Opioids and the progression of simian AIDS.

This review is a concise description of the study undertaken to examine the modulation by opioids of simian acquired immunodeficiency syndrome (SAIDS) induced by inoculation of rhesus monkeys with simian AIDS virus SIVmac239. The study showed that the replication rate of the virus was several times greater in monkeys made dependent on morphine than in those of non-morphine treated monkeys. Further, a significant change in the mutation rate of the infecting virus in morphine-treated monkeys resulted in the production of mutants that were silent to conventional serological screening tests as well as resistant to AZT. In addition, opioid and chemokine receptors involved were identified in immune cells and a full comparative spectrum of the compromise of the immune system was examined allowing subsequent studies to evaluate wherein the modulation of the development of the syndrome could be better characterized. The data gathered to date are unique and germane to furthering our understanding of AIDS in humans and its subsequent treatment thereof.

Methadone induces CCR5 and promotes AIDS virus infection.

Methadone, a regimen for the treatment of opioid dependency, was found to induce the expression of CCR5, a co-receptor for human immunodeficiency virus (HIV)/simian form of HIV (SIV) entry, on human CEM x174 lymphocytes. Both CCR5 mRNA and protein were elevated in methadone-treated cells. A concomitant increase of mu opioid receptors was also observed. Upon methadone exposure, SIVmac239-infected CEM x174 cells released greater amounts of virus particles as revealed by both the number of syncytia formation and reverse transcriptase activities. Similar methadone effect was not observed on CEM x174 cells infected with other simian retroviruses that do not depend on CCR5 for cellular entry. These studies raise concerns considering methadone as an innocuous morphine substitute.

Morphine suppresses lymphocyte apoptosis by blocking p53-mediated death signaling.

Opiates such as morphine or heroin may promote cell apoptosis and cause dysfunction of immune cells. In simian immunodeficiency virus (SIV)-infected lymphocytic cells, however, morphine may protect the cells from apoptotic lysis and allow the virus to continue to replicate. To further explore this apparently antithetical effect of opiates, we evaluated in the present study the effects of morphine on human lymphocytic CEM x174 cells induced to undergo apoptosis in the presence of actinomycin D. It was found that induction of apoptosis (characterized by DNA laddering) by actinomycin D was accompanied by a stimulation of the expression of active (phosphorylated) form of p53. Pretreatment of the cells with 10nM morphine caused a transient, naloxone-reversible suppression of the appearance of activated p53 and the generation of DNA laddering. Parallel evaluation of the growth of CEM x174 indicated that morphine treatment delays the inception of cell death triggered by actinomycin D. Inasmuch as Bcl-2 suppresses while Bax accelerates apoptosis, treatment of cells with morphine reduced the expression of Bax and enhanced the expression of Bcl-2. Taken together, morphine, through binding at the opioid receptor, may protect lymphocytic cells from apoptotic lysis if cell death is initiated by apoptosis-inducing agents such as human immunodeficiency virus (HIV), SIV or actinomycin D.

Interactions of opioid and chemokine receptors: oligomerization of mu, kappa, and delta with CCR5 on immune cells.

Activation of opioid receptors by morphine was previously shown to specifically induce the expression of chemokine receptor CCR5, promoting simian AIDS virus entry and replication in immune cells. The present study was undertaken to determine whether these two structurally and functionally distinct G-protein-coupled receptors are in close proximity and form an oligomeric complex in the cell membrane so that the activation of one triggers the activity of the other. Both human CEM x174 and monkey lymphocytes were used in this study and gave similar results. Immunoprecipitation experiments showed that CCR5, but not CD4 nor Na(+)/H(+) exchanger, coprecipitates with all three subtypes (mu, delta, and kappa) of opioid receptors. A single protein band immunoreactive with antibodies against both the CCR5 and the opioid receptors was identified after electrophoresis on nondenaturing polyacrylamide gels. Chemical crosslinking experiments using glutaraldehyde or BS(3) indicate that these receptors are closely situated on the cell membrane with an intermolecular distance less than 11.4A. Functional studies revealed that a combination treatment of cells with morphine, an agonist for mu, and MIP-1beta, a ligand for CCR5, suppresses the inhibitory effect of MIP-1beta and increases the stimulatory effect of morphine on CCR5 expression. These results suggest that oligomerization of chemokine receptor CCR5 and opioid receptors on the cell membrane of human or monkey lymphocytes may modulate receptor functions.

Morphine promotes simian acquired immunodeficiency syndrome virus replication in monkey peripheral mononuclear cells: induction of CC chemokine receptor 5 expression for virus entry.

Rhesus monkeys (Macaca mulatta) chronically administered opioids were more susceptible to simian immunodeficiency virus (SIV) strain mac239 (SIVmac239) infection than those without prior exposure to opioids. Increased plasma viremia in morphine-dependent monkeys allowed SIV to be detected in the animals' peripheral blood mononuclear cells (PBMC) without cocultivation with a tissue culture cell line. In contrast, virus titers from the PBMC of morphine-naive SIVmac239-infected animals were undetectable in the absence of cocultivation. PBMC isolated from noninfected animals and treated with morphine sulfate in vitro produced an increase in the expression of beta-chemokine receptor 5 (CCR5). Because both SIVmac239 and human immunodeficiency virus (HIV) require CCR5 for cell entry, the unique role of morphine in promoting SIV infection may provide a mechanism to account for the high incidence of HIV disease among drug-using populations.

Identification of opioid-regulated genes in human lymphocytic cells by differential display: upregulation of Krüppel-like factor 7 by morphine.

Morphine was previously found to compromise the chemotactic mobility of leukocytic cells toward inflammatory sites as well as induce the expression of chemokine receptor CCR5, a coreceptor for HIV entry, on lymphocytes. Both effects increase the host's susceptibility for HIV infection. In this report, we used a reverse transcription-polymerase chain reaction-based differential display (RT-PCR DD), an RNA finger printing procedure, coupled with Northern slot blot hybridization, in identifying gene or genes regulated by morphine in human lymphocytic cells. Ten positive gene segments, including two morphine downregulated genes and eight morphine upregulated genes, were identified. Analysis of the DNA sequences of these gene segments showed high homologies to known DNA and protein sequences on databases, of which two apoptosis/cell growth-related genes (PNAS-133 and Krüppel-like factor 7) were revealed. PNAS-133 gene was downregulated by morphine whereas the Krüppel-like factor 7 (KLF7), a zinc finger transcription factor, was upregulated by morphine. The upregulation of KLF7 by morphine was further studied and found to be at both the transcriptional and the translational levels in a naloxone-reversible manner. Cell growth was promoted by morphine up to day 4 (72 h) post treatment. These findings suggest a unique role for morphine in regulating cell proliferation and differentiation of immune cells.

Chemokine receptor CCR5: polymorphism at protein level.

Polymorphisms in chemokine receptor CCR5 genes have been implicated in HIV disease progression, resistance, or non-progressive infection. Multiple CCR5 transcripts and mRNA diversity have also been identified. This study presents evidence to show that two distinct forms of CCR5 protein, 62 and 42k Da, are present in human lymphocytic cells and monkey peripheral blood mononuclear cells. The ratio of these two forms of CCR5 changes with cell growth. The 62k Da CCR5 predominates if the electrophoresis sample buffer does not contain reducing agent. However, the 62 and the 42 kDa CCR5 are not interconvertible. Morphine sulfate induces the formation and expression of both forms of CCR5 whereas RANTES, MIP-1alpha or MIP-1beta inhibits them. Localization studies indicate that the 62 kDa CCR5 resides mainly on the cell membrane and the 42 kDa CCR5 is present solely in the cytoplasm of the cells.