Tobacco smoking is common in HIV-infected patients, and is prevalent among intravenous opiate abusers. Conversely, intravenous opiate abusers are more likely HIV-infected, and opiate abuse is associated with more severe neuroinflammation. Given the coincident use of tobacco smoking among HIV-infected intravenous drug users (IVDUs), we set out to study the effects of smoke exposure, chronic morphine administration, and HIV infection using the NSG humanized mouse model. Our results show that smoke, morphine, and the combination promotes the decline in CD4+ T cells in HIV-infected mice. Further, chronic morphine administration increases the numbers of circulating CD8+ T cells which express the inhibitory receptor PD-1, as well as the cytolytic proteins perforin and granzyme B in the infected mice. We also found that the combination of smoke and morphine inhibited the expression of IL-1α, IL-4 and IL-17A. Finally, the combination of smoke and morphine exposure induces microglial activation following infection, as well as in the absence of HIV infection. To our knowledge, this is the first report to assess the combined effects of smoke and chronic morphine exposure on the inflammation associated with HIV infection, and demonstrate that these two insults exert significant neuroinflammatory activity.