Lawrence E. Mathes

Methamphetamine enhances cell-associated feline immunodeficiency virus replication in astrocytes

Human immunodeficiency virus (HIV) infection among substance abusers is on the rise worldwide. Psychostimulants, and in particular methamphetamine (METH), have detrimental effects on the immune system as well as causing a progressive neurodegeneration, similar to HIV infection. Many Lentivirinae, including feline immunodeficiency virus (FIV), penetrate into the central nervous system early in the course of infection with astrocytes serving as a reservoir of chronic brain infection. We demonstrate that the FIV-Maryland isolate infects feline primary and cell line (G355-5)-cultured astrocytes only under cell-associated conditions. Infected astrocytes yielded a new astrocytotropic isolate, capable of cell-free infection (termed FIV-MD-A). This isolate contained four amino acid substitutions in the envelope polyprotein resulting in a change in net charge as compared to FIV-MD. Infection for both isolates was dependent upon a functional astrocyte CXCR4 receptor. Methamphetamine increased significantly FIV replication in feline astrocytes for cell-associated infection only, with no effect on peripheral blood mononuclear cells or astrocytes infected with FIV-MD-A. This viral replication was related to proviral copy number, suggesting the effect of METH is at the viral entry or integration into host genome levels, but not at the translational level. Thus, lentiviral infection of the brain in the presence of the psychostimulant METH may result in enhanced astrocyte viral replication, producing a more rapid and increased brain viral load.

Progressive encephalopathy associated with CD4/CD8 inversion in adult FIV-infected cats.

Experimental intravenous challenge of five adult cats with the feline immunodeficiency virus Maryland isolate (FIV-MD) was investigated for its ability to induce neurologic abnormalities associated with the onset of immunodeficiency. Five 8-month-old cats were inoculated with 1000 median tissue culture infective dose of FIV-MD isolate, with five age-matched cats serving as uninfected controls. All FIV-MD-infected cats tested positive for serum antiviral antibodies and plasma viral DNA as detected by polymerase chain reaction at 2, 4, 10, and 16 months postinfection (PI). At 10 and 16 months PI, there was a significant reduction in the CD4/CD8 lymphocyte ratio, with all cats having a CD4/CD8 ratio of 1 or less. Total protein electrophoretic analysis of cerebrospinal fluid demonstrated a significantly increased albumin quotient at 4 and 16 months PI, representing a disrupted blood-brain barrier (BBB). At 16 months PI, all cats demonstrated a preferential increase in frontal cortical slow-wave activity compared with control cats. Serial evaluation of brainstem auditory evoked potential recordings revealed a prolongation of the interpeak latencies times over the study time. At least one abnormality was found over time in visual and somatosensory evoked potential testing in three and four infected cats, respectively. Comparing lymphocyte subtype ratios with neurologic testing revealed that every FIV-MD-infected cat exhibited an abnormality in at least one neurologic functional test with a concurrent CD4/CD8 count ratio of 1 or less. Overall, this study demonstrated that FIV-MD infection in adult cats results in a delayed-onset, progressive encephalopathy that parallels the decline in the CD4/CD8 lymphocyte ratio. Compared with prior information from pediatric FIV-MD-infected cats, these results indicate that age of infection influences the onset and severity of disease and may be associated with CD4 cell depletion in FIV-MD-infected cats, as seen in HIV-1-infected humans.

Enhancement of LFA-1-Mediated T Cell Adhesion by Human T Lymphotropic Virus Type 1 p12I1

Cell-to-cell transmission of retroviruses, such as human T lymphotropic virus type 1 (HTLV-1), is well documented, but the roles of viral regulatory or other nonstructural proteins in the modulation of T cell adhesion are incompletely understood. In this study we tested the role of the HTLV-1 accessory protein, p12I, on LFA-1-mediated cell adhesion. p12I is critical for early HTLV-1 infection by causing the release of calcium from the endoplasmic reticulum to activate NFAT-mediated transcription. We tested the role of this novel viral protein in mediating LFA-1-dependent cell adhesion. Our data indicated that T cells expressing a mutant HTLV-1 provirus that does not produce p12I mRNA (ACH.p12I) exhibited reduced LFA-1-mediated adhesion compared with wild-type HTLV-1-expressing cells (ACH). Furthermore, the expression of p12I in Jurkat T cells using lentiviral vectors enhanced LFA-1-mediated cell adhesion, which was inhibited by the calcium chelator BAPTA-AM, the calcium channel blocker SK&F 96365, and calpeptin, an inhibitor of the calcium-dependent protease calpain. Similar to the intracellular calcium mobilizer, thapsigargin, the expression of p12I in Jurkat T cells induced cell surface clustering of LFA-1 without changing the level of integrin expression. Our data are the first to indicate that HTLV-1 p12I, in addition to enhancing T cell activation, promotes cell-to-cell spread by inducing LFA-1 clustering on T cells via calcium-dependent signaling.

The feline model of neuroAIDS: understanding the progression towards AIDS dementia.

Feline immunodeficiency virus (FIV) is a neurotropic lentivirus that produces a protracted state of immunodeficiency and encephalopathy in the cat. Recent evidence has shown several similarities to the natural progression of human immunodeficiency virus infection (HIV-1) associated degenerative effects on the central and peripheral nervous systems. Similar to HIV-1, FIV-induced encephalopathy neurovirulence is strain dependent, results in progressive immunodeficiency and increasing early peripheral but not brain viral load, preferentially affects the developing nervous system, produces quantifiable behavioural and neurophysiological impairment that is not directly linked to neuronal infectivity, and induces neuronal injury and loss both in vivo and in vitro. This paper highlights the cumulative scientific body of evidence supporting the use of the feline model of neuroAIDS.

A feline large granular lymphoma and its derived cell line.

SummaryA lymphoma cell line (MCC) was derived from an abdominal mass from a 13-yr-old castrated male cat. The cells resemble natural killer precursor cells, have membrane-bound granules, and are positive for chloroacetate esterase, α-naphthyl butyrate esterase, and tartrate-resistant acid phosphatase activities. The MCC cells are negative for rearranged feline T-cell receptor genes, negative for feline T-cytotoxic antigen, Ia, and surface μ, τ, and lambda chains and do not form E-rosettes. The MCC cell line is negative for the feline leukemia virus (FeLV); e.g., negative for exogenous FeLV (exU3) sequences, negative for cytoplasmic and surface FeLV major core protein of 27 000 daltons (p27) by indirect, immunofluorescence assay, negative for helper FeLV by clone 81 assay, and negative for release of soluble FeLV p27 by enzyme-linked immunosorbent assay. Electron microscopy reveals budding type C retrovirus particles and MCC cells react with anti-RD-114 (anti-endogenous feline retrovirus) reference serum. After in vitro infection, MCC replicate FeLV readily, but replication is noncytopathic.

Anti-human Immunodeficiency Virus (HIV) agents are also potent and selective inhibitors of Feline Immunodeficiency Virus (FIV)-induced cytopathic effect: Development of a new method for screening of anti-FIV substances in vitro

The ability of several known anti-HIV substances to inhibit feline immunodeficiency virus (FIV) was tested. The results showed that FIV infection of feline T-cells was almost completely blocked in the presence of all of the agents tested. However, FIV-induced syncytium formation between a human T-cell line (MT-2 cells) and a FIV-infected feline lymphocyte cell line (3201/FIV) was inhibited only by dextran sulfate and pradimicin A. The assay used to measure syncytium inhibition was rapid and did not use potentially hazardous human immunodeficiency virus (HIV)-infected cells. The efficacy results coincided with those of HIV studies.

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.

Frontal lobe neuronal injury correlates to altered function in FIV-infected cats.

Six cats infected intravenously at 8 weeks of age with feline immunodeficiency virus Maryland isolate (FIV-MD), were evaluated at 8 and 14 months of age (6 months and 12 months postinfection, respectively) with high spatial resolution proton magnetic resonance spectroscopy (MRS) of the frontal cortex. Two separate control cat groups were evaluated at 8 months and 16 months of age. Single voxel two-dimensional high-resolution proton magnetic resonance imaging was performed using the PRESS sequence by selecting a 0.125 ml volume of interest in the medial frontal cortex. A significant reduction in both N-acetylaspartate (NAA) and NAA: choline ratio was found in the FIV 14-month-old group compared with FIV 8-month-old cats, and to the respective age-matched control 16-month-old cats. A negative correlation between NAA and CD4 lymphocyte count was seen in the FIV-14 group only. This group of FIV cats also exhibited a higher proportion of quantitative electroencephalographic relative slow wave activity (RSWA) that correlated to lower NAA content in the frontal cortical voxel. Although peripheral blood proviral load increased over time of infection, no correlation was found between proviral blood or lymph node load and NAA values, CD4 lymphocyte counts, or frontal cortical RSWA. Thus, this study demonstrated that neurologic functional disruption of the frontal cortex correlated strongly with neuronal injury and/or loss in FIV-MD-infected cats independent of peripheral proviral load. The ability to define in vivo neurodegeneration further in this animal model helps in understanding the neuropathogenesis of lentivirus infection, and possibly, a means to follow progression and reversibility during the initial stages of brain infection as therapeutic agents are identified.

Immunoglobulin class response to canine distemper virus in gnotobiotic dogs

Serial serum samples from 27 gnotobiotic dogs infected with R252-canine distemper virus (CDV) were tested for anti-viral IgG, IgM and IgA immunoglobulins using an enzyme-linked immunosorbent assay (ELISA). The results were compared retrospectively to clinicopathological course of disease and to previously reported patterns of complement-fixing and virus neutralizing antibody titers determined in these same sera. Virus-specific IgA was never detected in the sera. High levels of IgG correlated with recovery from disease, whereas the antiviral IgM levels were equivalent in both persistently infected animals and those animals which recovered from disease. The inability to sustain a significant antiviral antibody response in either IgM or IgG classes was characteristic of dogs with fatal encephalitis. The data suggests that IgG is the most important Ig class for recovery from disease.

Experimental oncornavirus vaccines in the cat.

An experimental approach to the immunoprophylatic control of feline oncornavirus-mediated diseases has included induction of antivirus immunity and antibodies to the feline oncornavirus-associated membrane (tumor) antigens. A suitable model for exploring the effectiveness of killed oncornavirus vaccines in the cat has been provided by the use of feline sarcoma virus. Immunization of seven pregnant queens over a 6-week period with ultraviolet light-inactivated Gardner-Arnstein feline sarcoma virus resulted in significant protection among 12 kittens challenged with a tumor-forming Dose 90 at 7 days of age. This immunity was not present in kittens challenged at 35 days of age. Among 12 kittens born of queens immunized during pregnancy with ultraviolet light-inactivated Kawakami-Theilen feline leukemia virus and challenged with the same live virus at 4 days of age, significant protection was noted, ranging from prolongation of survival time to complete protection in 3 kittens. In general, the higher the antibody titer in the mother, the more effective the protection afforded the kittens. Immunization of 43 kittens during their first 5 weeks of life with the same vaccines used in adult cats did not immunize sufficiently to protect against feline sarcoma virus challenge at 5 weeks of age. Neutralizing antibody responses in these kittens were significantly lower than in pregnant queens. That kittens of this age are immunologically responsive was established, since complete protection of 9 kittens to feline sarcoma virus was obtained by immunization with a crude tumor extract inactivated with 5 to 7 megarads of gamma-irradiation. All these kittens developed feline oncornavirus-associated membrane antibodies while 3 developed demonstrable levels of virus-neutralizing antibodies. The results of these studies are believed indicative that killed virus vaccines and tumor vaccines can be effective immunoprophylatic measures in the control of RNA tumor virus oncogenesis in the cat. Developments in this model system should be relevant to any consideration given similar vaccines in humans.