target cells during treatment. The basic reproductive ratio (R0) for the spread of SIV infection in vivo, which represents the number of productively infected cells derived from each productively infected cell at the beginning of infection, was also estimated. This parameter quantifies the extent to which antiviral therapy or vaccination must limit the initial spread of virus to prevent establishment of chronic disseminated infection. The results thus provide an important guide for efforts to develop vaccines against SIV and, by extension, human immunodeficiency virus.     

Accumulation of human immunodeficiency virus-specific cytotoxic T lymphocytes away from the predominant site of virus replication during primary infection

Down-regulation of the initial burst of viremia during primary human immunodeficiency virus (HIV) infection is thought to be mediated predominantly by HIV-specific CD8+ cytotoxic T lymphocytes (CTL). This response is associated with major perturbations in the T cell receptor (TCR) repertoire. To investigate the failure of the cellular immune response to adequately control viral spread and replication and to prevent establishment of HIV infection, changes in the TCR repertoire and in the distribution of virus-specific CTL between blood and lymph node were analyzed in three patients with primary infection. By the combined use of clonotype-specific polymerase chain reaction and analysis of the frequency of in vivo activated HIV-specific CTL, it was shown that HIV-specific CTL clones preferentially accumulated in blood as opposed to lymph node. Accumulation of HIV-specific CTL in blood occurred prior to effective down-regulation of virus replication in both blood and lymph node. These findings should provide new insights into how HIV, and possibly other viruses, elude the immune response of the host during primary infection.