Chronic morphine administration differentially modulates viral reservoirs in SIVmac251 infected rhesus macaque model

HIV persists in cellular reservoirs despite effective anti-retroviral therapy with rebound of viremia upon therapy interruption. Opioids modulate the immune system and suppress antiviral gene responses, which significantly impact people living with HIV (PLWH). However, the effects of opioids on viral reservoir remains elusive. Herein, we describe a morphine dependent SIVmac251 infected Rhesus macaque (RM) model to study the impact of opioids on HIV reservoirs. RMs were ramped up with morphine (n=10) or saline (n=9) for two weeks to a final dosage of 5mg/kg administered twice daily, which was maintained for seven weeks, and then infected with SIVmac251. Combined anti-retroviral therapy (cART) was initiated in approximately half the animals in each group five weeks post-infection and morphine/saline administration continued for 10 months. Among drug naïve macaques, there were no differences in plasma/CSF viral load nor in cell-associated DNA/RNA loads. However, within the cART-suppressed macaques, there was a reduction in cell-associated DNA load, intact proviral DNA copy numbers, and inducible SIV reservoir in both peripheral blood and lymph nodes (LNs) of morphine-administered RMs compared to saline controls. Further, PBMCs of morphine administered RMs, a reduction in Th1 polarized CD4+ T cells and in LNs there was a reduction in the total Tfh and Th1 like Tfh cells were observed, indicating probably have impact on reduction of viral reservoirs. In distinction to PBMC and LNs, within the CNS size of latent SIV reservoirs was higher in the CD11b+ microglia/macrophages of morphine-dependent RMs. These data suggest that morphine plays a role in modulating SIV reservoirs, reduces the CD4+ T-cell reservoir in both peripheral blood and LNs, and increases microglia/macrophage reservoirs in CNS. These findings will aid in understanding of molecular mechanism(s) of opioid-mediated differential modulation of viral reservoirs and evaluation of therapeutic strategies to reduce/eliminate HIV reservoirs in opioid-dependent PLWH. Author summary Opioids are commonly used as well as abused by HIV infected individuals, are known to suppress immune responses. However, their effects on modulating viral reservoir dynamics is not known. Here we developed a morphine dependent SIVmac251 infected rhesus macaque (RM) model to study the impact of opioids on HIV reservoirs and immune cells. We found that there was no difference in viral loads or cell-associated DNA/RNA loads among morphine dependent vs. saline treated control macaques. On the other hand, when macaques were treated with cART, there was a reduction in SIV reservoirs both in periphery and lymphoid tissues in morphine administered RMs, and the size of latent SIV reservoir was higher in the CNS CD11b+ microglia/macrophages as compared to control macaques. Therefore, these new data form macaque models suggest that PLWH who suffering from opioid use disorders have higher reservoirs in CNS as compared to lymphoid system. Thus, through understanding these reservoirs among PLWH who uses opioids are critical for better designing HIV cure strategies. ### Competing Interest Statement The authors have declared no competing interest. * BAL : Bronchoalveolar lavage BBB : Blood Brain Barrier cART : combined Antiretroviral Therapy CCL2 : C-C Motif Chemokine Ligand 2 CCL5 : C-C Motif Chemokine Ligand 5 CCR5 : C-C chemokine receptor type 5 CD4+ : T cells Cluster of differentiation 4+ T cells CNS : Central Nervous System CSF : cerebrospinal fluid ddPCR : Droplet Digital Polymerase Chain Reaction DMEM : Dulbecco’s Modified Eagle Medium DNA : Deoxyribonucleic acid DRV : Darunavir DTG : Dolutegravir EDTA : Ethylenediaminetetraacetic acid FBS : Fetal bovine serum FTC : Emtricitabin GALT : Gut Associated Lymphoid Tissues HIV-1 Human Immunodeficiency Virus - 1 HCV : Hepatitis C virus IACUC : Institutional Animal Care and Use Committee IBC : Institutional biosafety Committee ICAM : Intercellular adhesion molecule IL-6 : Interleukin 6 IPDA : Intact proviral DNA assay IUPM : Infectious units per million LN : Lymph Node MØ-QVOA : Macrophage Quantitative Viral Outgrowth Assay NHPs : Non-human primates NK : cells Natural killer cells OUD : Opioid use disorder PBMCs : Peripheral blood mononuclear cells PBS : Phosphate-buffered saline PLWH : People living with HIV PMA : Phorbol Myristate Acetate QVOA : Quantitative Viral Outgrowth Assay RM : Rhesus Macaque RNA : Ribonucleic acid RPM : Revolutions per minute RTV : Ritonavir SIV : Simian immunodeficiency virus SNP : Single Nucleotide Polymorphisms SUD : Substance use disorders TFV : Tenofovir alafenamide TILDA : Tat/rev Induced Limiting Dilution Assay TLR4 : Toll-like receptor 4 TNF-α : Tumor necrosis factor alpha VCAM : Vascular cell adhesion protein β-ME : 2-Mercaptoethanol