Larry Foresman

Neuropathogenesis of Chimeric Simian/Human Immunodeficiency Virus infection In Pig-tailed and Rhesus Macaques

We recently reported that a chimeric simian/human immunodeficiency virus (SHIVKU‐1) developed in our laboratory caused progressive depletion of CD4+T lymphocytes and AIDS within 6 months of inoculation into pig‐tailed macaques (M.nemestrina). None of the pig‐tailed macaques showed productive SHIV infection in the central nervous system (CNS). In this report, we show that by further passage of the pathogenic virus in rhesus macaques [M. mulatta], we have derived a new strain of SHIV (SHIVKU‐2) that has caused AIDS and productive CNS infection in 3 of 5 rhesus macaques infected with the virus. Productive replication of SHIV in the CNS was clearly shown by high infectivity titers and p27 protein levels in brain homogenates, and in 2 of the 3 rhesus macaques this was associated with disseminated, nodular, demyelinating lesions, including focal multinucleated giant cell reaction, largely confined to the white matter. These findings were reminiscent of HIV‐1 associated neurological disease, and our immunohistochemical and in situ hybridization data indicated that the neuropathological lesions were associated with the presence of SHIV‐specific viral antigens and nucleic acid respectively. However, the concomitant reactivation of opportunistic infections in these macaques suggested that such pathogens may have influenced the replication of SHIV in the CNS, or modified the neuropathological sequelae of SHIV infection in the rhesus species, but not in pig‐tailed macaques. Our findings in the two species of macaques highlight the complexities of lentiviral neuropathogenesis, the precise mechanisms of which are still elusive.

Chimeric SHIV that causes CD4+ T cell loss and AIDS in rhesus macaques

Abstract: By animal to animal passage in rhesus and pig‐tailed macaques, we developed a rhesus model of HIV‐1 disease in humans. Rhesus macaques infected with a cell‐free stock of SHIVKU‐2 developed CD4+ T cell loss, primary lentiviral encephalitis and pneumonia, and AIDS. Six of nine rhesus macaques died within eight months post‐inoculation, while the remaining three are at five, five, and eight months post‐inoculation, respectively. Animals infected by either mucosal or parenteral routes of infection had a similar course of infection.

Initial characterization of viral sequences from a SHIV-inoculated pig-tailed macaque that developed AIDS.

Abstract: In this study, we report on the derivation of a pathogenic SIV‐HIV chimeric virus (SHIV) and the initial characterization of the viral sequences from the first (macaque PPc) of a series of pig‐tailed macaques that developed CD4+ T cell loss and AIDS. Viral genes were amplified by PCR from the brain, lymphoid, and kidney tissues and their sequences compared to the original SHIV used to initiate passages in macaques. Our results show that the vpu gene, which was nonfunctional in the original SHIV, now coded for functional protein in macaque PPc. The tat and rev genes had no consensus changes but the nef gene had 4–5 consensus changes, depending on the tissue examined. The gp 120 gene had the highest number of nucleotide and amino acid substitution rates that varied from 0.64% to 1.44% and 1.17% to 3.71%, respectively, again depending on the tissue examined. These results suggest that a constellation of changes accumulated at the genomic level during the derivation of a SHIV that was pathogenic for pig‐tailed macaques.

Early activation of PBMC and appearance of antiviral CD8+ cells influence the prognosis of SIV-induced disease in rhesus macaques.

We studied 15 macaques inoculated with SIV and identified three phases of infection. Phase 1 was characterized by activated lymphocytes in blood and infected cells in the CSF. In phase 2, activated cells were not detected but virus was recovered from mitogen‐stimulated PBMC, while in phase 3, virus was recovered from mitogen‐stimulated PBMC only after depletion of CD8+ lymphocytes, indicating effective control of the virus in peripheral blood. Early development of phase 3 status correlated with a longer period of clinical normalcy.

Simple and Choice Reaction Time Performance in SIV-Infected Rhesus Macaques

It is well established that HIV infection can lead to motor/cognitive disorders in humans. A number of studies have shown that simian immunodeficiency virus (SIV) infection in rhesus macaques parallels many aspects of HIV disease in humans. The purpose of this study was to define further the SIV-infected rhesus macaque as a model of neuro-AIDS. Our objective was to detect movement-related impairments in behaviorally trained, SIV-infected macaques using both simple and choice reaction time tasks. Reaction times (RTs), movement times (MTs), and error types were examined. Nine monkeys were infected with neurovirulent strains of SIVmac, four of which served initially as controls before their inoculation. Seven of the nine monkeys developed simian AIDS within 4 months of inoculation (rapid progressors), while two monkeys survived for more than 1 year postinoculation (slow progressors). Of the rapid progressors, four exhibited slowed reaction times and six showed movement time slowing. One rapid progressor showed evidence of a strategy shift to overcome impaired motor abilities. Monkeys with rapidly progressing SIV-related disease consistently show behavioral abnormalities reflecting underlying neuronal injury. Although the slow progressors also showed RT and/or MT slowing, a role for nonspecific factors related to late-stage simian AIDS could not be ruled out in these cases. The results demonstrate that motor impairments associated with SIV infection in rhesus macaques can be detected using RT and MT measures, further establishing the SIVmac-infected macaque monkey as a viable model of neuro-AIDS.

Significance of macrophage tropism of SIV in the macaque model of HIV disease.

Microglia, alveolar macrophages, and Langerhans cells are representatives of cells of macrophage lineage that are susceptible to infection with HIV‐1 and they play important roles in the pathogenesis of AIDS dementia, lymphoid interstitial pneumonia, and systemic viral invasion from mucosal surfaces, respectively. In contrast, elimination of CD4+ T cells with resultant development of immunosuppression and AIDS is thought to be reflective of the exclusive tropism of the virus for CD4+ T cells. Examination of these concepts in macaques infected with molecularly cloned strains of SIVmac suggested that all strains of the virus are both macrophage‐ and lymphocyte‐tropic and that all aspects of pathogenesis including loss of CD4+ T cells are dependent on infection in both cell types. However, viral clones that caused productive lytic infection in macrophages were less virulent than those which caused persistent nonproductive infection. The former caused subclinical and even immunizing infections, whereas the latter caused activation and productive infection in CD4+ T cells, AIDS, and systemic infection, even after inoculation of the virus on mucosal surfaces. If these findings on SIVmac are relevant to HIV‐1 disease, then demonstration that HIV‐1 isolates are macrophage‐tropic probably does not necessarily correlate with their pathogenic potential. J. Leukoc. Biol. 62: 12–19; 1997.

Passively Administered Neutralizing Serum that Protected Macaques against Infection with Parenterally Inoculated Pathogenic Simian-Human Immunodeficiency Virus Failed to Protect against Mucosally Inoculated Virus

Macaques inoculated orally, vaginally, or parenterally with SHIV(KU-1) develop severe systemic infection, acute loss of CD4+ T cells, and AIDS. We showed in a previous report that passive immunization with neutralizing serum protected macaques against infection with parenterally inoculated pathogenic SHIV given 24 hr later. In the study reported here we asked whether the identical passive immunization protocol would protect macaques against infection with pathogenic SHIV following oral inoculation of the virus. Ten pigtail macaques were inoculated orally with one animal infectious dose of SHIV(KU-1). Four of the 10 had been given pooled anti-SHIV plasma (15 ml/kg) 24 hr earlier, 4 others were given the same dose of anti-SHIV plasma 2 hr after virus challenge, and the 2 remaining animals were used as controls. The neutralizing antibodies failed to protect macaques against infection after mucosal challenge with SHIV(KU-1).

Auditory brainstem responses in a Rhesus Macaque model of neuro-AIDS

Nine rhesus macaques (Macaca mulatta) were inoculated with a combination of two passaged strains of SIVmac (R71 and 17E), both of which are known to be neurovirulent. Auditory brainstem responses (ABRs) were recorded at regular intervals from these animals both before and after inoculation. Increases in ABR peak and interpeak latency were observed corresponding to progression of SIV disease. Post-inoculation increases in latency were observed for all five peaks of the ABR and for interpeak intervals I-V and III-V. The largest increases in latency were associated with end-stage disease. Within 14 weeks of inoculation, all but two animals developed end-stage simian AIDS and were euthanized. Histopathological examination revealed multifocal lesions in the cerebral gray and white matter as well as in the auditory structures of the brainstem. In most animals, ABR changes were accompanied by evidence of underlying neuropathology. However, cases of severe neuropathology with no ABR abnormalities and vice versa were also noted. Though in a much shorter time frame, SIVmac R71/17E produced both physiological and histopathological abnormalities similar to those associated with HIV disease in humans. These results further support the SIVmac R71/17E infected rhesus macaque as an animal model of HIV related neurological disease in humans.

MOTOR SKILL IMPAIRMENT IN SIV-INFECTED RHESUS MACAQUES WITH RAPIDLY AND SLOWLY PROGRESSING DISEASE

Abstract: A number of studies have shown that simian immunodeficiency virus (SIV) infection in rhesus macaques parallels many aspects of HIV disease in humans. The purpose of this study was to further characterize the rhesus macaque infected with neurovirulent SIV as a model of neuroAIDS. Using a motor skill task, our objective was to detect SIV‐related movement impairments in behaviorally trained macaques. The motor skill task required retrieval of a food pellet from a cup in a rotating turntable across a range of speeds. Nine monkeys were infected with neurovirulent strains of SIVmac (R71/17E); four monkeys served initially as controls pre‐inoculation. Seven monkeys developed simian AIDS within 4 months of inoculation (rapid progressors), and two survived more than 18 months post‐inoculation (slow progressors). Of the rapid progressors, five exhibited significant deficits in this task, most showing a gradual decline in performance terminating in a sharp drop to severely impaired levels of performance. One slow progressor (AQ15) showed no performance declines. The other slow progressor (AQ94) showed a significant decrease in maximum speed that was concurrent with the onset of clinical signs. For AQ94, the role of sickness behavior related to late stage simian AIDS could not be ruled out. These results demonstrate that motor system impairment can be detected early in the course of SIV infection in rhesus macaques, further establishing the SIVmac‐infected macaque monkey as a viable model of neuroAIDS.

Motor evoked potentials in a rhesus macaque model of neuro-AIDS

Previous work using bone marrow passaged SIVmac239 (simian immunodeficiency virus) has shown that macrophage tropic strains of this virus enter the rhesus macaque brain early following inoculation (Sharma et al, 1992; Desrosiers et al, 1991; Zhu et al, 1995; and Narayan et al, 1997). As part of an effort to more fully characterize the extent of neurologic impairment associated with SIV infection of the brain, we used transcranial electrical stimulation of motor cortex and the spinal cord to evoke EMG potentials in two forelimb (EDC and APB) and two hindlimb (LG and AH) muscles. The latencies, magnitudes and thresholds of motor evoked potentials (MEPs) recorded from nine monkeys infected with neurovirulent SIVmac R71/17E were compared to pre-inoculation records from the same monkeys. Seven of nine monkeys developed simian AIDS within 4 months of inoculation and were euthanized. Two monkeys remained free of AIDS-related clinical illness for over 18 months following inoculation. Six of the seven monkeys with rapidly progressing disease showed post-inoculation latency increases ( > or = 2 s.d. of control) in at least one cortical MEP. Increases in cortical MEP latency ranged from 21-97% in different monkeys. All seven rapidly progressing animals showed post-inoculation increases in at least one spinal cord MEP latency. Maximum spinal cord MEP latency increases ranged from 22-147%. Increases in central conduction time (CCT) ranged up to 204% and exceeded two standard deviations of control in four monkeys. Neither of the two monkeys with slowly progressing disease showed significant increases in either cortical or spinal cord MEP latency or CCT. Only the monkeys with rapidly progressing disease exhibited classic AIDS-related neuropathology, although there was no consistent relationship between the severity of neuropathology and the extent of MEP abnormalities. In conclusion, our results demonstrate clear deficits in the functional integrity of both central and peripheral motor system structures associated with SIV infection and further support the use of SIV-infected rhesus macaques as a model of neuro-AIDS.