LIF upregulates expression of NK-1R in NHBE cells - PubMed (original) (raw)

LIF upregulates expression of NK-1R in NHBE cells

Cheng-Ping Hu et al. Mediators Inflamm. 2006.

Abstract

Leukemia inhibitory factor (LIF), a cytokine at the interface between neurobiology and immunology, is mainly mediated through JAK/STAT pathway and MAPK/ERK pathway. Evidence suggested LIF is related to the higher expression of neurokinin-1 receptor (NK-1R) in asthma. In this study, the immunohistochemistry stain showed the expressions of NK-1R, LIF, p-STAT3, and p-ERK1/2 in the lung tissues of allergic rats were increased compared with the controls, and the main positive cell type was airway epithelial cell. Normal human bronchial epithelial cells were treated with LIF in the presence or absence of AG490 (JAK2 inhibitor), PD98059 (MEK inhibitor), and the siRNA against STAT3. Western blot and RT-PCR indicated that LIF induced the expression of NK-1R, which was inhibited by the inhibitors mentioned above. No significant interaction was found between JAK/STAT pathway and MAPK/ERK pathway. In summary, bronchial epithelial cell changes in asthma are induced by LIF which promotes the expression of NK-1R, and JAK/STAT pathway and MAPK/ERK pathway may participate in this process.

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Figures

Figure 1

Figure 1

Expression of NK-1R, LIF, p-STAT3, and p-ERK1/2 in lung tissues of asthmatic rats (SABC × 200). Immunohistochemistry was performed on lung tissue of asthmatic rats. There were higher expressions for NK-1R ((a) control group, (b) asthmatic group) and LIF ((c) control group, (d) asthmatic group) in the asthmatic rats than those in the control rats, and the similar changes were observed for p-STAT3 ((e) control group, (f) asthmatic group) and p-ERK1/2 ((g) control group, (h) asthmatic group). The main positive cell type was airway epithelial cell.

Figure 2

Figure 2

Effects of AG-490, PD-98059, or PMA on LIF-induced activation of signal transduction and activation of transcription_(STAT3) and ERK1/2. Western blot was performed on NHBE cells that had been preincubated with or without AG-490, PD-98059, or PMA and then stimulated with LIF. (a) LIF induced activation of tyrosine phosphorylation of STAT3, and tyrosine phosphorylation of STAT3 was inhibited by AG-490, but not by PD-98059, and not affected by PMA. (b) LIF did not enhance the expression of total-STAT3, and its expression was not affected by AG-490, PD-98059, and PMA. (c) LIF induced activation of phosphorylation of ERK1/2, and ERK1/2 activation was inhibited by PD-98059, but not by AG-490; PMA increased the expression of p-ERK1/2 in NHBE cells, but there were no significant differences between the cells stimulated with LIF and the cells stimulated with LIF in the presence of PMA. (d) and (e) LIF did not enhance the expression of total-ERK1/2, furthermore, AG-490, PD-98059, and PMA also did not affect it. Experiments were repeated three times with similar results, and the data was expressed as the mean ratio (target/GAPDH) ± SD.

Figure 3

Figure 3

Effects of AG-490, PD-98059, PMA, or siRNA-1(STAT3) on LIF-induced expression of NK-1R detected by immunocytochemistry_(SABC × 200). Immunocytochemistry was performed on cells that had been preincubated with or without AG-490, PD-98059, PMA, or siRNA-1(STAT3) and then stimulated with LIF ((a) control, (b) PD-98059, (c) AG-490, (d) LIF, (e) PMA, (f) LIF + PD-95059, (g) LIF + PMA, (h) LIF + AG-490, (i) LIF + control siRNA, (j) LIF + sham plasmid, (k) LIF + siRNA-1 against STAT3). LIF induced expression of NK-1R, which was inhibited by AG-490, PD-98059, and siRNA-1 against STAT3, but was affected neither by the control siRNA nor the sham plasmid. Experiments were repeated three times with similar results, and the data was expressed as the mean ratio (positive cells number/total cells number) ± SD.

Figure 4

Figure 4

Effects of AG-490, PD-98059, PMA, or siRNA-1(STAT3) on LIF-induced expression of NK-1R detected by RT-PCR. RT-PCR was performed on cells that had been preincubated with or without AG-490, PD-98059, PMA, or siRNA-1(STAT3) and then stimulated with LIF. (a) LIF induced expression of NK-1R mRNA, and that was inhibited by AG-490 and PD-98059; PMA increased the expression of NK-1R mRNA in NHBE cells. (b) LIF-induced expression of NK-1R mRNA was inhibited by siRNA-1 against STAT3, but was affected neither by the control siRNA nor the sham plasmid. Experiments were repeated three times with similar results, and the data was expressed as the mean ratio_(target/β-actin) ± SD.

Figure 5

Figure 5

Effects of siRNA(STAT3) on LIF-induced activation of signal transduction and activation of transcription (STAT3) and ERK1/2. Western blot was performed on NHBE cells that had been preincubated with or without siRNA(STAT3) and then stimulated with LIF. (a) LIF induced activation of tyrosine phosphorylation of STAT3, and tyrosine phosphorylation of STAT3 was inhibited by siRNA-1 and siRNA-2, but neither by control siRNA nor sham plasmid. (b) Similiar to phosphorylation of STAT3, total-STAT3 was inhibited by siRNA-1 and siRNA-2, but neither by control siRNA nor sham plasmid. (c) LIF induced activation of phosphorylation of ERK1/2, but ERK1/2 activation was not inhibited by siRNA-1 against STAT3. (d) Similiar to phosphorylation of ERK1/2, the expression of total-ERK1/2 was not affected by siRNA-1 against STAT3. Experiments were repeated three times with similar results, and the data was expressed as the mean ratio(target/GAPDH) ± SD.

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