Mucosal-Associated Invariant T Cell Deficiency in Chronic Obstructive Pulmonary Disease

<p>Mucosal-associated invariant T (MAIT) cells have been reported to play an important role in mucosal immunity. However, little is known about the roles of MAIT cells in chronic obstructive pulmonary disease (COPD). The aims of this study were to examine the levels of circulating MAIT cells and their subsets in COPD patients and to investigate the potential relationship between clinical parameters and MAIT cell levels. Forty-five COPD patients and 57 healthy control subjects were enrolled in the study. Circulating MAIT cells and their subset levels in the peripheral blood were measured by flow cytometry. Disease grades were classified according to the GOLD criteria for the assessment of severity of COPD. Circulating MAIT cell levels were found to be significantly reduced in COPD patients. In particular, this MAIT cell deficiency was more prominent in CD8+ and double-negative T cell subsets. Interestingly, elevated serum C-reactive protein level and reduced FEV₁/FVC ratio were associated with MAIT cell deficiency in COPD patients. Furthermore, the circulating MAIT levels were found to be significantly lower in patients with moderate to severe COPD than in patients with mild COPD. Our data shows that MAIT cells are numerically deficient in the peripheral blood of patients with COPD. In addition, this MAIT cell deficiency was found to reflect inflammatory activity and disease severity. These findings provide important information for monitoring the changes in MAIT cell levels and for predicting the prognosis during the disease course.</p

Increased level and interferon-γ production of circulating natural killer cells in patients with scrub typhus

<div><p>Background</p><p>Natural killer (NK) cells are essential immune cells against several pathogens. Not much is known regarding the roll of NK cells in <i>Orientia tsutsugamushi</i> infection. Thus, this study aims to determine the level, function, and clinical relevance of NK cells in patients with scrub typhus.</p><p>Methodology/Principal findings</p><p>This study enrolled fifty-six scrub typhus patients and 56 health controls (HCs). The patients were divided into subgroups according to their disease severity. A flow cytometry measured NK cell level and function in peripheral blood. Circulating NK cell levels and CD69 expressions were significantly increased in scrub typhus patients. Increased NK cell levels reflected disease severity. In scrub typhus patients, tests showed their NK cells produced higher amounts of interferon (IFN)-γ after stimulation with interleukin (IL)-12 and IL-18 relative to those of HCs. Meanwhile, between scrub typhus patients and HCs, the cytotoxicity and degranulation of NK cells against K562 were comparable. CD69 expressions were recovered to the normal levels in the remission phase.</p><p>Conclusions</p><p>This study shows that circulating NK cells are activated and numerically increased, and they produced more IFN-γ in scrub typhus patients.</p></div

Regression coefficients of NK cell percentages with respect to clinical and laboratory parameters in scrub typhus patients.

<p>Regression coefficients of NK cell percentages with respect to clinical and laboratory parameters in scrub typhus patients.</p

Changes in NK cell levels and CD 69 expression in scrub typhus patients.

<p>The percentages of NK cells (panel A) and CD69-expressing NK cells (panel B) in the peripheral blood of 11 scrub typhus patients during active disease and remission were determined by flow cytometry. Symbols represent individual subjects. *p < 0.005 by the Wilcoxon matched-pairs signed rank test.</p

Activation, Impaired Tumor Necrosis Factor-α Production, and Deficiency of Circulating Mucosal-Associated Invariant T Cells in Patients with Scrub Typhus

<div><p>Background</p><p>Mucosal-associated invariant T (MAIT) cells contribute to protection against certain microorganism infections. However, little is known about the role of MAIT cells in <i>Orientia tsutsugamushi</i> infection. Hence, the aims of this study were to examine the level and function of MAIT cells in patients with scrub typhus and to evaluate the clinical relevance of MAIT cell levels.</p><p>Methodology/Principal Findings</p><p>Thirty-eight patients with scrub typhus and 53 health control subjects were enrolled in the study. The patients were further divided into subgroups according to disease severity. MAIT cell level and function in the peripheral blood were measured by flow cytometry. Circulating MAIT cell levels were found to be significantly reduced in scrub typhus patients. MAIT cell deficiency reflects a variety of clinical conditions. In particular, MAT cell levels reflect disease severity. MAIT cells in scrub typhus patients displayed impaired tumor necrosis factor (TNF)-α production, which was restored during the remission phase. In addition, the impaired production of TNF-α by MAIT cells was associated with elevated CD69 expression.</p><p>Conclusions</p><p>This study shows that circulating MAIT cells are activated, numerically deficient, and functionally impaired in TNF-α production in patients with scrub typhus. These abnormalities possibly contribute to immune system dysregulation in scrub typhus infection.</p></div

Cytotoxicity of NK cells in scrub typhus patients.

<p>Panel A and B: NK cytotoxicity. Freshly isolated PBMCs (panel A) or purified NK cells (panel B) from 30 HCs and 20 patients with scrub typhus were cocultured with K562 cells for 4 hours, and then stained with FITC-conjugated anti-CD45 mAb and PI. Cytotoxicity was determined as the percentage of apoptotic K562 cells by flow cytometry. Panel C: CD107a expression in NK cells. PBMCs obtained from 15 HCs and 15 patients with scrub typhus patients were stained with FITC-conjugated anti-CD107a or isotype control mAbs and then incubated with K562 cells. After 1 hour, monensin was added and the cells were incubated for an additional 4 hours. The cells were then stained with PerCP-conjugated anti-CD3 and PE-conjugated anti-CD56 mAbs, and then CD107a expression in NK cells was analyzed by flow cytometry. Symbols represent individual subjects and horizontal lines indicate median values. ns = not significant by the ANCOVA test. Panel D: NK cell cytotoxicity of purified CD69- and CD69+ NK cell subsets in scrub typhus patients. Results are representative of 3 independent experiments.</p

Plasma levels of IFN-γ, IL-12, IL-17, IL-18 and TNF-α in patients with scrub typhus.

<p>Plasma samples of the patients were collected before specific treatment on admission. Plasma levels of IFN-γ (panel A), IL-17 (panel B), IL-12 (panel C), IL-18 (panel D), and TNF-α (panel E) were determined by Luminex and ELISA. Data were obtained from 15 age- and sex-matched HCs and 25 patients with scrub typhus. Symbols represent individual subjects and horizontal lines indicate median values. *p < 0.005, **p < 0.001, ***p < 0.0001 by the Mann-Whitney U test.</p

Clinical and laboratory characteristics of 38 scrub typhus patients.

<p>Clinical and laboratory characteristics of 38 scrub typhus patients.</p

Increased circulating NK cell numbers in the peripheral blood of scrub typhus patients.

<p>Freshly isolated PBMCs from 56 HCs and 56 patients with scrub typhus were stained with FITC-conjugated anti-CD56, PE-conjugated anti-CD3, and PerCP-conjugated anti-CD45 mAbs, and then analyzed by flow cytometry. Percentages of NK cells were calculated using a CD45/SSC gate. Panel A: Representative NK cell percentages as determined by flow cytometry. Panel B: NK cell percentages among peripheral blood lymphocytes. Panel C: Absolute NK cell numbers (per microliter of blood). Symbols represent individual subjects and horizontal lines indicate median values. Panel D: The ratios of CD3-CD56^bright/CD3-CD56^dim NK cell percentages among peripheral blood lymphocytes. Data are shown as box plots. Each box represents the 25th and 75th percentiles. The line inside the boxes represent the median. Whiskers represent the 10th and 90th percentiles. *p < 0.05, **p < 0.0005 by the ANCOVA test.</p

Expression of CD69 and PD-1 and apoptosis of MAIT cells after stimulation with IL-12 and IL-18.

<p>PBMCs (1 × 10⁶/well) were incubated for 24 hours in the presence of IL-12 (50 ng/mL) and IL-18 (50 ng/mL), and then stained with FITC-conjugated anti-CD3, FITC-conjugated annexin V, APC-conjugated anti-TCR Vα7.2, PE-conjugated anti-CD3, PE-conjugated anti-CD69, PE-conjugated anti-PD-1 and PE-Cy5-conjugated anti-CD161 monoclonal antibodies. Percentages of CD69-expressing cells (<u>panel A</u>), annexin V-positive cells (<u>panel C</u>), and PD-1-expressing cells (<u>panel E</u>) among MAIT cells were determined by flow cytometry. Data in <u>panels B, D and F</u> were obtained from 6 HCs. Values are expressed as the mean ± SEM. *p < 0.005 by paired t test.</p