Abstract
Background
Primary biliary cholangitis (PBC) is a slowly progressing autoimmune liver disease characterized by immune destruction of the interlobular bile ducts. While the etiology of PBC is unknown, it is thought to involve an environmental trigger in a genetically susceptible individual. Among environmental factors studied to date, only the human betaretrovirus (HBRV) has been reproducibly detected in PBC patients’ biliary epithelium. Recent studies have shown that the majority of PBC patients as well as proportion of patients with AIH and cryptogenic cirrhosis have evidence of HBRV RNA and proviral integrations in their biliary epithelium
Aims
Previous studies suggest that PBC patiens make proinflammatory responses to HBRV proteins. Using overlapping peptides of the HBRV Gag and Env, we have found that 38% of PBC patients had memory CD8+ T-cell responses to HBRV Gag with intracellular IFN-γ and TNF-α production. Whereas only 7% of patients demonstrated proinflammtory responses by T-cells to HBRV Env stimulation in identical experiments. The finding of significant diminished proinflammatory T-cell response to HBRV Env versus HBRV Gag in PBC patients and our prior experience of finding diminished humoral responses to HBRV Env suggested a hypothesis that the HBRV Env may contain an immunosuppressive domain (ISD), a highly conserved sequence in transmembrane (TM) protein of retroviruses (such as HIV-1, MuLV and HERV-K) that induces immunoregulatory effect on the immune system
Methods
In order to determine whether HBRV Env contains an ISD, we used 85 overlapping 18-mer individual HBRV peptides corresponding to SU and TM proteins to stimulate healthy PBMC. We examined whether individual peptides induced IL-10, IL-4, and IL-6 secretion. PBMCs (3 × 105 cells/well) from a healthy donor were incubated with individual HBRV SU and TM peptides for 24 hrs prior to collection of the supernatants. IL-10, IL-4, and IL-6 levels were then measured by Mesoscale analysis
Results
We identified a single peptide homologous to other retroviral ISDs located in HBRV TM protein that lead to IL-10, IL-4, and IL-6 production. In order to validate the identification of the HBRV ISD we repeated the experiment using HBRV ISD sequences with a single AA difference. The mutations were selected based on the homology of the potential HBRV ISD with known conserved ISDs in other retroviruses. Accordingly, we were able to identify the functionally important AAs in the ISD within the HBRV TM that triggered IL-10, IL-4, and IL-6 production by selectively altering the AAs in the HBRV ISD sequence
Conclusions
These studies are important because they show how HBRV potentially avoids the immune attack to HBRV Env and tolerizes the host to viral infection. Further studies will be required to determine its other functional properties as well as the implications of this finding surrounding the pathophysiology underlying PBC
Funding Agencies
CIHR