The Epigenetic Eeffects of Alcohol, AM630, and JWH-015 on Mmonocyte-Derived Dendritic Cell Function

Previous studies have demonstrated that substances of abuse such as alcohol (Aroor et al., 2014) and marijuana (Yang et al., 2014) play a role in epigenetically modifying gene expression in immune system cells through site-specific histone modifications. Interestingly, THC, the main psychotropic constituent in marijuana, is also known to interact with differentially associated genes responsible for cellular functions such as cell cycle regulation and metabolism (Yang et al., 2014). THC and related synthetic cannabinoids such as JWH-015 and AM630 are functionally similar in that they all bind to the two main cannabinoid receptors, CB1 and CB2 (Fattore & Fratta, 2011). Our own preliminary data analyzing histone modifications clearly revealed the ability of alcohol and the synthetic cannabinoid, JWH-015, to alter the presence of histones H3 and H4 in a previous model where treatments were administered chronically. In this model, alcohol and cannabinoids will be administered to monocyte-derived dendritic cells (MDDCs) at specific time points (24, 48, and 72 hours). Cells will be treated in-vitro with varying concentrations of alcohol (0.05, 0.1, 0.2, 0.3, and 0.4 %), the CB2 receptor agonist: JWH-015 (1,5, and 10 μM), and the CB2 receptor antagonist: AM630 (1,5,10 μM), in order to assess the ability of these substances to epigenetically modify the function of these key immune system cells. H3 and H4 histone quantification will be performed after treatments. The role of histone deacetylases will be confirmed by the use of histone deacetylase inhibitors, TSA and MGCD0103. Results emanating from this study will elucidate the epigenetic mechanisms of alcohol and cannabinoids on dendritic cell regulation

The Immunomodulatory Role of Alcohol on HIV-Infected Monocyte-Derived Dendritic Cells

Alcohol is known to induce inflammation in the presence of the human immunodeficiency virus (HIV). In our previous studies, we revealed that alcohol induces cannabinoid receptors which play a role in the regulation of inflammatory cytokine production in monocyte-derived dendritic cells (MDDC). However, the ability of alcohol to alter MDDC function during HIV infection has not been clearly elucidated yet. To study the potential impact of alcohol on HIV-infected MDDC (confirmed by p24 ELISA), monocytes were isolated from commercially available buffy coats and cultured for 7 days with GM-CSF and IL-4. MDDC were infected with HIV- 1Ba-L and treated with different concentrations of alcohol (0.1% band 0.2%) for 4-7 days. MDDC phenotype, endocytosis, cytokine production, and ability to transmit HIV to T cells were analyzed. Uninfected CD4+ T cells were co-cultured for 7 days with either infected/treated MDDC or the supernatants from infected/treated MDDC. Inflammatory cytokine arrays were performed using supernatants from HIV-infected MDDC treated with alcohol. Results showed that HIV positive MDDC treated with alcohol had higher levels of infection compared to untreated HIV positive controls. CD4+ T cells exposed to HIV-infected MDDC acquired 100-fold higher levels of p24 compared to CD4+ T cells exposed to only supernatants. CD4+ T cells exposed to HIV-infected and alcohol-treated MDDC had higher levels of infection compared to controls. Cytokine array data show dysregulation of cytokine production by alcohol. In addition, MDDC phenotype and endocytic capacity were altered in the alcohol treated MDDC. Our results indicate a crucial role of MDDC in HIV transmission to T cells and provide insights into the inflammatory role alcohol exerts on dendritic cell function in the context of HIV infection. Supported by the National Institute on Alcohol Abuse and Alcoholism award R00AA021264, the National Institute on Drug Abuse award R01DA034547, and the Institute on NeuroImmune Pharmacology at FIU

Alcohol and Cannabinoids Differentially Affect HIV Infection and Function of Human Monocyte-Derived Dendritic Cells(MDDC)

During human immunodeficiency virus (HIV) infection, alcohol has been known to induce inflammation while cannabinoids have been shown to have an anti-inflammatory role. For instance cannabinoids have been shown to reduce susceptibility to HIV-1 infection and attenuate HIV replication in macrophages. Recently, we demonstrated that alcohol induces cannabinoid receptors and regulates cytokine production by monocyte-derived dendritic cells (MDDC). However, the ability of alcohol and cannabinoids to alter MDDC function during HIV infection has not been clearly elucidated yet. In order to study the potential impact of alcohol and cannabinoids on differentiated MDDC infected with HIV, monocytes were cultured for 7 days with GM-CSF and IL-4, differentiated MDDC were infected with HIV-1Ba-L and treated with EtOH (0.1 and 0.2%), THC (5 and 10 μM), or JWH-015 (5 and 10 μM) for 4-7 days. HIV infection of MDDC was confirmed by p24 and Long Terminal Repeats (LTR) estimation. MDDC endocytosis assay and cytokine array profiles were measured to investigate the effects of HIV and substances of abuse on MDDC function. Our results show the HIV + EtOH treated MDDC had the highest levels of p24 production and expression when compared with the HIV positive controls and the cannabinoid treated cells. Although both cannabinoids, THC and JWH-015 had lower levels of p24 production and expression, the HIV + JWH-015 treated MDDC had the lowest levels of p24 when compared to the HIV + THC treated cells. In addition, MDDC endocytic function and cytokine production were also differentially altered after alcohol and cannabinoid treatments. Our results show a differential effect of alcohol and cannabinoids, which may provide insights into the divergent inflammatory role of alcohol and cannabinoids to modulate MDDC function in the context of HIV infection