Single cell RNA transcriptome response to fentanyl use in persons with hepatitis C virus infection

Krishna Roskin¹Heidi L Meeds²Janani Madhuravasal Krishnan²Matthew Juhascik³John M Cafardi⁴Jennifer L Brown⁵Caroline Freiermuth⁶Michael S Lyons⁷Kenneth E Sherman²Jason Blackard^8*

• ¹Division of Biomedical Informatics, Cincinnati Children’s Hospital Medical Center, Cincinnati, Ohio, United States
• ²Division of Digestive Diseases, Department of Internal Medicine, College of Medicine, University of Cincinnati, Cincinnati, Ohio, United States
• ³Miami Valley Regional Crime Laboratory / Montgomery County Coroner's Office, Dayton, United States
• ⁴Christ Hospital, Cincinnati, Ohio, United States
• ⁵Department of Psychological Sciences, College of Health and Human Sciences, Purdue University, West Lafayette, Indiana, United States
• ⁶Department of Emergency Medicine, College of Medicine, University of Cincinnati, Cincinnati, Ohio, United States
• ⁷Department of Emergency Medicine, College of Medicine, The Ohio State University, Columbus, Ohio, United States
• ⁸College of Medicine, University of Cincinnati, Cincinnati, United States

The US has experienced a major drug epidemic in recent years, attributed in large part to synthetic opioids such as fentanyl. Here, we evaluated how recreational / non-prescribed fentanyl use in persons living with HCV infection impacted gene expression profiles in the peripheral blood. Whole blood was collected from 11 individuals, including 4 HCV-negative healthy controls, 4 HCV-positive individuals with current fentanyl use, and 3 HCV-positive individuals with no current fentanyl / no opioid use. The median HCV RNA level was 5.6 log10 copies/mL. Cell frequencies were not different by fentanyl status except for non-CD4+, non-CD8+ T cells (higher for fentanyl use; p = 0.052). When comparing HCV-positive persons with / without fentanyl detected in their blood, 106 differentially expressed genes (DEGs) were identified, including 11 in CD4+ T lymphocytes, 46 in CD8+ T lymphocytes, 5 in monocytes, 13 in B lymphocytes (excluding plasmablasts), 24 in plasmablasts, 2 in dendritic cells, and 13 in NK cells. Seven DEGs – DHRS4L2, GZMA, H1-3, HLA-C, ISG15, PARP8, PRKX – were shared across multiple cell types, with the majority being involved in host defenses against viruses. Enrichment analysis of differentially expressed genes identified genes involved in multiple cellular processes and phenotypes. Expression of the HCV entry factor CD81 was high in PBMCs; however, other HCV entry factors were expressed at low levels, and none were differentially expressed in HCV-positive persons with / without fentanyl detected in their blood. These results highlight multiple pathways by which commonly abused opioids may affect HCV pathogenesis and may reveal additional pathways for novel target-specific therapeutic interventions and enhance the clinical management of this difficult-to-treat population.