LIF Up-regulates NK-1R Expression In NHBE Cells Via STAT Pathway And MAPK Pathway

Background: leukemia inhibitory factor(LIF) has been implicated invarious processes of neuronal development, differentiation, survival andneurogenesis, and it was indicated that LIF could increase the expressionof substance P and its receptor(neurokinin-1 receptor, NK-1R). Highlevels of LIF were found in atopic patients and asthmatic rats, and LIFmay be an important signal molecule in the airway response toinflammation. As a kind of cytokines, the biological effects of LIFdepend on signal transduction pathway such as janus kinase/signaltransducer and activator of transcription (JAK/STAT) pathway and theras-mitogen activated protein kinase (MAPK) pathway. It is indicated thatboth JAK/STAT pathway and MAPK pathway are related to asthma.Bronchial epithelial cell is a barrier to airway structure, and it is animportant target cell type in most respiratory diseases such as asthma.High levels of LIF and NK-1R were observed in bronchial epithelial cellsof asthmatic rats, so the bronchial epithelial cell is considered as thetarget cell to explore the signal transduction mechanism of NK-1Rregulation by LIF in this study.Chapter One LIF up-regulates NK-1R expression in lungs ofasthmatic rats via JAK/STAT pathway and MAPK pathway.Objective: To explore the regulation mechanism between LIF andNK-1R in asthma through comparing the expression of NK-1R, LIF,p-STAT3 and p-ERK1/2 between asthmatic rats and asthmatic rats treatedwith anti-LIF.Methods: 30 Sprague-Dawley rats were divided into 3 groups atrandom(asthmatic group, anti-LIF group and control group, n=10), andthey were housed under specific pathogen-free conditions. Sensitizationand provocation(the asthmatic group and the anti-LIF group) wereproduced with OVA, in this process, the anti-LIF group were treated withanti-LIF. Then the rats were killed and lung tissues were fixed in 4%polyoxymethylene, embedded in paraffin, and finally sliced into sections. Immunoreactivity for p-STAT3, p-ERK1/2, NK-1R and LIF proteins weredetected in the lungs of these rats.Results: Compared with the control, inflammatory cell infiltration werefound around airway in asthmatic rats, in addition, alveolar septum andbronch-wall were thickened in asthmatic rats. Compared with theasthmatic, the extent of inflammatory cells infiltration was relieved in therats treated with anti-LIF, and the pathological changes of bronch-wallwere also improved. Immunohistochemistry indicated a higher expressionof LIF in the asthmatic rats compared to that in the control group.Consistent with that, similar changes were observed for NK-1R, p-STAT3and p-ERK1/2. The main positive cell type was airway epithelial cell, andother positive types were also observed such as lymphocyte.Conclusions: NK-1R expression in lungs of asthmatic rats may beregulated by LIF, and it is possible that the effect of LIF is mediated byJAK/STAT pathway and MAPK pathway.Chapter Two LIF up-regulates NK-1R expression in NHBE cellsvia JAK/STAT pathway and MAPK pathway.Objective: To investigate whether LIF induces the expression of NK-1Rin bronchial epithelial cells and whether JAK/STAT pathway andMAPK/ERK pathway participate in this process.Methods: we treated normal human bronchial epithelial (NHBE) cellswith LIF in the presence or absence of AG490(JAK2 inhibitor),PD98059(ERK inhibitor), PMA(the activator of protein kinase C) and thesiRNA against STAT3. Then the expression of NK-1R, p-STAT3,total-STAT3, p-ERK1/2 and total-ERK1/2 in NHBE cells were detectedby Western-blot, RT-PCR or immunocytochemistry.Results: LIF induced activation of p-STAT3, and p-STAT3 expressionwas inhibited by AG-490 but not by PD-98059. Nevertheless, theexpression of total-STAT3 was not affected by the factors mentionedabove. LIF induced activation of p-ERK1/2, and p-ERK1/2 was inhibited by PD-98059 but not by AG-490. Similar to total-STAT3, the expressionof total-ERK1/2 did not change. PMA increased the expression ofp-ERK1/2 in NHBE cells, but there were no significant differencebetween the cells stimulated with LIF and the cells stimulated with LIF inthe presence of PMA. LIF induced expression of NK-1R in NHBE cells,both AG-490 and PD-98059 suppressed the LIF-induced expression ofNK-1R; on the contrary, PMA increased the expression of NK-1R inNHBE cells. LIF-induced p-STAT3 was inhibited by STAT3-siRNA, inaddition, the expression of total-STAT3 was also inhibited bySTAT3-siRNA. However, STAT3-siRNA did not affect the expression ofp-ERK1/2 and total-ERK1/2. The LIF-induced expression of NK-1R wasinhibited by STAT3-siRNA both at the mRNA level and the pretein level,but it was not affected by negative control siRNA and sham plasmid.Conclusions: LIF up-regulates NK-1R expression in NHBE cells viaJAK2/STAT3 pathway and MAPK pathway, in this process, noobservable interaction was found between the two pathways.