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Nicotinic receptor β2 determines NK cell-dependent metastasis in a murine model of metastatic lung cancer.

Hao J, Shi FD, Abdelwahab M, Shi SX, Simard A, Whiteaker P, Lukas R, Zhou Q - PLoS ONE (2013)

Bottom Line: Further, nicotinic suppression of NF-κB-induced transcriptional activity in NK cells is dependent on nAChR β2.This nAChR subtype also plays a large role in the NK cell-mediated control of melanoma lung metastasis, in a murine lung metastasis model exposed to nicotine.Our findings suggest nAChR β2 as a prominent pathway for nicotine induced impairment of NK cell functions which contributes to the occurrence of smoking-related pathologies.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Tianjin Neurological Institute, Tianjin Medical University General Hospital, Tianjin, China. jhao0216@gmail.com

ABSTRACT
Cigarette smoke exposure markedly compromises the ability of the immune system to protect against invading pathogens and tumorigenesis. Nicotine is a psychoactive component of tobacco products that acts as does the natural neurotransmitter, acetylcholine, on nicotinic receptors (nAChRs). Here we demonstrate that natural killer (NK) cells strongly express nAChR β2. Nicotine exposure impairs the ability of NK cells to kill target cells and release cytokines, a process that is largely abrogated by nAChR β2 deficiency. Further, nicotinic suppression of NF-κB-induced transcriptional activity in NK cells is dependent on nAChR β2. This nAChR subtype also plays a large role in the NK cell-mediated control of melanoma lung metastasis, in a murine lung metastasis model exposed to nicotine. Our findings suggest nAChR β2 as a prominent pathway for nicotine induced impairment of NK cell functions which contributes to the occurrence of smoking-related pathologies.

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Nicotine impairs NK cell proliferation and killing of target cells by acting on nAChR β2.NK cells were sorted from splenocyte single cell suspensions of wild type (β2+/+) or nAChR β2 knock-out (β2−/−). A. Cells were then cultured in the presence of various concentrations of nicotine (0.1, 1, 10 or 100 µM), and [3H]thymidine incorporation was measured (103 cpm + SEM; ordinate). B. NK cell numbers were assessed from β2+/+ or β2−/− mice in the presence or absence of nicotine. (N = 6–8 per group; Student’s t-test, * p<0.05). C. 51Cr-labbed target cell YAC-1 were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine, at the indicated effector/target cell ratio. Killing of target cells are measured by 51Cr release assay. D. B16 cells were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine. D-Luciferin was added to each well and the plate was imaged to obtain photons/s per cell. Wells with cells alone (no nicotine) or cells (added nicotine) were included as controls. The killing of B16 cells are measured via bioluminescence imaging. The data of one experiment out of two performed is shown, n = 3–4 mice/group. P values, Student’s t-test, *p<0.05. E. Production of IFN-γ was measured by intracellular cytokine staining. Dot plots are representative of two separate experiments (n = 6–18 mice). F. Production of IFN-γ, TNF-α, GM-CSF, MIP-1α and MIP-β by sorted NK cells was measured by ELISA. P values, Student’s t-test, *p<0.05.
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pone-0057495-g003: Nicotine impairs NK cell proliferation and killing of target cells by acting on nAChR β2.NK cells were sorted from splenocyte single cell suspensions of wild type (β2+/+) or nAChR β2 knock-out (β2−/−). A. Cells were then cultured in the presence of various concentrations of nicotine (0.1, 1, 10 or 100 µM), and [3H]thymidine incorporation was measured (103 cpm + SEM; ordinate). B. NK cell numbers were assessed from β2+/+ or β2−/− mice in the presence or absence of nicotine. (N = 6–8 per group; Student’s t-test, * p<0.05). C. 51Cr-labbed target cell YAC-1 were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine, at the indicated effector/target cell ratio. Killing of target cells are measured by 51Cr release assay. D. B16 cells were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine. D-Luciferin was added to each well and the plate was imaged to obtain photons/s per cell. Wells with cells alone (no nicotine) or cells (added nicotine) were included as controls. The killing of B16 cells are measured via bioluminescence imaging. The data of one experiment out of two performed is shown, n = 3–4 mice/group. P values, Student’s t-test, *p<0.05. E. Production of IFN-γ was measured by intracellular cytokine staining. Dot plots are representative of two separate experiments (n = 6–18 mice). F. Production of IFN-γ, TNF-α, GM-CSF, MIP-1α and MIP-β by sorted NK cells was measured by ELISA. P values, Student’s t-test, *p<0.05.

Mentions: To understand the biological consequences of altered NK cell receptor expression induced by nicotine exposure, we investigated the effects of nicotine on NK cell mediated killing of target cells, as well as release of cytokines by NK cells. NK cells normal represent 5–10% of peripheral blood mononuclear cells in human and 1–3% of monocytes in mouse spleen or lymph node. For NK cells to perform their functions, these cells must proliferate under pathological situations. Thus, we first assessed the influence of nicotine on NK cell proliferation and found that nicotine reduced NK cell proliferation and numbers (Figure 3A, B). nAChR β2 deficiency largely restored the capacity of NK cells to proliferate and numbers in the presence of nicotine (Figure 3A, B).


Nicotinic receptor β2 determines NK cell-dependent metastasis in a murine model of metastatic lung cancer.

Hao J, Shi FD, Abdelwahab M, Shi SX, Simard A, Whiteaker P, Lukas R, Zhou Q - PLoS ONE (2013)

Nicotine impairs NK cell proliferation and killing of target cells by acting on nAChR β2.NK cells were sorted from splenocyte single cell suspensions of wild type (β2+/+) or nAChR β2 knock-out (β2−/−). A. Cells were then cultured in the presence of various concentrations of nicotine (0.1, 1, 10 or 100 µM), and [3H]thymidine incorporation was measured (103 cpm + SEM; ordinate). B. NK cell numbers were assessed from β2+/+ or β2−/− mice in the presence or absence of nicotine. (N = 6–8 per group; Student’s t-test, * p<0.05). C. 51Cr-labbed target cell YAC-1 were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine, at the indicated effector/target cell ratio. Killing of target cells are measured by 51Cr release assay. D. B16 cells were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine. D-Luciferin was added to each well and the plate was imaged to obtain photons/s per cell. Wells with cells alone (no nicotine) or cells (added nicotine) were included as controls. The killing of B16 cells are measured via bioluminescence imaging. The data of one experiment out of two performed is shown, n = 3–4 mice/group. P values, Student’s t-test, *p<0.05. E. Production of IFN-γ was measured by intracellular cytokine staining. Dot plots are representative of two separate experiments (n = 6–18 mice). F. Production of IFN-γ, TNF-α, GM-CSF, MIP-1α and MIP-β by sorted NK cells was measured by ELISA. P values, Student’s t-test, *p<0.05.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585320&req=5

pone-0057495-g003: Nicotine impairs NK cell proliferation and killing of target cells by acting on nAChR β2.NK cells were sorted from splenocyte single cell suspensions of wild type (β2+/+) or nAChR β2 knock-out (β2−/−). A. Cells were then cultured in the presence of various concentrations of nicotine (0.1, 1, 10 or 100 µM), and [3H]thymidine incorporation was measured (103 cpm + SEM; ordinate). B. NK cell numbers were assessed from β2+/+ or β2−/− mice in the presence or absence of nicotine. (N = 6–8 per group; Student’s t-test, * p<0.05). C. 51Cr-labbed target cell YAC-1 were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine, at the indicated effector/target cell ratio. Killing of target cells are measured by 51Cr release assay. D. B16 cells were incubated with NK cells derived from β2+/+ or β2−/− mice in the presence or absence of nicotine. D-Luciferin was added to each well and the plate was imaged to obtain photons/s per cell. Wells with cells alone (no nicotine) or cells (added nicotine) were included as controls. The killing of B16 cells are measured via bioluminescence imaging. The data of one experiment out of two performed is shown, n = 3–4 mice/group. P values, Student’s t-test, *p<0.05. E. Production of IFN-γ was measured by intracellular cytokine staining. Dot plots are representative of two separate experiments (n = 6–18 mice). F. Production of IFN-γ, TNF-α, GM-CSF, MIP-1α and MIP-β by sorted NK cells was measured by ELISA. P values, Student’s t-test, *p<0.05.
Mentions: To understand the biological consequences of altered NK cell receptor expression induced by nicotine exposure, we investigated the effects of nicotine on NK cell mediated killing of target cells, as well as release of cytokines by NK cells. NK cells normal represent 5–10% of peripheral blood mononuclear cells in human and 1–3% of monocytes in mouse spleen or lymph node. For NK cells to perform their functions, these cells must proliferate under pathological situations. Thus, we first assessed the influence of nicotine on NK cell proliferation and found that nicotine reduced NK cell proliferation and numbers (Figure 3A, B). nAChR β2 deficiency largely restored the capacity of NK cells to proliferate and numbers in the presence of nicotine (Figure 3A, B).

Bottom Line: Further, nicotinic suppression of NF-κB-induced transcriptional activity in NK cells is dependent on nAChR β2.This nAChR subtype also plays a large role in the NK cell-mediated control of melanoma lung metastasis, in a murine lung metastasis model exposed to nicotine.Our findings suggest nAChR β2 as a prominent pathway for nicotine induced impairment of NK cell functions which contributes to the occurrence of smoking-related pathologies.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Tianjin Neurological Institute, Tianjin Medical University General Hospital, Tianjin, China. jhao0216@gmail.com

ABSTRACT
Cigarette smoke exposure markedly compromises the ability of the immune system to protect against invading pathogens and tumorigenesis. Nicotine is a psychoactive component of tobacco products that acts as does the natural neurotransmitter, acetylcholine, on nicotinic receptors (nAChRs). Here we demonstrate that natural killer (NK) cells strongly express nAChR β2. Nicotine exposure impairs the ability of NK cells to kill target cells and release cytokines, a process that is largely abrogated by nAChR β2 deficiency. Further, nicotinic suppression of NF-κB-induced transcriptional activity in NK cells is dependent on nAChR β2. This nAChR subtype also plays a large role in the NK cell-mediated control of melanoma lung metastasis, in a murine lung metastasis model exposed to nicotine. Our findings suggest nAChR β2 as a prominent pathway for nicotine induced impairment of NK cell functions which contributes to the occurrence of smoking-related pathologies.

Show MeSH
Related in: MedlinePlus