Presence of Endogenous Viral Elements Negatively Correlates with Feline Leukemia Virus Susceptibility in Puma and Domestic Cat Cells.

While feline leukemia virus (FeLV) has been shown to infect felid species other than the endemic domestic cat host, differences in FeLV susceptibility among species has not been evaluated. Previous reports have noted a negative correlation between endogenous FeLV (enFeLV) copy number and exogenous FeLV (exFeLV) infection outcomes in domestic cats. Since felids outside the genus Felis do not harbor enFeLV genomes, we hypothesized absence of enFeLV results in more severe disease consequences in felid species lacking these genomic elements. We infected primary fibroblasts isolated from domestic cats (Felis catus) and pumas (Puma concolor) with FeLV and quantitated proviral and viral antigen loads. Domestic cat enFeLV env and long terminal repeat (LTR) copy numbers were determined for each individual and compared to FeLV viral outcomes. FeLV proviral and antigen levels were also measured in 6 naturally infected domestic cats and 11 naturally infected Florida panthers (P. concolor coryi). We demonstrated that puma fibroblasts are more permissive to FeLV than domestic cat cells, and domestic cat FeLV restriction was highly related to enFeLV-LTR copy number. Terminal tissues from FeLV-infected Florida panthers and domestic cats had similar exFeLV proviral copy numbers, but Florida panther tissues have higher FeLV antigen loads. Our work indicates that enFeLV-LTR elements negatively correlate with exogenous FeLV replication. Further, Puma concolor samples lacking enFeLV are more permissive to FeLV infection than domestic cat samples, suggesting that endogenization can play a beneficial role in mitigating exogenous retroviral infections. Conversely, presence of endogenous retroelements may relate to new host susceptibility during viral spillover events.IMPORTANCE Feline leukemia virus (FeLV) can infect a variety of felid species. Only the primary domestic cat host and related small cat species harbor a related endogenous virus in their genomes. Previous studies noted a negative association between the endogenous virus copy number and exogenous virus infection in domestic cats. This report shows that puma cells, which lack endogenous FeLV, produce more virus more rapidly than domestic cat fibroblasts following cell culture challenge. We document a strong association between domestic cat cell susceptibility and FeLV long terminal repeat (LTR) copy number, similar to observations in natural FeLV infections. Viral replication does not, however, correlate with FeLV env copy number, suggesting that this effect is specific to FeLV-LTR elements. This discovery indicates a protective capacity of the endogenous virus against the exogenous form, either via direct interference or indirectly via gene regulation, and may suggest evolutionary outcomes of retroviral endogenization.

A soluble envelope protein of endogenous retrovirus (FeLIX) present in serum of domestic cats mediates infection of a pathogenic variant of feline leukemia virus.

T-lymphotropic feline leukemia virus (FeLV-T), a highly pathogenic variant of FeLV, induces severe immunosuppression in cats. FeLV-T is fusion defective because in its PHQ motif, a gammaretroviral consensus motif in the N terminus of an envelope protein, histidine is replaced with aspartate. Infection by FeLV-T requires FeLIX, a truncated envelope protein encoded by an endogenous FeLV, for transactivation of infectivity and Pit1 for binding FeLIX. Although Pit1 is present in most tissues in cats, the expression of FeLIX is limited to certain cells in lymphoid organs. Therefore, the host cell range of FeLV-T was thought to be restricted to cells expressing FeLIX. However, because FeLIX is a soluble factor and is expressed constitutively in lymphoid organs, we presumed it to be present in blood and evaluated its activities in sera of various mammalian species using a pseudotype assay. We demonstrated that cat serum has FeLIX activity at a functional level, suggesting that FeLIX is present in the blood and that FeLV-T may be able to infect cells expressing Pit1 regardless of the expression of FeLIX in vivo. In addition, FeLIX activities in sera were detected only in domestic cats and not in other feline species tested. To our knowledge, this is the first report to prove that a large amount of truncated envelope protein of endogenous retrovirus is circulating in the blood to facilitate the infection of a pathogenic exogenous retrovirus.

Refrex-1, a soluble restriction factor against feline endogenous and exogenous retroviruses.

The host defense against viral infection is acquired during the coevolution or symbiosis of the host and pathogen. Several cellular factors that restrict retroviral infection have been identified in the hosts. Feline leukemia virus (FeLV) is a gammaretrovirus that is classified into several receptor interference groups, including a novel FeLV-subgroup D (FeLV-D) that we recently identified. FeLV-D is generated by transduction of the env gene of feline endogenous gammaretrovirus of the domestic cat (ERV-DCs) into FeLV. Some ERV-DCs are replication competent viruses which are present and hereditary in cats. We report here the determination of new viral receptor interference groups and the discovery of a soluble antiretroviral factor, termed Refrex-1. Detailed analysis of FeLV-D strains and ERV-DCs showed two receptor interference groups that are distinct from other FeLV subgroups, and Refrex-1 specifically inhibited one of them. Refrex-1 is characterized as a truncated envelope protein of ERV-DC and includes the N-terminal region of surface unit, which is a putative receptor-binding domain, but lacks the transmembrane region. Refrex-1 is efficiently secreted from the cells and appears to cause receptor interference extracellularly. Two variants of Refrex-1 encoded by provirus loci, ERV-DC7 and DC16, are expressed in a broad range of feline tissues. The host retains Refrex-1 as an antiretroviral factor, which may potentially prevent reemergence of the ERVs and the emergence of novel ERV-related viruses in cats. Refrex-1 may have been acquired during endogenization of ERV-DCs and may play an important role in retroviral restriction and antiviral defense in cats.

The surface glycoprotein of feline leukemia virus isolate FeLV-945 is a determinant of altered pathogenesis in the presence or absence of the unique viral long terminal repeat.

Feline leukemia virus (FeLV) is a naturally transmitted gammaretrovirus that infects domestic cats. FeLV-945, the predominant isolate associated with non-T-cell disease in a natural cohort, is a member of FeLV subgroup A but differs in sequence from the FeLV-A prototype, FeLV-A/61E, in the surface glycoprotein (SU) and long terminal repeat (LTR). Substitution of the FeLV-945 LTR into FeLV-A/61E resulted in pathogenesis indistinguishable from that of FeLV-A/61E, namely, thymic lymphoma of T-cell origin. In contrast, substitution of both FeLV-945 LTR and SU into FeLV-A/61E resulted in multicentric lymphoma of non-T-cell origin. These results implicated the FeLV-945 SU as a determinant of pathogenic spectrum. The present study was undertaken to test the hypothesis that FeLV-945 SU can act in the absence of other unique sequence elements of FeLV-945 to determine the disease spectrum. Substitution of FeLV-A/61E SU with that of FeLV-945 altered the clinical presentation and resulted in tumors that demonstrated expression of CD45R in the presence or absence of CD3. Despite the evident expression of CD45R, a typical B-cell marker, T-cell receptor beta (TCRß) gene rearrangement indicated a T-cell origin. Tumor cells were detectable in bone marrow and blood at earlier times during the disease process, and the predominant SU genes from proviruses integrated in tumor DNA carried markers of genetic recombination. The findings demonstrate that FeLV-945 SU alters pathogenesis, although incompletely, in the absence of FeLV-945 LTR. Evidence demonstrates that FeLV-945 SU and LTR are required together to fully recapitulate the distinctive non-T-cell disease outcome seen in the natural cohort.

Identification of novel subgroup A variants with enhanced receptor binding and replicative capacity in primary isolates of anaemogenic strains of feline leukaemia virus.

BACKGROUND: The development of anaemia in feline leukaemia virus (FeLV)-infected cats is associated with the emergence of a novel viral subgroup, FeLV-C. FeLV-C arises from the subgroup that is transmitted, FeLV-A, through alterations in the amino acid sequence of the receptor binding domain (RBD) of the envelope glycoprotein that result in a shift in the receptor usage and the cell tropism of the virus. The factors that influence the transition from subgroup A to subgroup C remain unclear, one possibility is that a selective pressure in the host drives the acquisition of mutations in the RBD, creating A/C intermediates with enhanced abilities to interact with the FeLV-C receptor, FLVCR. In order to understand further the emergence of FeLV-C in the infected cat, we examined primary isolates of FeLV-C for evidence of FeLV-A variants that bore mutations consistent with a gradual evolution from FeLV-A to FeLV-C. RESULTS: Within each isolate of FeLV-C, we identified variants that were ostensibly subgroup A by nucleic acid sequence comparisons, but which bore mutations in the RBD. One such mutation, N91D, was present in multiple isolates and when engineered into a molecular clone of the prototypic FeLV-A (Glasgow-1), enhanced replication was noted in feline cells. Expression of the N91D Env on murine leukaemia virus (MLV) pseudotypes enhanced viral entry mediated by the FeLV-A receptor THTR1 while soluble FeLV-A Env bearing the N91D mutation bound more efficiently to mouse or guinea pig cells bearing the FeLV-A and -C receptors. Long-term in vitro culture of variants bearing the N91D substitution in the presence of anti-FeLV gp70 antibodies did not result in the emergence of FeLV-C variants, suggesting that additional selective pressures in the infected cat may drive the subsequent evolution from subgroup A to subgroup C. CONCLUSIONS: Our data support a model in which variants of FeLV-A, bearing subtle differences in the RBD of Env, may be predisposed towards enhanced replication in vivo and subsequent conversion to FeLV-C. The selection pressures in vivo that drive the emergence of FeLV-C in a proportion of infected cats remain to be established.

Feline leukemia virus infection requires a post-receptor binding envelope-dependent cellular component.

Gammaretrovirus receptors have been suggested to contain the necessary determinants to mediate virus binding and entry. Here, we show that murine NIH 3T3 and baby hamster kidney (BHK) cells overexpressing receptors for subgroup A, B, and C feline leukemia viruses (FeLVs) are weakly susceptible (10(1) to 10(2) CFU/ml) to FeLV pseudotype viruses containing murine leukemia virus (MLV) core (Gag-Pol) proteins, whereas FeLV receptor-expressing murine Mus dunni tail fibroblast (MDTF) cells are highly susceptible (10(4) to 10(6) CFU/ml). However, NIH 3T3 cells expressing the FeLV subgroup B receptor PiT1 are highly susceptible to gibbon ape leukemia virus pseudotype virus, which differs from the FeLV pseudotype viruses only in the envelope protein. FeLV resistance is not caused by a defect in envelope binding, low receptor expression levels, or N-linked glycosylation. Resistance is not alleviated by substitution of the MLV core in the FeLV pseudotype virus with FeLV core proteins. Interestingly, FeLV resistance is alleviated by fusion of receptor-expressing NIH 3T3 and BHK cells with MDTF or human TE671 cells, suggesting the absence of an additional cellular component in NIH 3T3 and BHK cells that is required for FeLV infection. The putative FeLV-specific cellular component is not a secreted factor, as MDTF conditioned medium does not alleviate the block to FeLV infection. Together, our findings suggest that FeLV infection requires an additional envelope-dependent cellular component that is absent in NIH 3T3 and BHK cells but that is present in MDTF and TE671 cells.

Distinctive receptor binding properties of the surface glycoprotein of a natural feline leukemia virus isolate with unusual disease spectrum.

BACKGROUND: Feline leukemia virus (FeLV)-945, a member of the FeLV-A subgroup, was previously isolated from a cohort of naturally infected cats. An unusual multicentric lymphoma of non-T-cell origin was observed in natural and experimental infection with FeLV-945. Previous studies implicated the FeLV-945 surface glycoprotein (SU) as a determinant of disease outcome by an as yet unknown mechanism. The present studies demonstrate that FeLV-945 SU confers distinctive properties of binding to the cell surface receptor. RESULTS: Virions bearing the FeLV-945 Env protein were observed to bind the cell surface receptor with significantly increased efficiency, as was soluble FeLV-945 SU protein, as compared to the corresponding virions or soluble protein from a prototype FeLV-A isolate. SU proteins cloned from other cohort isolates exhibited increased binding efficiency comparable to or greater than FeLV-945 SU. Mutational analysis implicated a domain containing variable region B (VRB) to be the major determinant of increased receptor binding, and identified a single residue, valine 186, to be responsible for the effect. CONCLUSIONS: The FeLV-945 SU protein binds its cell surface receptor, feTHTR1, with significantly greater efficiency than does that of prototype FeLV-A (FeLV-A/61E) when present on the surface of virus particles or in soluble form, demonstrating a 2-fold difference in the relative dissociation constant. The results implicate a single residue, valine 186, as the major determinant of increased binding affinity. Computational modeling suggests a molecular mechanism by which residue 186 interacts with the receptor-binding domain through residue glutamine 110 to effect increased binding affinity. Through its increased receptor binding affinity, FeLV-945 SU might function in pathogenesis by increasing the rate of virus entry and spread in vivo, or by facilitating entry into a novel target cell with a low receptor density.

Identification of a feline leukemia virus variant that can use THTR1, FLVCR1, and FLVCR2 for infection.

The pathogenic subgroup C feline leukemia virus (FeLV-C) arises in infected cats as a result of mutations in the envelope (Env) of the subgroup A FeLV (FeLV-A). To better understand emergence of FeLV-C and potential FeLV intermediates that may arise, we characterized FeLV Env sequences from the primary FY981 FeLV isolate previously derived from an anemic cat. Here, we report the characterization of the novel FY981 FeLV Env that is highly related to FeLV-A Env but whose variable region A (VRA) receptor recognition sequence partially resembles the VRA sequence from the prototypical FeLV-C/Sarma Env. Pseudotype viruses bearing FY981 Env were capable of infecting feline, human, and guinea pig cells, suggestive of a subgroup C phenotype, but also infected porcine ST-IOWA cells that are normally resistant to FeLV-C and to FeLV-A. Analysis of the host receptor used by FY981 suggests that FY981 can use both the FeLV-C receptor FLVCR1 and the feline FeLV-A receptor THTR1 for infection. However, our results suggest that FY981 infection of ST-IOWA cells is not mediated by the porcine homologue of FLVCR1 and THTR1 but by an alternative receptor, which we have now identified as the FLVCR1-related protein FLVCR2. Together, our results suggest that FY981 FeLV uses FLVCR1, FLVCR2, and THTR1 as receptors. Our findings suggest the possibility that pathogenic FeLV-C arises in FeLV-infected cats through intermediates that are multitropic in their receptor use.

Identification of a retroviral receptor used by an envelope protein derived by peptide library screening.

This study demonstrates the power of a genetic selection to identify a variant virus that uses a new retroviral receptor protein. We screened a random peptide library within the receptor-binding domain of a feline leukemia virus retroviral Envelope (FeLV Env) protein for productive infection of feline AH927 cells. One variant, A5, obtained with altered tropic properties acquired the ability to use the solute carrier protein family 35 member F2 (SLC35F2) as a receptor. The SLC35F2 protein is a presumed transporter of unknown function predicted to encode 8 to 10 transmembrane-spanning regions and is not homologous to any identified retroviral receptor. Expression of the feline SLC35F2 cDNA in nonpermissive cells renders the cells susceptible to infection by A5 virus, with remarkably high titers in the range of 10(5) infectious units per ml. The human SLC35F2 ORF also functioned as the retroviral receptor, albeit at lower efficiency than the feline homologue. The successful selection of a novel molecule, the SLC35F2 transporter/channel-type protein, as a receptor by the FeLV Env backbone suggests that multipass transmembrane proteins may be particularly suited for use in productive viral entry and fusion. The analysis of retroviral Env libraries randomized in the receptor-binding domain offers a viable means to develop viral vectors targeted to specific cell types in the absence of known targeting ligands.

What's New in Feline Leukemia Virus Infection.

Feline leukemia virus (FeLV) is a retrovirus with global impact on the health of domestic cats that causes tumors (mainly lymphoma), bone marrow disorders, and immunosuppression. The importance of FeLV is underestimated due to complacency associated with previous decline in prevalence. However, with this comes lowered vigilance, which, along with potential for regressively infected cats to reactivate viremia and shed the virus or develop clinical signs, can pose a risk to feline health. This article summarizes knowledge on FeLV pathogenesis, courses of infection, and factors affecting prevalance, infection outcome, and development of FeLV-associated diseases, with special focus on regressive FeLV infection.

Molecular cloning of a feline leukemia virus that induces fatal immunodeficiency disease in cats.

A replication-defective variant of feline leukemia virus was molecularly cloned directly from infected tissue and found to induce a rapid and fatal immunodeficiency syndrome in cats. Studies with cloned viruses also showed that subtle mutational changes would convert a minimally pathogenic virus into one that would induce an acute form of immunodeficiency. The data suggest that acutely pathogenic viruses may be selected against by current methods for isolation of the human and simian immunodeficiency viruses.

Enhanced immunosuppressive activity associated with metastatic lymphoma cells.

Earlier reports from our laboratory showed that Abelson virus-induced, highly malignant and liver metastatic RAW117-H10 cells, but not the parental, less metastatic RAW117-P cells, inhibited both T-cell and B-cell mitogen-induced proliferation of syngeneic normal murine spleen cells. Similar inhibition was also noted when RAW117-H10 cell surface molecules extracted with butanol were used instead of whole tumor cells. In this report we describe the suppressive properties of the butanol-extracted RAW117-H10 cell surface molecules on other immune functions and the isolation/purification of a molecule from RAW117-H10 cell butanol extract which shows inhibitory activity. The immunosuppressive molecules also inhibit natural killer cell-mediated cytotoxicity, lymphokine-activated killer cell-mediated cytotoxicity, and bone marrow colony-forming unit-granulocyte-macrophage colony formation, but not colony-forming unit-fibroblast colony formation. The suppressive molecules inhibit interleukin 2 production by the T-lymphocytes. One of the molecules responsible for some of the immunosuppressive activity has been isolated and purified from butanol extracts of the metastatic RAW117-H10 cells by preparative isoelectrofocusing techniques. The suppressive molecule has an isoelectric point of 4.3 with an approximate molecular weight of 70,000. Metastatic RAW117-H10 lymphoma cells therefore express immunosuppressive molecules, which may facilitate their growth and metastasis in vivo.

Inhibition of erythroid colony-forming cells by a Mr 15,000 protein of feline leukemia virus.

The effects of concentrated ultraviolet-inactivated feline leukemia virus (FeLV), the purified Mr 15,000 envelope protein (p15E) of FeLV, or the purified Mr 27,000 structural protein (p27) of FeLV on feline bone marrow mononuclear cells were studied in vitro in methylcellulose cultures. Whole virus and purified viral proteins were from the Kawakami-Theilen isolate of FeLV, which induces erythroid aplasia in cats. Bone marrow mononuclear cells from FeLV-negative young adult cats were preincubated with a medium control, ultraviolet-inactivated whole virus, or the p15E or p27 of FeLV, incubated in methylcellulose cultures for 2 days, and then observed for the formation of colony-forming units-erythroid (CFU-E) and colony-forming units-granulocyte/macrophage. The ultraviolet-inactivated Kawakami-Theilen isolate of FeLV at concentrations of 10 or 20 micrograms of viral protein/5 X 10(4) cells suppressed CFU-E to 66 to 56% of control values but had no significant effect on proliferation of colony-forming units-granulocyte/macrophage. p15E at concentrations of 0.1 to 0.2 micrograms/5 X 10(4) cells decreased CFU-E numbers to 0 to 1% of control values, whereas the same concentration of p27 did not alter CFU-E growth when compared with controls. Neither p15E nor p27 had a significant effect on growth of colony-forming units-granulocyte/macrophage. The erythrosuppressive effects of whole virus and an envelope-derived protein but not a structural core protein suggest that FeLV envelope proteins are important in the selective inhibition of erythrogenesis observed in vivo in FeLV-infected cats.

Biology of feline leukemia virus in the natural environment.

The feline leukemia virus (FeLV) was discovered in 1964 in a cluster of cats with lymphosarcoma. The observed clustering of cases of feline lymphosarcoma suggested that FeLV was an infectious agent for cats. The development of a simple immunofluorescent test for FeLV permitted a seroepidemiological study to be undertaken on the distribution of the virus in cats living in their natural environment. Over 2000 cats were tested, and the results showed conclusively that FeLV is an infectious agent for cats. This finding has now been independently confirmed using three different test procedures. After the infectious nature of FeLV was discovered, a simple FeLV test and removal program was devised to control the spread of the virus in the natural environment. The spread of FeLV was controlled in 45 households by removing the FeLV-infected cats, while in 25 households, where the infected cats were left in contact with the uninfected cats, 12% of the uninfected cats became infected. The ultimate control of FeLV awaits the development of an effective FeLV vaccine, which now seems feasible since we have already experimentally immunized some cats with attenuated FeLV. Although FeLV is infectious for cats there is no evidence that FeLV can infect humans.

Pathogenicity of feline leukemia virus is commonly associated with variant viruses.

Many of the serious diseases resulting from feline leukemia virus (FeLV) infection are associated with the generation of novel variant viruses. The prototype FeLV-A virus is highly stable and circulates in the cat population without apparent antigenic change. However, recombination with cellular oncogenes produces viruses which cause leukemia or other malignant diseases. Other recombinants within the env genes of FeLV-A and endogenous FeLV are recognized as belonging to a second subgroup, FeLV-B, the presence of which is correlated with an increased risk of infection with FeLV and a higher incidence of leukemia. Mutants of FeLV which affect the env gene are phenotypically of a third subgroup, FeLV-C, and have a close association with erythroid aplasia. None of these viruses, apart from FeLV-B, is transmitted further in nature. Therefore the generation of these novel viruses and the production of disease is an inadvertent consequence of FeLV infection.

Feline immunodeficiency syndrome--a comparison between feline T-lymphotropic lentivirus and feline leukemia virus.

A feline T-lymphotrophic lentvirus (FTLV) has recently been isolated from a domestic cat free of feline leukemia virus (FeLV). This virus is distinct from FeLV (an oncornavirus), although they share a common denominator, namely, the ability to cause immunosuppression and induce lymphadenopathy and anemia. Their differences can be revealed by examining the following: the metal requirement for reverse transcriptase activity, the antigenic comparison by Western blot analysis, the different susceptibilities of a variety of feline cells, and the morphology based on electron microscopy. In the serological survey of 1,612 cats surveyed in the USA, 232 (14.4%) were seropositive for antibodies to FTLV, which was lower than for the 42 Canadian cats surveyed of which 8 (19%) were seropositive. Of the 61 cats positive for FeLV, 15 (25%) were also positive for FTLV, giving the impression that coinfection between these two retroviruses plays an important role in the cliniocpathological signs of what was previously thought to be solely an FeLV syndrome.

In vitro selective suppression of feline myeloid colony formation is attributable to molecularly cloned strain of feline leukemia virus with unique long terminal repeat.

Molecularly cloned feline leukemia virus (FeLV)-clone 33 (C-33), derived from a cat with acute myelocytic leukemia (AML), was examined to assess its relation to the pathogenesis of AML and myelodysplastic syndrome (MDS). To evaluate in vitro pathogenicity of FeLV C-33, bone marrow colony-forming assay was performed on marrow cells infected with FeLV C-33 or an FeLV subgroup A strain (61E, a molecularly cloned strain with minimal pathogenicity). The myeloid colony-forming activity of feline bone marrow mononuclear cells infected with FeLV C-33 was significantly lower than that of cells infected with 61E. This suggests that FeLV C-33 has myeloid lineage-specific pathogenicity for cats, and that FeLV C-33 infection is useful as an experimental model for investigating pathogenesis of MDS and AML.

Influence of naturally acquired feline leukemia virus (FeLV) infection on the phagocytic and respiratory burst activity of neutrophils and monocytes of peripheral blood.

The purpose of this study was cytometric evaluation of phagocytic and oxidative burst activity of neutrophils and monocytes in cats naturally infected with FeLV. To conduct the study, the peripheral blood was obtained from 33 cats naturally infected with FeLV. The control group consisted of 30 FeLV-, FIV-, clinically healthy cats. The percentage of phagocytizing neutrophils of peripheral blood was lower in FeLV+ than in FeLV- cats. The percentage of neutrophils and monocytes in which an oxidative burst occurred was lower in FeLV+ than in FeLV-animals. Also an oxidative product formation in neutrophils after E. coli and PMA stimulation was lower in FeLV+ than in FeLV-animals. Obtained results allow to conclude that diminished phagocytic and oxidative burst activity of peripheral blood leukocytes may cause impairment of innate immunity in cats infected with FeLV.

Feline leukemia virus infection downmodulates the production of growth-inhibitory activity by marrow stromal cells.

To determine the role of infected marrow accessory cells in the pathogenesis of viral-associated hematologic disorders, we evaluated whether feline leukemia virus (FeLV) infection alters the cytoadhesive properties of long-term marrow culture (LTMC) stromal cells, the support of stromal-associated progenitors in LTMCs, and the production of progenitor growth-promoting and -inhibiting activities by marrow stromal cells. Our previous studies demonstrated that LTMCs containing FeLV-infected stromal cells generated two- to three-fold higher numbers of total nonadherent cells and nonadherent granulocyte-macrophage progenitors (CFU-GM) compared with uninfected LTMCs. In the present studies, CFU-GM and primitive erythroid progenitors (BFU-E) bound equivalently to FeLV-infected or uninfected LTMC stromal cells in a 2-hour adherence assay. In recharge LTMC studies, the numbers of adherent CFU-GM maintained in cultures containing stromal cells infected with FeLV-A/61E were not significantly different from controls (range 84-191% of uninfected control cultures, p > 0.1); however, the percentages of adherent CFU-GM in S phase of the cell cycle were consistently increased (range 42-62% compared with controls, range 5-23%). FeLV infection had no significant effect on the cell-cycle status of the nonadherent CFU-GM in LTMCs. Agar co-culture assays revealed that multilineage colony-stimulating activity was constitutively and equivalently produced by feeder cell layers consisting of either uninfected or FeLV-infected irradiated heterogeneous LTMC stromal cells, homogeneous marrow stromal fibroblasts, or a fibroendothelial marrow stromal cell line. However, FeLV infection significantly attenuated the soluble progenitor growth-inhibitory activity associated with higher densities of these stromal cells. Assays of conditioned medium from cultures of irradiated stromal cells demonstrated that FeLV infection or hydrocortisone exposure decreased the utilization of glucose, the production of acidic metabolic products, and the constitutive production of active and latent transforming growth factor beta (TGF-beta) bioactivity and TGF-beta 2 immunoreactivity. Levels of macrophage inflammatory protein 1 alpha (MIP-1 alpha) and tumor necrosis factor alpha (TNF-alpha) were undetectable and unchanged in CM samples. Together, these observations suggest that downmodulation of TGF-alpha and/or the basal metabolic status of stromal cells may be responsible for the high basal proliferative activity of adherent CFU-GM in FeLV-infected LTMCs, and by extension, that retroviral infection in vivo could alter hematopoiesis by perturbing the progenitor growth-regulatory and -supportive function of marrow stromal cells.

No benefit of therapeutic vaccination in clinically healthy cats persistently infected with feline leukemia virus.

Therapeutic vaccinations have a potential application in infections where no curative treatment is available. In contrast to HIV, efficacious vaccines for a cat retrovirus, feline leukemia virus (FeLV), are commercially available. However, the infection is still prevalent, and no effective treatment of the infection is known. By vaccinating persistently FeLV-infected cats and presenting FeLV antigens to the immune system of the host, e.g., in the form of recombinant and/or adjuvanted antigens, we intended to shift the balance toward an advantage of the host so that persistent infection could be overcome by the infected cat. Two commercially available FeLV vaccines efficacious in protecting naïve cats from FeLV infection were tested in six experimentally and persistently FeLV-infected cats: first, a canarypox-vectored vaccine, and second, an adjuvanted, recombinant envelope vaccine was repeatedly administered with the aim to stimulate the immune system. No beneficial effects on p27 antigen and plasma viral RNA loads, anti-FeLV antibodies, or life expectancy of the cats were detected. The cats were unable to overcome or decrease viremia. Some cats developed antibodies to FeLV antigens although not protective. Thus, we cannot recommend vaccinating persistently FeLV-infected cats as a means of improving their FeLV status, quality of life or life expectancy. We suggest testing of all cats for FeLV infection prior to FeLV vaccination.