Morbidly obese human subjects have increased peripheral blood CD4+ T cells with skewing toward a Treg- and Th2-dominated phenotype

Kim van der Weerd, Willem A Dik, Benjamin Schrijver, Dave H Schweitzer, Anton W Langerak, Hemmo A Drexhage, Rosalie M Kiewiet, Maarten O van Aken, Astrid van Huisstede, Jacques J M van Dongen, Aart-Jan van der Lelij, Frank J T Staal, P Martin van Hagen, Kim van der Weerd, Willem A Dik, Benjamin Schrijver, Dave H Schweitzer, Anton W Langerak, Hemmo A Drexhage, Rosalie M Kiewiet, Maarten O van Aken, Astrid van Huisstede, Jacques J M van Dongen, Aart-Jan van der Lelij, Frank J T Staal, P Martin van Hagen

Abstract

Obesity is associated with local T-cell abnormalities in adipose tissue. Systemic obesity-related abnormalities in the peripheral blood T-cell compartment are not well defined. In this study, we investigated the peripheral blood T-cell compartment of morbidly obese and lean subjects. We determined all major T-cell subpopulations via six-color flow cytometry, including CD8+ and CD4+ T cells, CD4+ T-helper (Th) subpopulations, and natural CD4+CD25+FoxP3+ T-regulatory (Treg) cells. Moreover, molecular analyses to assess thymic output, T-cell proliferation (T-cell receptor excision circle analysis), and T-cell receptor-β (TCRB) repertoire (GeneScan analysis) were performed. In addition, we determined plasma levels of proinflammatory cytokines and cytokines associated with Th subpopulations and T-cell proliferation. Morbidly obese subjects had a selective increase in peripheral blood CD4+ naive, memory, natural CD4+CD25+FoxP3+ Treg, and Th2 T cells, whereas CD8+ T cells were normal. CD4+ and CD8+ T-cell proliferation was increased, whereas the TCRB repertoire was not significantly altered. Plasma levels of cytokines CCL5 and IL-7 were elevated. CD4+ T-cell numbers correlated positively with fasting insulin levels. The peripheral blood T-cell compartment of morbidly obese subjects is characterized by increased homeostatic T-cell proliferation to which cytokines IL-7 and CCL5, among others, might contribute. This is associated with increased CD4+ T cells, with skewing toward a Treg- and Th2-dominated phenotype, suggesting a more anti-inflammatory set point.

Figures

FIG. 1.
FIG. 1.
Absolute counts of leukocyte subpopulations (A), PBMC subpopulations (B), and CD4+ and CD8+ T cells (C) in peripheral blood of morbidly obese and lean subjects. CD4, CD4+ T cells; CD8, CD8+ T cells. White bars represent lean subjects (n = 11, BMI 23.2 ± 1.4 kg/m2); gray bars represent morbidly obese subjects (n = 8, BMI 42.4 ± 6.7 kg/m2). *P < 0.05.
FIG. 2.
FIG. 2.
A: Absolute counts of CD4+ and CD8+ T-cell subpopulations in peripheral blood of morbidly obese and lean subjects. B: Absolute counts of natural CD4+CD25+FoxP3+ Treg cells, IFN-γ–producing (Th1) cells, IL-4–producing (Th2) cells, and IL-17A–producing (Th17) cells in peripheral blood of morbidly obese and lean subjects within the CD4+ T-cell gate. TN, naive T cells; TCM, central memory T cells; TEM, effector memory T cells; TTD, terminally differentiated T cells. White bars represent lean subjects (n = 11, BMI 23.2 ± 1.4 kg/m2); gray bars represent morbidly obese subjects (n = 8, BMI 42.4 ± 6.7 kg/m2). *P < 0.05, †P < 0.01.
FIG. 3.
FIG. 3.
A: sjTREC content in total αβ+-T cells and T-cell subpopulations of morbidly obese and lean subjects. αβ+ T, αβ+-T cells; CD4 N, CD4+ naive T cells; CD4 M, CD4+ memory T cells; CD8 N, CD8+ naive T cells; CD8 M, CD8+ memory T cells; ND, not detectable. B: Correlation between sjTREC content and BMI (left) and age (right) in morbidly obese and lean subjects. White bars or dots represent lean subjects, αβ+-T cells (n = 10, BMI 24.0 ± 2.2 kg/m2), T-cell subpopulations (n = 4, BMI 23.6 ± 1.0 kg/m2); gray bars or dots represent morbidly obese subjects, αβ+-T cells (n = 8, BMI 42.4 ± 6.7 kg/m2), T-cell subpopulations (n = 5, BMI 41.6 ± 5.1 kg/m2). *P < 0.05.
FIG. 4.
FIG. 4.
GeneScan analysis of Vβ–Jβ rearrangements in CD4+ and CD8+ naive and memory T-cell subpopulations in a representative lean (A) and morbidly obese (B) subject. Tube A: Vβ+Jβ1.1 to Jβ1.6+Jβ2.2+Jβ2.6+Jβ2.7; Tube B: Vβ+Jβ2.3+Jβ2.4+Jβ2.5. Primers for the Jβ1 cluster were HEX-labeled (green line); primers for the Jβ2 cluster were FAM-labeled (blue line). CD4 N, CD4+ naive T cells; CD4 M, CD4+ memory T cells; CD8 N, CD8+ naive T cells; CD8 M, CD8+ memory T cells. Lean subjects (n = 4, BMI 23.6 ± 1.0 kg/m2); morbidly obese subjects (n = 5, BMI 41.6 ± 5.1 kg/m2).
FIG. 5.
FIG. 5.
Plasma levels of IL-6 and TNF-α (A); Th1 cytokines IL-12p70 and IFN-γ, Th2 cytokines IL-4 and IL-10, and the Th17 cytokine IL-17A (B); and IL-2, IL-7, and CCL5 in morbidly obese and lean subjects (C). D: Correlation between IL-7 plasma levels and CD4+ or CD8+ T-cell counts in peripheral blood. E: Correlation between CCL5 plasma levels and CD4+ or CD8+ T-cell numbers in peripheral blood. White dots represent lean subjects (n = 11, BMI 23.2 ± 1.4 kg/m2); gray dots represent morbidly obese subjects (n = 8, BMI 42.4 ± 6.7 kg/m2). *P < 0.05.
FIG. 6.
FIG. 6.
Correlation between CD4+ T-cell count in peripheral blood and fasting insulin levels (A) and insulin sensitivity (fasting glucose–to–fasting insulin ratio) (B). Correlation between CD8+ T-cell count in peripheral blood and fasting insulin levels (C) and insulin sensitivity (fasting glucose–to–fasting insulin ratio) (D). Gray dots represent morbidly obese subjects (n = 8, BMI 42.4 ± 6.7 kg/m2).

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