Aberrant populations of circulating T follicular helper cells and regulatory B cells underlying idiopathic pulmonary fibrosis

Abstract

Background: T follicular helper (Tfh) cells have been identified as a new category of helper T cells, which express CXCR5 on their surface and induce the production of antigen-specific antibodies. Many investigations have found morbid proliferation and/or activation of Tfh cells in systemic autoimmune and allergic diseases. It is also known that Tfh cells are regulated by regulatory B (Breg) cells in the deteriorating such diseases. Recently, CXCL13, a ligand of CXCR5, has been reported to increase in the peripheral blood and lungs of patients with idiopathic pulmonary fibrosis (IPF). This study aimed to investigate the involvement of Tfh cells and Breg cells in IPF.

Methods: Peripheral blood samples were obtained from 18 patients with IPF. We isolated heparinized peripheral blood mononuclear cells and investigated the proportions of Breg cells, Tfh cells, PD-1⁺ICOS⁺ Tfh cells (activated form of Tfh cells), and the Tfh-cell subsets by flow cytometry. These cell profiles were compared with those of 21 healthy controls. Furthermore, we investigated the correlations between profiles of lymphocytes and lung physiology.

Results: The median proportions of Tfh cells per total CD4⁺ T cells and of PD-1⁺ICOS⁺ proportion of Tfh cells per total Tfh cells was significantly more in the IPF patients (20.4 and 5.2%, respectively) compared with healthy controls (15.4 and 2.1%, respectively; p = 0.042 and p = 0.004, respectively). The proportion of Tfh2 cells per total Tfh cells was significantly higher and the proportion of Tfh17 was smaller in the IPF patients than healthy controls. The percentage of Breg cells to total B cells was significantly decreased in the IPF patients (median, 8.5%) compared with that in the controls (median, 19.7%; p < 0.001). The proportion of Breg cells was positively correlated with the annual relative change in diffusing capacity of the lungs for carbon monoxide in the IPF patients (r = 0.583, p = 0.018).

Conclusion: Proliferation and activation of Tfh cells and a decrease in Breg cells were observed in the peripheral blood of patients with IPF. The profile of the Tfh-cell subset also changed. Specific humoral immunity aberration would likely underlie complicated pathophysiology of IPF.

Keywords: CXCR5 (C-X-C motif chemokine receptor 5); ICOS (inducible co-stimulatory molecule); Idiopathic pulmonary fibrosis (IPF); PD-1 (programmed death 1); T follicular helper cell (Tfh cell); autoimmunity immunity; regulatory B cell (Breg cell).

Fig. 1

Ratios of circulating total Tfh cells in IPF and healthy cases. a Representative fluorescence-activated cell sorting profiles indicating total Tfh cells (CD3⁺CD4⁺CXCR5⁺). Plots were pregated on CD3⁺CD4⁺ cells and examined by the levels of CXCR5. The numbers indicate the proportion of cells in the gate. b The proportion of total Tfh cells in CD3⁺CD4⁺ cells is shown in the panel. Tfh, follicular helper T; IPF, idiopathic pulmonary fibrosis

Fig. 2

Ratios of circulating PD1⁺ICOS⁺Tfh cells in IPF and healthy cases. a Representative fluorescence-activated cell sorting profiles indicating PD-1⁺ICOS⁺Tfh cells. Plots were pregated on CD3⁺CD4⁺CXCR5⁺ cells and examined by the levels of PD-1 and ICOS. The numbers indicate the proportion of cells in the gate. b The proportion of PD-1⁺ICOS⁺Tfh cells in Tfh cells is shown in the panel. Tfh, follicular helper T; IPF, idiopathic pulmonary fibrosis

Fig. 3

Polarization of circulating Tfh-cell subsets in IPF and healthy cases. a Representative fluorescence-activated cell sorting profiles indicating Tfh1 cells (CXCR3⁺CCR6⁻), Tfh2 cells (CXCR3⁻CCR6⁻), and Tfh17 cells (CXCR3⁻CCR6⁺). Plots were pregated on CD3⁺CD4⁺CXCR5⁺ cells and examined by the levels of CXCR3 and CCR6. The numbers indicate the proportion of cells in the gate. b–d The proportions of Tfh-cell subsets among all Tfh cells are shown in the panel. b Tfh1 cells, (c) Tfh2 cells and (d) Tfh17 cells. Tfh, follicular helper T; IPF, idiopathic pulmonary fibrosis

Fig. 4

Proportions of circulating Breg cells in IPF and healthy cases. a Representative fluorescence-activated cell sorting profiles indicating Breg cells (CD24^hiCD27⁺). Plots were pregated on CD3⁻CD19⁺ cells and examined according to the levels of CD24 and CD27. The numbers indicate the proportion of cells in the gate. b The proportion of Breg cells among all CD3⁻CD19⁺ B cells is shown in the panel. Breg, regulatory B; IPF, idiopathic pulmonary fibrosis

Fig. 5

Correlation between the proportion of Breg cells and ΔDLCO in patients with idiopathic pulmonary fibrosis. ΔDLCO: means annual changes in DLCO. Breg: regulatory B; DLCO: diffusing capacity of the lung for carbon monoxide