Prophylactic ZDV therapy prevents early viremia and lymphocyte decline but not primary infection in feline immunodeficiency virus-inoculated cats.

Prophylactic zidovudine (ZDV) therapy was evaluated in the feline immunodeficiency virus (FIV)-inoculated cat model for HIV-1 infection in humans. ZDV treatment (30 mg/kg/day via continuous subcutaneous infusion) was initiated 48 h prior to virus inoculation and continued for 28 days. Transient plasma antigenemia evident in six of six untreated cats at week 2 post-inoculation (pi) was absent in the ZDV-treated cats although at 10 and 14 weeks pi (6 and 10 weeks after drug treatment), one of the ZDV-treated cats had low-level antigenemia. Both CD4 and CD8 lymphocyte numbers were consistently higher in the ZDV-treated cats when compared to both the FIV-inoculated untreated cats and the virus-naive, age-matched controls. CD4:CD8 ratios were lower for the ZDV-treated cats than either the FIV-inoculated untreated or virus-naive, control cats. The decreased CD4:CD8 ratios were the result of an increase in CD8 lymphocytes in the ZDV-treated cats while decreased ratios in the FIV-inoculated untreated cats were due to cell loss. Both ZDV-treated and untreated cats showed nearly identical FIV-specific antibody responses beginning 2 weeks pi. Polymerase chain reaction (PCR) results from blood lymphocytes showed that six of six ZDV-treated and six of six untreated cats were positive for FIV-specific gag sequences. Although primary infection was not prevented, these results suggest that prophylactic ZDV therapy deterred early systemic spread of infection mediated by viremia and delayed absolute CD4 and CD8 lymphocyte decline.

Fusion activity dissociated from replication ability in feline immunodeficiency virus (FIV) in human cells.

Multinucleated-giant-cell formation followed by cell killing was observed after cocultivation of the feline immunodeficiency virus (FIV)-producing feline T-cell line 3201/FIV with various human cells, including T-cell lines carrying human T-cell lymphotropic virus type I (HTLV-I). The susceptibility to giant cell formation varied with the cell lines tested. Cocultivation of irradiated 3201/FIV cells with MT-2 cells resulted in giant cell formation as early as 2 h in culture, with striking resemblance to that induced by human immunodeficiency virus (HIV). MT-4 cells (HTLV-I positive) and H9 cells (HTLV-I negative) were less susceptible than MT-2 to the induction of syncytia. MOLT-4 cells (HTLV-I negative) had intermediate sensitivity to syncytia formation. No syncytia were observed in the monocytic cell line U-937 (HTLV-I negative). Syncytia formation between 3201/FIV and MT-2 cells was inhibited by polyclonal cat anti-FIV antisera but not polyclonal cat anti-feline leukemia virus (FeLV) antisera, goat anti-FeLV, uninfected specific-pathogen-free cat serum, human anti-HTLV-I antisera, or normal human and goat serum. Concentrated cell-free FIV supernatant from 3201/FIV also induced giant cells of MT-2 cells that were indistinguishable from those induced by cocultivation. Giant cells and extensive cell killing associated with giant cell formation declined and disappeared within 10 days. Surviving cells appeared to be of normal size and grew continuously without expressing FIV antigen or releasing infective virus. Although Southern blot analysis using probes specific for FIV could not detect proviral DNA in any of the five human cell lines cocultured with irradiated 3201/FIV cells, the polymerase chain reaction (PCR) technique detected FIV-specific DNA in MOLT-4 cells. DNA from the FIV-PCR positive MOLT-4 cells was PCR negative for endogenous FeLV-specific sequences, indicating that the MOLT-4 cell DNA was not contaminated with DNA from feline cells (i.e., 3201 cells). The FIV-MOLT-4 cells remained PCR positive for FIV after 40 passages, suggesting stable integration in the human cell line. These findings indicate that FIV is capable of forming proviral DNA in human T-lymphoid cells by cocultivation, although this FIV-carrying human cell line failed to produce replication-competent viruses.

Loss of neutrophil and natural killer cell function following feline immunodeficiency virus infection.

Experimental infection with the Mt. Airy isolate of feline immunodeficiency virus (FIVMA), a lentivirus isolated from a domestic cat exhibiting signs of an immunodeficiency-like syndrome, results in transient lymphadenopathy, fever, stomatitis, enteritis, neurologic abnormalities, and immunosuppression. The effects of FIVMA infection on neutrophil and natural killer cell (NK) function were examined in vitro. Suppression of neutrophil chemiluminescence (CL) responses, as well as reduction in NK-mediated cytotoxicity were demonstrated. Neutrophil CL was decreased by 50% in infected cats when compared to control values. This loss of CL was present through 6 months after infection. In addition, NK-mediated cytotoxicity was approximately 50% less in FIVMA infected cats than in controls. Loss of innate immunity was paralleled with inversion in feline CD4/CD8 lymphocyte ratios and decreases in lymphocyte mitogenesis seen as early as 5 weeks after infection. These results suggest that FIVMA infection induces an immunodeficiency disorder in infected cats similar to that seen in human immunodeficiency virus infections.

Polymorphonuclear leukocyte function in HIV-1-infected chimpanzees.

The chemiluminescent characteristics of enriched populations of neutrophils from control and HIV-infected chimpanzees were assessed. Neutrophils from HIV-infected chimpanzees were suppressed in their ability to generate a normal response to particulate and soluble stimuli when compared to normal and hepatitis non-A, non B-infected controls. Particulate (latex beads) stimulation of neutrophils resulted in an aberrant response when contrasted with controls. Normal control responses were characteristically biphasic while the response from hepatitis NANB HIV-infected chimpanzees was not biphasic. Neutrophils challenged with a soluble (phorbol ester) stimulant also demonstrated a suppressed response. These data suggest that HIV infection has an additive suppressive effect on neutrophil function in chimpanzees previously infected with hepatitis NANB. The suppression of chimpanzee neutrophil function following HIV infection is similar to that seen in other non-primate viral and retroviral infections.

Immunodeficiency in latent feline leukemia virus infections.

Challenge of naive experimental animals with a retroviral inoculum may result in one of two broad sequelae. The first is the establishment of an appropriate humoral and cellular immune response leading to a condition of immunity to subsequent infection with the retrovirus. Alternatively, the host may fail to develop a successful immune response, resulting in a chronic viremia associated with immunosuppression and ultimately death due to secondary pathogens. An alternate disease course is the establishment of a latent infection characterized by the presence of neutralizing antibody and strong cellular immune reactivity. Recent data from the feline leukemia virus (FeLV) system suggest that cats infected with this virus may develop immunosuppression in the form of persistent neutrophil dysfunction. The potential effect of this cellular dysfunction is the possible susceptibility of the host to the same opportunistic pathogens which are responsible for the increased mortality noted in chronic FeLV infections. These data demonstrate that persistent retroviremia is not essential for the establishment of immunosuppression. This overview presents data accumulated from the feline model of the human acquired immunodeficiency syndrome (AIDS) and discusses its relationship to human retroviral infections.

FeLV-induced immunosuppression through alterations in signal transduction: down regulation of protein kinase C.

Activation of protein kinase C by a phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), was shown to stimulate the respiratory burst in normal cat neutrophils. Neutrophils from feline leukemia virus (FeLV)-exposed viremic and nonviremic cats had significant suppression of their respiratory burst when stimulated with TPA. The addition of whole ultraviolet light-inactivated FeLV and FeLV proteins to normal cat neutrophils produced no significant suppression of the respiratory burst. These data suggest two possible mechanisms for suppression. The first is partially due to viral alterations of the neutrophil as seen in viremic cats, but, because exogenously applied FeLV or FeLV proteins had no effect on the respiratory burst, an additional mechanism is present. The second mechanism may be caused by a latent FeLV infection residing in nonviremic cat bone marrows which alters their immune system, resulting in immunosuppression.

Evaluation of terminal deoxynucleotidyl transferase expression in bone marrow of clinically normal and feline leukemia virus-exposed cats.

Bone marrow cells from 8 specific-pathogen-free and 11 feline leukemia virus-exposed cats were examined for the expression of the nuclear antigen terminal deoxynucleotidyl transferase (TdT). Using a standard indirect immunofluorescence technique, feline leukemia-exposed cats had increased expression of TdT in bone marrow aspirates (2.0% to 29.0%) when compared with that in bone marrow cells from specific-pathogen-free cats (2.5% to 6.0%). Seemingly, TdT can be used as an antigenic marker in leukemogenesis of FeLV-infected cats.

Biological effects of staphylococcal protein A immunotherapy in cats with induced feline leukemia virus infection.

Biological effects of staphylococcal protein A (SPA) immunotherapy were studied in 5 viremic and 6 nonviremic cats with induced FeLV infection and in 6 control cats. The SPA therapy neither reversed FeLV viremia nor resulted in consistent improvement in humoral immune responses to FeLV antigens. However, SPA immunotherapy induced a proliferative response in bone marrow granulocytic lineage, possibly resulting in expression of FeLV-free mature neutrophils in the blood. Seemingly, viral burden and chemiluminescent responses were reversed in viremic cats during SPA immunotherapy.

Evaluation of a feline leukemia virus vaccine in a controlled natural transmission study.

Twenty-six 8-week-old specific-pathogen-free cats were vaccinated subcutaneously with 2 doses of a commercially available FeLV vaccine, and 26 age-matched specific-pathogen-free cats were similarly vaccinated with a placebo vaccine containing the same adjuvant as the FeLV vaccine. Cats then were randomly assigned to 2 groups of 26 cats (13 FeLV-vaccinated cats and 13 control cats), and each group was housed with 5 cats previously inoculated with FeLV. All cats were tested biweekly for the next 26 weeks for evidence of FeLV antigenemia. Five of the 26 control cats developed antigenemia. However, 4 of these cats were only transiently antigenemic (positive results for 3 consecutive biweekly samples) and only 1 was persistently antigenemic. None of the FeLV-vaccinated cats developed antigenemia. Preventable fraction was calculated to be 100%.

Demonstration of depressed polymorphonuclear leukocyte function in nonviremic FeLV-infected cats.

Polymorphonuclear leukocytes (PMN) from healthy, normal control cats and FeLV-infected cats were analyzed for differences in phagocytic capabilities. One week after experimental infection, PMNs from FeLV-infected cats exhibited a marked decrease in phagocytic function as determined by the luminol-dependent chemiluminescent response. Depressed PMN function was observed in these cats during the viremic stage of infection and subsequently remained depressed after the cessation of viremia. The data presented here suggest that while nonviremic cats are reported as clinically normal, they may in fact be exhibiting a suppressed PMN function.

Inhibition of phorbol ester-induced neutrophil chemiluminescence by FeLV.

Receptor-mediated activation is accompanied by phospholipid metabolism and by calcium fluctuation resulting in a chemiluminescence (CL) response in the neutrophil. This pathway involves activation of protein kinase C (PKC) and the NADPH oxidase. Artificial stimulants such as phorbol esters, specifically 12-O-tetradecanylphorbol-13-acetate (TPA), circumvent the receptor-mediated pathway and activate PKC resulting in a measurable CL response. Neutrophils from feline leukemia virus (FeLV) exposed cats were tested for their ability to generate a TPA-induced CL response. As compared to the non-FeLV-exposed specific-pathogen-free (SPF) control cat neutrophil CL responses, both viremic and nonviremic FeLV-exposed cats showed significant decreases in their CL responsiveness. Neither ultraviolet light-inactivated FeLV (UV-FeLV) nor protein components (FeLV-p15E and FeLV-p27) caused a significant decrease in the CL responses of the SPF cat neutrophils. The suppressed TPA-induced CL response from FeLV-infected cats may involve an intracellular mechanism not affected in vitro by exposure of the neutrophil to the virus or viral components.

Evaluation of a HTLV-1 subunit vaccine in prevention of experimental STLV-I infection in Macaca nemestrina.

Pig-tailed macaques were vaccinated with a human T-cell lymphotropic virus type I (HTLV-I) subunit vaccine. Vaccinates and controls were challenged with simian T-cell lymphotropic virus type I (STLV-I)-infected cells. Vaccination yielded antibody responses to HTLV-I and STLV-I gag and env precursors. Controls developed HTLV-I and STLV-I antibody to gag and tax protein. Immunization produced syncytium inhibiting antibody and cellular cytotoxicity to virus-infected cells. Reverse transcriptase activity was present in control macaques only, implying that the subunit vaccine was protective against STLV-I infection.

Suppression of in vitro neutrophil function by feline leukaemia virus (FeLV) and purified FeLV-p15E.

Feline neutrophils (PMN) were isolated and exposed to ultraviolet light-inactivated feline leukaemia virus (UV-FeLV) and purified envelope component p15E (FeLV-p15E). Functional capacity of exposed PMN was measured in vitro utilizing the chemiluminescence (CL) response. PMN exposed to UV-FeLV demonstrated depressed CL responses to Ca2+-ionophore A23187 and latex particles. However, FeLV-p15E produced significant suppression in the CL response to A23187 but failed to produce significant alterations in response to latex particles. The data indicate that FeLV-p15E may, in part, be responsible for increased morbidity and mortality among FeLV-infected cats through suppression of the PMN population.

Polymorphonuclear leukocyte dysfunction associated with feline leukaemia virus infection.

The chemiluminescent characteristics of enriched (greater than 95%) peripheral blood polymorphonuclear leukocyte populations (PMN) from normal and feline leukaemia virus (FeLV)-infected cats were investigated. FeLV-infected cats demonstrated a significantly lower (P less than 0.001) PMN chemiluminescent response when compared to the response of normal age-matched controls. Normal PMN treated with FeLV-infected cat serum exhibited a depressed response in comparison to control cells. A titration of serum from infected cats supplemented with normal serum revealed a titratable suppression of chemiluminescence with increasing concentration of serum from the infected cats. However, PMN from FeLV-infected cats treated with normal serum displayed a slight increase in chemiluminescence over the same cells in autologous serum. The addition of inactivated FeLV to normal PMN caused a titratable decrease in chemiluminescence.

Chronic feline leukemia virus infection alters arachidonic acid proportions in vivo and in vitro.

The polyunsaturated omega-6 fatty acid, arachidonic acid ([AA] 20:4n-6), is both the key of the immunoregulatory substances, prostaglandins, and leukotrienes, and an essential component of immune cell membrane phospholipids, providing stability and flexibility to ensure cellular function. To explore possible effects of the physiological burden of viral replication in chronic viral infections on AA availability, plasma total esterified fatty acid (FA) proportions were measured in the feline leukemia (FeLV) model. Plasma FA profiles of 12 specific-pathogen-free cats with chronic infections with Rickard strain feline leukemia virus (FeLV-R) were compared with 12 age- and sex-matched uninfected specific-pathogen-free cats at 4 months after infection. A significant decrease from normal of average AA proportion was found in FeLV-R-infected cat plasma, while other major FA (palmitic, stearic, and oleic and omega-3 FA normally remained present until near death. Since plasma FA content rapidly affects circulating immune cell membrane composition and since FeLV infection also targets immune cells, we compared FA profiles of feline T4-thymic lymphoma 3201 cell membranes that were infected with virulent FeLV-R or less virulent FeLV-A, at 20 days after viral inoculation with sham-inoculated uninfected 3201 cells. Significantly altered FA proportions and ratios of saturated to unsaturated FA found in infected cell membranes were similar to plasma FA changes and paralleled the virulence of the FeLV inoculum. Altered postinfection FA proportions may impart serious functional defects to the immune cells of chronic FeLV-infected cats, contributing to the inability of their immune systems to eliminate FeLV by depleted plasma AA stores and modified cell membrane composition. Decreased AA availability may be an important factor in the cachexia and fatal outcome of FeLV infection.

Feline leukemia virus: current status of the feline induced immune depression and immunoprevention.

This is a review of the current knowledge of feline leukemia virus (FeLV) associated with immune depression observed in cats. It will focus on the clinical and experimental observations associated with feline retroviral infection and presence in vivo and in vitro. We will briefly describe retroviral-associated acquired immune deficiency syndrome associated with FeLV infection in the cat and specific cellular pathology associated with FeLV latency. In addition, we will focus on the action of FeLV-p15E in vitro and describe possible mechanisms of the FeLV-associated immunosuppression observed both in vivo and in vitro. Lastly, we will evaluate the current status of immunoprevention of FeLV. We will not attempt an in-depth analysis of the current literature; our focus is to review current findings as they relate to feline AIDS and immunotherapy.

Lymphocyte and neutrophil dysfunction associated with hepatitis B virus and hepatitis non-A, non-B virus infection in the chimpanzee.

Chimpanzees were examined for the effect of viral hepatitis infections on specific and nonspecific immune response mechanisms. The data suggest that infection with either hepatitis B virus or hepatitis non-A, non-B virus may result in suppression of cellular immune response components. Mitogen-induced lymphocyte proliferation was lower in virus-infected chimpanzees than in naive animals. Neutrophils from virus infected animals exhibited decreased or altered chemiluminescence kinetics.

Expression, release, and characterization of soluble human T-lymphotropic virus-I (HTLV-I) antigens from an infected cell line.

The Human T-lymphotropic virus type I (HTLV-I) infected cell line MT-2 was studied to obtain HTLV-I or related proteins for the purpose of producing an effective vaccine for HTLV-I infection. The cells were characterized as to HTLV-I antigen expression during the cell cycle and antigens released into the culture fluids. MT-2 cell grown in fetal calf supplemented media produced more HTLV-I related antigens during the G2/M phase of the cell cycle. To determine the conditions for maximal release and harvest of HTLV-I associated proteins, the MT-2 cells were grown in RPMI 1640 medium supplemented with serum-free medium. Cell- and virus-free supernatants were collected on day 4, lyophilized, and concentrated 50-fold. The proteins in these supernatants were characterized using SDS-PAGE and western blot using rabbit anti-HTLV-I sera and human adult T-cell leukemia sera. The western-blot analysis indicated that the supernatants obtained from the MT-2 cells grown in serum-free supplemented medium contained detectable amounts of proteins which reacted with human ATL and rabbit anti-HTLV-I sera. The molecular weights of these proteins observed are 68kd, 46kd, 28kd, 24kd, 19kd, and 15kd indicating that gag, env, and pX gene products are present.

Saliva as a source of feline leukemia virus antigen for diagnosis of disease.

An enzyme-linked immunosorbent assay (ELISA) for detection of feline leukemia virus (FeLV) p27 in saliva was tested for its accuracy and sensitivity in diagnosing FeLV infections. Saliva and serum samples from 564 clinical cases were tested with a 99.2% specificity. The overall accuracy of the saliva ELISA reactive to the serum ELISA was 97.9%. Experimentally, the ELISA saliva was the least sensitive in diagnosing early FeLV infections. However, the overall accuracy, ease of use, and simplicity of the test support its use as a screening procedure in clinical practice.