Cigarette smoke exposure promotes differentiation of CD4⁺ T cells toward Th17 cells by CD40–CD40L costimulatory pathway in mice

Figures & data

Figure 1 The in vitro cultures of CD4⁺ T cells and BMDCs.

Notes: (1) the in vitro culture contained CD4⁺T cells only; (2) the in vitro culture contained CD4⁺T cells and CSE; (3) the in vitro culture contained CD4⁺T cell and BMDC without cigarette smoke exposure; (4) the in vitro culture contained CD4⁺T cell and BMDC with cigarette smoke exposure; (5) the in vitro culture contained CD4⁺T cell and BMDC without cigarette smoke exposure and CSE; (6) the in vitro culture contained CD4⁺T cell and BMDC with cigarette smoke exposure and CD40 antibody; (7) the in vitro culture contained CD4⁺T cell and BMDC without cigarette smoke exposure and CSE and CD40 antibody.
Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract.

Table 1 BMDCs proliferation exposed to varied concentrations of CSE(A_{490 nm}, $\overline{x}\pm s$)

Time (hours)	Control group	5 mL/L CSE	10 mL/L CSE	15 mL/L CSE	20 mL/L CSE
12	0.531±0.084	0.549±0.053	0.599±0.041	0.391±0.041a,b	0.274±0.034a,b
24	0.584±0.064	0.591±0.061	0.715±0.090a	0.428±0.036a,b	0.320±0.053a,b
48	0.621±0.041	0.619±0.070	0.693±0.057c	0.278±0.045a,b	0.133±0.038a,b
72	0.671±0.031	0.603±0.044	0.602±0.054a	0.148±0.014a,b	0.157±0.017a,b

Notes:

a P<0.01 vs control group.

b P<0.01 vs 10 mL/L CSE.

c P<0.05 vs control group.

Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract.

Figure 2 The number of CD4⁺IL-17⁺ T lymphocytes in lungs of the A group^a and the B group^b.

Notes: ^aThe A group in which mice were exempt from cigarette smoke exposure. ^bThe B group in which mice were exposed to cigarette smoke for 4 weeks. (A) Lymphocytes were marked with a circle in lung mononuclear cells by the flow cytometry. (B) The number of CD4⁺IL-17⁺ T lymphocytes in lungs of the A group. (C) The number of CD4⁺IL-17⁺ T lymphocytes in lungs of the B group.

Abbreviations: FSC, forward scatter; PE, phycoerythrin.

Figure 3 CD40 on surfaces of lung mononuclear cells of the A group^a and the B group^b.

Notes: ^aThe A group in which mice were exempt from cigarette smoke exposure. ^bThe B group in which mice were exposed to cigarette smoke for 4 weeks. (A) CD40 on lung mononuclear cells in the A group, red line as blank control. (B) CD40 on lung mononuclear cells in the B group, red line as blank control.
Abbreviation: PE, phycoerythrin.

Table 2 CD40 expression on the surfaces of BMDCs

Groups	CD40 expression
BMDCs from the A groupa	10.39±2.81
BMDCs from the B groupb	51.61±2.23c
BMDCs from the A group +10 mL/L CSEd	44.93±5.71c

Notes:

a The A group in which mice were exempt from cigarette smoke exposure.

b The B group in which mice were exposed to cigarette smoke for 4 weeks.

c P<0.01 vs the A group.

d BMDC from the control group and exposed to 10 mL/L CSE for 24 hours. Data presented as mean ± standard deviation.

Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract.

Figure 4 CD 40 expression on surfaces of BMDCs.

Notes: The A group in which mice were exempt from cigarette smoke exposure. The B group in which mice were exposed to cigarette smoke for 4 weeks. (A) CD40 on BMDCs in A group, red line as blank control. (B) CD40 on BMDCs in B group, red line as blank control. (C) CD40 on BMDCs in A group exposed to 10 mL/L CSE, red line as blank control.
Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract; PE, phycoerythrin.

Table 3 Th17 cell expressions in the in vitro cultures of CD4⁺ T cells and BMDCs

Group	CD4^+IL-17^+ T lymphocytes
CD4^+ T group	1.39±0.48
CD4^+ T + CSE group	3.85±0.73a
CD4^+ T + BMDC group	3.91±0.42a
CD4^+ T + smoke BMDC group	5.35±1.03b
CD4^+ T + smoke BMDC + CD40 antibody group	3.27±0.80c
CD4^+ T + BMDC + CSE group	7.78±0.76b
CD4^+ T + BMDC + CSE + CD40 antibody group	3.88±0.41d

Notes:

a P<0.01 vs the CD4⁺T group.

b P<0.01 vs the CD4⁺ T + BMDC group.

c P<0.01 vs the CD4⁺ T + smoke BMDC group.

d P<0.01 vs the CD4⁺ T + BMDC + CSE group. Data presented as mean ± standard deviation.

Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract.

Table 4 Concentrations of IL-17A and IL-27 in the in vitro cultures of CD4⁺ T cells and BMDCs

Group	IL-17A (pg/mL)	IL-27 (pg/mL)
CD4^+ T group	38.42±9.41	–
CD4^+ T + CSE group	93.83±14.71##	–
CD4^+ T + BMDC group	109.29±16.09##	8.91±1.98
CD4^+ T + smoke BMDC group	313.13±49.35*	34.92±5.03*
CD4^+ T + smoke BMDC + CD40 antibody group	174.24±35.21**	14.37±4.95**
CD4^+ T + BMDC + CSE group	329.36±45.18*	40.07±8.54*
CD4^+ T + BMDC + CSE + CD40 antibody group	141.18±29.27#	13.75±5.11#

Notes:

* P<0.01 vs the CD4⁺ T + BMDC group.

** P<0.01 vs the CD4⁺ T + smoke BMDC group.

# P<0.01 vs the CD4⁺ T + BMDC + CSE group.

## P<0.01 vs the CD4⁺ T group. “–” = value is too little to be quantified. Data presented as mean ± standard deviation.

Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract.

Figure 5 CD4⁺IL-17⁺ T lymphocytes in the seven in vitro cultures of CD4⁺ T cells and BMDCs.

Notes: (A) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cell group. (B) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cells + CSE group. (C) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cells + BMDCs group. (D) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cells + smoke BMDCs group. (E) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cells + smoke BMDCs + CD40 antibody group. (F) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺T cells + CSE group. (G) CD4⁺IL-17⁺ T lymphocytes in the CD4⁺ T cells + BMDCs + CSE + CD40 antibody group.
Abbreviations: BMDCs, bone marrow-derived dendritic cells; CSE, cigarette smoke extract; PE, phycoerythrin.