Neuroimmunity, Drugs of Abuse, and neuroAIDS

It has long been postulated that drugs of abuse may represent significant cofactors in the progression of human immunodeficiency virus (HIV)-induced disease. Both HIV infection and drugs of abuse have significant effect on the immune system as well as on the nervous system. In HIV infection, abnormalities in these systems intersect to lead to a constellation of symptoms known as neuroAIDS. Drugs of abuse may synergize with such damage, acting on immune and/or neural cells. However, definitive epidemiological evidence for such an interaction is lacking. Here we review such studies as well as the use of the nonhuman primate/simian immunodeficiency virus system to investigate the interaction of neuroAIDS with drugs of abuse. Furthermore, recent findings on mechanisms of actions of selected drugs reveal the possibility of protective as well as detrimental effects on the central nervous system damage induced by HIV.     

Low pre-infection levels and loss of central memory CD4+ T cells may predict rapid progression in SIV-infected pigtail macaques

CD4⁺ T lymphocyte subsets are targeted to different degrees by SIV infection. We studied central memory, effector memory, naïve, and regulatory T cell levels longitudinally in 11 SIV_mac251-infected pigtail macaques. Depletion of CD28⁺CD95⁺ central memory CD4⁺ T cells, but not other populations, correlated with both SIV viral load and disease progression. A low pre-infection level of central memory CD4⁺ T cells was also predictive of rapid disease progression. If confirmed in larger studies, our results suggest stratifying macaques for baseline central memory CD4⁺ T cells would be useful in defining both the pathogenesis of SIV disease and SIV vaccine efficacy.     

Development and use of SIV-based Integrase defective lentiviral vector for immunization

Integrase (IN) defective lentiviral vectors have a high safety profile and might prove useful as immunizing agents especially against HIV-1. However, IN defective SIV-based vectors must be developed in order to test their potential in the non-human primate models (NHP) of AIDS. To this aim we tested a novel SIV-based IN defective lentiviral vector for its ability to induce sustained immune responses in mice. BALB/c mice were immunized once intramuscularly with a SIV-based IN defective lentiviral vector expressing the model antigen enhanced green fluorescence protein (eGFP). Immune responses were evaluated 90 days after the injection and compared with those elicited with the IN competent counterpart. The IN defective vector was able to efficiently elicit specific and long-lasting polyfunctional immune responses as evaluated by enzyme-linked immunospot (ELISPOT) assays for interferon-γ (IFN-γ) in spleens, bone marrow (BM) and draining lymph nodes, and by intracellular staining (ICS) for IFN-γ, Interleukin-2 (IL-2) and tumor necrosis factor (TNF-α) in both splenocytes and BM cells without integration of the vector into the host genome. This is the first demonstration that an IN defective SIV-based lentiviral vector provides effective immunization, thus paving the way for the construction of IN defective vectors expressing SIV antigen(s) and test their efficacy against a SIV virus challenge in the NHP model of AIDS.     

Cross-modal circuitry between auditory and somatosensory areas of the cat anterior ectosylvian sulcal cortex: a 'new' inhibitory form of multisensory convergence

Examples of convergence of visual and auditory, or visual and somatosensory, inputs onto individual neurons abound throughout the brain, but substantially fewer incidences of auditory-somatosensory neurons have been reported. The present experiments sought to examine auditory-somatosensory convergence to assess whether there is a feature of this type of convergence that might obscure it from conventional methods of multisensory detection. Auditory-somatosensory convergence was explored in cat anterior ectosylvian sulcus (AES) cortex, where higher-order somatosensory area IV (SIV) and auditory field of the anterior ectosylvian sulcus (FAES) share a common border. While neuroanatomical tracers documented a projection from FAES to SIV, physiological studies failed to reveal the bimodal neurons expected from such cross-modal connectivity. Stimulation of FAES through indwelling electrodes also failed to excite any of the SIV neurons examined. However, when stimulation of auditory FAES was combined with somatosensory stimulation, a large majority (66%) of SIV neurons showed a significant response attenuation. FAES-induced response suppression was specific to SIV, could not be elicited by activating other auditory regions and was blocked by the microiontophoretic application of the GABAergic antagonist bicuculline methiodide. Based on these data, a novel, cross-modal circuit is proposed involving projections from auditory FAES to somatosensory SIV, where local inhibitory interneurons 'reverse the sign' of the cross-modal signals to produce auditory-somatosensory suppression. This form of excitatory-inhibitory multisensory convergence has not been reported before and suggests that the level of interaction between auditory and somatosensory modalities has been substantially underestimated.     

SIVmac239对恒河猴B淋巴细胞生长繁殖的抑制作用

目的探讨恒河猴免疫缺陷病毒SIVmac239感染CD4 T淋巴细胞后可否产生某些抑制B淋巴细胞的生长活性因子,为HIV/AIDS进一步的病原学研究和控制提供有益的线索。方法用MTT实验观察不同时间收集的SIVmac239感染的CD4 T淋巴细胞(C8166)上清和未感染SIVmac239的CD4 T淋巴细胞的上清对恒河猴B淋巴细胞(MM133)增殖的抑制作用情况。结果SIVmac239感染的CD4 T淋巴细胞(C8166)上清含有抑制猕猴B淋巴细胞生长的因子,且其抑制作用效果随着作用时间的延长而增加。结论SIVmac239感染CD4 T淋巴细胞(C8166)可产生抑制B淋巴细胞(MM133)生长的因子。     

Structural determinants of insert retention of poliovirus expression vectors with recombinant IRES elements

Although picornaviruses provide attractive vectors for expression of foreign genes, poor genetic stability restricts their use for immunization purposes. A new prototype vector was generated to increase foreign insert retention, by shifting of the initiation codon to a cryptic AUG within the internal ribosomal entry site (IRES) and replacement of IRES domain VI with foreign ORFs. Using our strategy to replace regulatory noncoding sequences with unrelated foreign genetic material, we generated stable poliovirus-based expression vectors with robust long-term expression of foreign ORFs. Our studies revealed that size and predicted secondary structure formed by the heterologous sequences govern long-term retention and efficiency of expression of foreign inserts replacing IRES structures. These observations indicate that, with certain limitations imposed by structural preferences, foreign sequences can functionally replace IRES substructures in stable picornavirus immunization vectors.     

Computer analysis of the amino acid sequences in gp41 of apathogenic African green monkey (AGM) virus,less pathogenic HIV-2 and highly pathogenic SIV and HIV-1 lentiviruses

The bestfit computer program was used to compare the amino acid sequence of the gp160 envelope glycoprotein of an apathogenic AGM and the pathogenic SIV_AGM monkey lentiviruses. It was found that the gp120 envelope glycoproteins of these viruses resembled each other in their functional domains. However, an insert of 40 amino acids was found in the gp41 envelope glycoproteins of the pathogenic SIV_AGM virus in the amino acid sequence between the membrane anchoring sequence and the carboxyterminus. The insert introduced a new RRIR proteolytic cleavage signal into gp41. Comparing HIV-1 gp41 to that of the pathogenic SIV_AGM virus revealed that the HIV-1 sequence contains an RR sequence that also serves as a signal for proteolytic cleavage. Comparing HIV-2 gp41 to the apathogenic and pathogenic simian immunodeficiency viruses revealed that HIV-2 gp41 lacks the above proteolytic cleavage signal. It is hypothesized that the pathogenic human and simian immunodeficiency lentiviruses can be proteolytically cleaved at the carboxyterminus of gp41, releasing two peptides: a) an immunodeficiency 58 amino acid peptide and b) an IL-2-like peptide. The apathogenic AGM virus and the less pathogenic HIV-2 lack one proteolytic cleavage signal in the gp41 amino acid sequence and therefore can release only the IL-2-like peptide but not the immunodeficiency peptide. If indeed the pathogenic SIV_AGM and HIV-1 do release an immunodeficiency peptide, then such a peptide can be regarded as a toxin. Immunization of healthy individuals or HIV-1 patients against the toxic effect of the viral gp41 toxic peptide might prevent damage to the immune system when the virus reactivation leads to ARC and AIDS in infected individuals. Synthetic peptides modeled according to the immunodeficiency peptide (the toxin) can be used to produce anti-toxin antibodies in healthy HIV-1 infected individuals. Such anti-toxin antibodies can be used for passive immunization of AIDS patients or for active immunization of HIV-1 positive individuals prior to ARC or AIDS.     

Productive infection of dendritic cells by simian immunodeficiency virus in macaque intestinal tissues

Dendritic cells (DCs) are potent antigen-presenting cells that likely play multiple roles in human immunodeficiency virus-1 (HIV-1) and simian immunodeficiency virus (SIV) pathogenesis. This paper describes the effects of pathogenic SIV infection on the networks of DCs in rhesus macaque (Macaca mulatta) intestinal tissues. Intestinal tissues were obtained from macaques at different stages of disease following infection with the pathogenic SIV/DeltaB670 isolate. The patterns and levels of expression of SIV and DC-associated mRNAs were examined and quantitated directly in intestinal tissue sections. In situ hybridization was performed for SIV, DC-specific ICAM3-grabbing non-integrin (DC-SIGN), DC-specific lysosome-associated membrane glycoprotein (DC-LAMP), DC-specific C-type lectin 1 (DECTIN-1), CC chemokine receptor 6 (CCR6), CCR7, and macrophage inflammatory protein 3alpha (MIP-3alpha/CCL20) mRNAs and quantitative image analysis was performed to measure mRNA expression levels. To identify the cell types productively infected by SIV, simultaneous in situ hybridization and immunohistochemical staining were performed. The DC networks in macaque intestinal tissues were found to be extensive and although they generally remained intact during the course of SIV infection, there were alterations in the expression of markers for immature DCs. One alteration was an increase in the expression in intestinal submucosa of DC-SIGN, a molecule that binds to HIV-1/SIV and increases its infectivity. Concomitant with this increase, it was found that during AIDS, the population of productively infected cells included DCs, based on co-expression of DC-SIGN and DECTIN-1 mRNAs. These data indicate that SIV infection affects subpopulations of macaque intestinal DCs, including productive infection of DC-SIGN+ DCs, the consequences of which are likely to be ongoing viral propagation and decreased immunostimulatory function.     

In situ tetramer staining

The development of MHC tetramer staining has opened the doors to multiple avenues of new research [Science 274 (1996) 94]. In this review, we will discuss the development and application of in situ MHC tetramer (IST) staining. We describe two independently developed IST staining methodologies and discuss current uses, limitations, future uses and the interesting biology revealed by the use of IST staining.