Demographic, clinical and laboratory characteristics of study participants.

<p>Footnotes: Statistical analyses performed using (A) Fisher’s Exact test and (B) Kruskal-Wallis non-parametric ANOVA test:</p><p>*p<0.05</p><p>**p<0.01</p><p>***p<0.001</p><p>CPTN: HIV/TB co-infection treatment naïve; CPTP: HIV/TB co-infection treatment positive; HVTN: HIV mono-infection treatment naïve; HVTP: HIV mono-infection treatment positive; NA: non- applicable. All values are expressed as mean ±SD.</p><p>Demographic, clinical and laboratory characteristics of study participants.</p

Expression levels of different markers by CD161⁺⁺CD8⁺ T-cell subsets in the study population.

<p><b>(A)</b> Zebra plots of double-gating strategy (gated on the CD161⁺⁺ CD8⁺ T cells) show staining with 4 different markers (CD103, PD-1, CCR6, CCR5) on representative samples from a HC and a CPTN. HCs showed increased amount of CCR6-expressing MAIT cells and decreased amount of PD-1 expressing MAIT cells compared to CPTNs. <b>(B)</b> HC showed significantly lower expression level of inhibitory receptor, PD-1 while HIV/TB co-infected patients show significantly increased PD-1 expressing MAIT cells. <b>(C)</b> Significantly increased CCR6-expressing MAIT cells were found in HCs compared to HIV and TB infected groups. <b>(D)</b> No significant difference was observed in CCR5 expression levels by MAIT cells among the different study subjects. The significant difference in CCR5 expression between HVTPs and HCs may be due to the limited number of samples. All graphs show median (red bars) and range (blue whiskers); <i>P</i> values are reported for two-sided Mann-Whitney tests with threshold for significance <i>P</i> = 0.025 after Bonferroni correction for 2 comparisons. (Note: TN, treatment naïve; TP, treatment positive; HC, healthy control; CP, HIV/TB co-infection; HV, HIV mono-infection).</p

Percentage of CD8⁺ T cells expressing CD161⁺⁺ across different study groups.

<p><b>(A)</b> Scatter plots (gated on the CD3⁺ T-cell population) show co-staining with CD8 and CD161 on representative samples from 5 different clinical groups: CPTNs, CPTPs, HVTNs, HVTPs, and HCs. <b>(B)</b> CD161⁺⁺CD8⁺ T (MAIT cell) frequency in HCs showed significantly increased MAIT cell levels compared to other study groups. <b>(C)</b> CD161⁺CD8⁺ T cell frequency showed no difference across the different study groups. <b>(D-E)</b> CD161⁺⁺CD8⁺ MAIT cell frequency in subjects with HIV mono-infection and HIV/TB co-infection shows no significant correlation with either HIV plasma viral load (copies/mL) or CD4⁺ T-cell counts (cells/mm³). All graphs show median (red bars) and range (blue whiskers); <i>P</i> values are reported for two-sided Mann-Whitney tests with threshold for significance <i>P</i> = 0.025 after Bonferroni correction for 2 comparisons. Correlations between MAIT cell frequency and markers of HIV disease progression were assessed using two-tailed non-parametric Spearman’s rank. (Note: TN, treatment naïve; TP, treatment positive; HC, healthy control; CP, HIV/TB co-infection; HV, HIV mono-infection).</p

Profile of expression of CD161 on MAIT cells.

<p><b>(A)</b> The zebra plots depict the gating strategy for the analysis of expression of CD161 on MAIT cells. CD8⁺ T cells were gated for TCR Vα7.2 specific for MAIT cells. After gating on CD8⁺ TCR Vα7.2⁺ MAIT cell population, CD161 expression was subsequently analyzed. <b>(B-C)</b> MAIT cells of HCs showed significant increase of CD161 compared to other infected groups. Conversely, HCs has the lowest amount of CD161^- MAIT cells. All graphs show median (red bars) and range (blue whiskers); <i>P</i> values are reported for two-sided Mann-Whitney tests with threshold for significance <i>P</i> = 0.025 after Bonferroni correction for 2 comparisons. (Note: TN, treatment naïve; TP, treatment positive; HC, healthy control; CP, HIV/TB co-infection; HV, HIV mono-infection).</p

Image_1.tif

<p>Mucosal-associated invariant T (MAIT) cells, defined as CD161⁺⁺TCR iVα7.2⁺ T cells, play an important role in the innate defense against bacterial infections, and their functionality is impaired in chronic viral infections. Here, we investigated the frequency and functional role of MAIT cells in chronic hepatitis B virus (HBV) infection. The peripheral CD3⁺CD161⁺⁺TCR iVα7.2⁺ MAIT cells in chronic HBV-infected patients and healthy controls were phenotypically characterized based on CD57, PD-1, TIM-3, and CTLA-4, as well as HLA-DR and CD38 expression. The frequency of MAIT cells was significantly decreased among chronic HBV-infected individuals as compared to controls. Expression of CD57, PD-1, CTLA-4, as well as HLA-DR and CD38 on MAIT cells was significantly elevated in chronic HBV-infected individuals relative to controls. The percentage of T cell receptor (TCR) iVα7.2⁺ CD161⁺ MAIT cells did not correlate with HBV viral load but inversely with HLA-DR on CD4⁺ T cells and MAIT cells and with CD57 on CD8⁺ T cells suggesting that decrease of MAIT cells may not be attributed to direct infection by HBV but driven by HBV-induced chronic immune activation. The percentage and expression levels of PD-1 as well as CTLA-4 on MAIT cells inversely correlated with plasma HBV-DNA levels, which may suggest either a role for MAIT cells in the control of HBV infection or the effect of HBV replication in the liver on MAIT cell phenotype. We report that decrease of TCR iVα7.2⁺ MAIT cells in the peripheral blood and their functions were seemingly impaired in chronic HBV-infected patients likely because of the increased expression of PD-1.</p

Image_2.tif

<p>Mucosal-associated invariant T (MAIT) cells, defined as CD161⁺⁺TCR iVα7.2⁺ T cells, play an important role in the innate defense against bacterial infections, and their functionality is impaired in chronic viral infections. Here, we investigated the frequency and functional role of MAIT cells in chronic hepatitis B virus (HBV) infection. The peripheral CD3⁺CD161⁺⁺TCR iVα7.2⁺ MAIT cells in chronic HBV-infected patients and healthy controls were phenotypically characterized based on CD57, PD-1, TIM-3, and CTLA-4, as well as HLA-DR and CD38 expression. The frequency of MAIT cells was significantly decreased among chronic HBV-infected individuals as compared to controls. Expression of CD57, PD-1, CTLA-4, as well as HLA-DR and CD38 on MAIT cells was significantly elevated in chronic HBV-infected individuals relative to controls. The percentage of T cell receptor (TCR) iVα7.2⁺ CD161⁺ MAIT cells did not correlate with HBV viral load but inversely with HLA-DR on CD4⁺ T cells and MAIT cells and with CD57 on CD8⁺ T cells suggesting that decrease of MAIT cells may not be attributed to direct infection by HBV but driven by HBV-induced chronic immune activation. The percentage and expression levels of PD-1 as well as CTLA-4 on MAIT cells inversely correlated with plasma HBV-DNA levels, which may suggest either a role for MAIT cells in the control of HBV infection or the effect of HBV replication in the liver on MAIT cell phenotype. We report that decrease of TCR iVα7.2⁺ MAIT cells in the peripheral blood and their functions were seemingly impaired in chronic HBV-infected patients likely because of the increased expression of PD-1.</p