Malin H. Meyer-Myklestad (1,2,3), Mari Kaarboe (1), Mingyi Yang (1), Birgitte Stiksrud (2), Anne Ma Dyrhol-Riise (2,3), Dag Kvale (2,3), Marius Trøseid (3,4,5), Pål Aukrust (3,5), Magnar Bjørås (1,6) and Dag Henrik Reikvam (2,3)

Affiliates

(1) Dep. of Microbiology, Oslo University Hospital, Oslo, Norway. (2) Dep. of Infectious diseases, Oslo University Hospital, Oslo, Norway. (3) Institute for Clinical Medicine, University of Oslo, Oslo, Norway. (4) Section of Clinical Immunology and Infectious diseases, Oslo University Hospital Rikshospitalet, Norway (5) Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway. (6) Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology, Trondheim, Norway.

Abstract

Background: Approximately 15% of people living with HIV (PLHIV) on antiretroviral therapy (ART) and persistent viral suppression fail to restore CD4+ T cell levels. These immunological nonresponders (INR) have increased risk of non-AIDS-related morbidity and mortality. The etiology of the INR phenotype remains unknown. We have recently shown that compared to immunological responders (IR) and HIV negative controls, INR have signs of increased enterocyte damage and gut mucosal immune dysfunction restricted to colon.
Methods: We performed mRNA sequencing and global proteome analyses by mass spectrometry (MS) of gut mucosal biopsies from sigmoid colon and terminal ileum of INR (ART>4 years with HIV RNA <50 copies/ml and CD4 count 3.5 years), IR (ART>4 years with HIV RNA 600 cells/µL for >3.5 years) matched on nadir CD4 count and age (n=5 of each group). Differential expression analyses were performed using negative binomial GLM fitting and Wald test for RNA-sequencing and empirical Bayes statistics test for analyses of proteomics MS data. Targeted differentially expressed genes were assessed by quantitative PCR.
Results: In the sigmoid colon, approximately 3300 mRNA transcripts were significantly differentially expressed in INR compared to IR. In contrast, no differential expression was observed between INR and IR in terminal ileum. We detected about 3700 proteins by global proteomic analyses that were evaluated for differential expression. Consistently, global proteomic analyses showed a higher level of differential regulation between INR and IR in colon than in terminal ileum (12 versus 4). In the colon, the protein periostin, involved in cell survival, and Musashi RNA binding protein 2 (MSI2), involved in cell cycle regulation, were both significantly downregulated in INR compared to IR. The apoptotic factor protein CASP3 was upregulated in colon of INR compared to IR. There was no differential regulation of these proteins in the terminal ileum.
Conclusion: Sigmoid colon, as opposed to terminal ileum, is implicated as an anatomic site linked to mechanisms causing complete immune recovery in PLHIV. The differentially regulated genes and proteins identified in sigmoid colon may contribute to the INR phenotype through reduced cell survival, cell cycle regulation and increased apoptosis. These factors may be candidates for adjuvant therapy to improve the prognosis and quality of life for PLHIV.