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Interleukin-17A promotes MUC5AC expression and goblet cell hyperplasia in nasal polyps via the Act1-mediated pathway.

Xia W, Bai J, Wu X, Wei Y, Feng S, Li L, Zhang J, Xiong G, Fan Y, Shi J, Li H - PLoS ONE (2014)

Bottom Line: IL-17A significantly stimulated the expression of IL-17RA, IL-17RC, act1 and MUC5AC, and the activation of the MAPK pathway in cultured PECs and NCI-H292 cells (p<0.05).In addition, IL-17RA, IL-17RC and act1 siRNA significantly blocked IL-17A-induced MUC5AC production in vitro (p<0.05).Our results suggest that IL-17A plays a crucial role in stimulating the production of MUC5AC and goblet cell hyperplasia through the act1-mediated signaling pathway and may suggest a promising strategy for the management of Th17-dominant NP patients.

View Article: PubMed Central - PubMed

Affiliation: Allergy and Cancer Center, Otorhinolarygology Hospital, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China; Department of Otolaryngology, Head and Neck Surgery, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.

ABSTRACT

Background: Recent studies demonstrated that nasal polyps (NP) patients in China and other Asian regions possessed distinct Th17-dominant inflammation and enhanced tissue remodeling. However, the mechanism underlying these observations is not fully understood. This study sought to evaluate the association of interleukin (IL)-17A with MUC5AC expression and goblet cell hyperplasia in Chinese NP patients and to characterize the signaling pathway underlying IL-17A-induced MUC5AC expression in vitro.

Method: We enrolled 25 NP patients and 22 normal controls and examined the expression of IL-17A, MUC5AC and act1 in polyp tissues by immunohistochemical (IHC) staining, quantitative polymerase chain reaction (qPCR) and western blot. Moreover, by using an in vitro culture system of polyp epithelial cells (PECs), IL-17A-induced gene expression was screened in cultured PECs by DNA microarray. The expression of IL-17RA, IL-17RC, act1 and MUC5AC and the activation of the MAPK pathway (ERK, p38 and JNK), were further examined in cultured PECs and NCI-H292 cells by qPCR and western blotting, respectively.

Results: We found that increased IL-17A production was significantly correlated with MUC5AC and act1 expression and goblet cell hyperplasia in polyp tissues (p<0.05). IL-17A significantly stimulated the expression of IL-17RA, IL-17RC, act1 and MUC5AC, and the activation of the MAPK pathway in cultured PECs and NCI-H292 cells (p<0.05). In addition, IL-17RA, IL-17RC and act1 siRNA significantly blocked IL-17A-induced MUC5AC production in vitro (p<0.05).

Conclusion: Our results suggest that IL-17A plays a crucial role in stimulating the production of MUC5AC and goblet cell hyperplasia through the act1-mediated signaling pathway and may suggest a promising strategy for the management of Th17-dominant NP patients.

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MAPK signaling mediated IL-17A induced MUC5AC in PECs and NCI-H292 cells in vitro.(A) Representative western blot result of phosphorylated p38, ERK and JNK in PECs after IL-17A stimulation. (B) Representative western blot result of phosphorylated p38, ERK and JNK in NCI-H292 cells after IL-17A stimulation. (C, D) MUC5AC protein level in cultured PECs and NCI-H292 cells after IL-17A stimulation for 24 h in the presence of specific inhibitors of p38, ERK and JNK. The data are expressed the means (SEM) of 3 independent experiments. * p<0.05 when compared with control PECs, #p<0.05 when compared with control NCI-H292 cells.
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pone-0098915-g005: MAPK signaling mediated IL-17A induced MUC5AC in PECs and NCI-H292 cells in vitro.(A) Representative western blot result of phosphorylated p38, ERK and JNK in PECs after IL-17A stimulation. (B) Representative western blot result of phosphorylated p38, ERK and JNK in NCI-H292 cells after IL-17A stimulation. (C, D) MUC5AC protein level in cultured PECs and NCI-H292 cells after IL-17A stimulation for 24 h in the presence of specific inhibitors of p38, ERK and JNK. The data are expressed the means (SEM) of 3 independent experiments. * p<0.05 when compared with control PECs, #p<0.05 when compared with control NCI-H292 cells.

Mentions: We next examined the importance of the MAPK pathway in IL-17A-induced MUC5AC expression in cultured PECs and NCI-H292 cells. Based on the preliminary experiment, we used 10 ng/mL of IL-17A as the optimal concentration for stimulation. As shown in Fig. 5A and B, IL-17A significantly increased p-p38, p-ERK and p-JNK in both PECs and NCI-H292 cells in a time-dependent manner. When adding specific inhibitors of p-p38 (SB203580), p-ERK (U0126) and p-JNK (SP600125), we found SB203580 and U0126, but not SP600125, significantly inhibited IL-17A-induced MUC5AC production (Fig. 5C–E, p<0.05), suggesting activated p38 and ERK were involved in MUC5AC expression in response to IL-17A stimulation. To further evaluate the importance of IL-17RA, IL-17RC and act1 in IL-17A-induced MUC5AC expression in vitro, we examined IL-17A-induced MUC5AC production in cultured NCI-H292 in the presence of IL-17RA, IL-17RC and act1 siRNA. Consequently, we found both IL-17RA and IL-17RC siRNA significantly inhibited the mRNA and protein levels of MUC5AC in IL-17A induced NCI-H292 cells (Fig S3 in File SI). As to act1 expression, IL-17A stimulation significantly increased act1 protein expression in cultured PECs and NCI-H292 cells in a time-dependent manner (Fig. 6A and B, p<0.05). When adding act1 siRNA, we found that act1, p-p38 and p-ERK protein levels were significantly inhibited in NCI-H292 cells compared with the control group (Fig. 6C–G, p<0.05). Consistently, MUC5AC mRNA and protein levels were significantly inhibited in NCI-H292 cells compared with the control group (Fig 6H, p<0.05). These findings suggested that IL-17RA and IL-17RC and act1 were required for IL-17A-induced MUC5AC production in airway epithelial cells.


Interleukin-17A promotes MUC5AC expression and goblet cell hyperplasia in nasal polyps via the Act1-mediated pathway.

Xia W, Bai J, Wu X, Wei Y, Feng S, Li L, Zhang J, Xiong G, Fan Y, Shi J, Li H - PLoS ONE (2014)

MAPK signaling mediated IL-17A induced MUC5AC in PECs and NCI-H292 cells in vitro.(A) Representative western blot result of phosphorylated p38, ERK and JNK in PECs after IL-17A stimulation. (B) Representative western blot result of phosphorylated p38, ERK and JNK in NCI-H292 cells after IL-17A stimulation. (C, D) MUC5AC protein level in cultured PECs and NCI-H292 cells after IL-17A stimulation for 24 h in the presence of specific inhibitors of p38, ERK and JNK. The data are expressed the means (SEM) of 3 independent experiments. * p<0.05 when compared with control PECs, #p<0.05 when compared with control NCI-H292 cells.
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getmorefigures.php?uid=PMC4043856&req=5

pone-0098915-g005: MAPK signaling mediated IL-17A induced MUC5AC in PECs and NCI-H292 cells in vitro.(A) Representative western blot result of phosphorylated p38, ERK and JNK in PECs after IL-17A stimulation. (B) Representative western blot result of phosphorylated p38, ERK and JNK in NCI-H292 cells after IL-17A stimulation. (C, D) MUC5AC protein level in cultured PECs and NCI-H292 cells after IL-17A stimulation for 24 h in the presence of specific inhibitors of p38, ERK and JNK. The data are expressed the means (SEM) of 3 independent experiments. * p<0.05 when compared with control PECs, #p<0.05 when compared with control NCI-H292 cells.
Mentions: We next examined the importance of the MAPK pathway in IL-17A-induced MUC5AC expression in cultured PECs and NCI-H292 cells. Based on the preliminary experiment, we used 10 ng/mL of IL-17A as the optimal concentration for stimulation. As shown in Fig. 5A and B, IL-17A significantly increased p-p38, p-ERK and p-JNK in both PECs and NCI-H292 cells in a time-dependent manner. When adding specific inhibitors of p-p38 (SB203580), p-ERK (U0126) and p-JNK (SP600125), we found SB203580 and U0126, but not SP600125, significantly inhibited IL-17A-induced MUC5AC production (Fig. 5C–E, p<0.05), suggesting activated p38 and ERK were involved in MUC5AC expression in response to IL-17A stimulation. To further evaluate the importance of IL-17RA, IL-17RC and act1 in IL-17A-induced MUC5AC expression in vitro, we examined IL-17A-induced MUC5AC production in cultured NCI-H292 in the presence of IL-17RA, IL-17RC and act1 siRNA. Consequently, we found both IL-17RA and IL-17RC siRNA significantly inhibited the mRNA and protein levels of MUC5AC in IL-17A induced NCI-H292 cells (Fig S3 in File SI). As to act1 expression, IL-17A stimulation significantly increased act1 protein expression in cultured PECs and NCI-H292 cells in a time-dependent manner (Fig. 6A and B, p<0.05). When adding act1 siRNA, we found that act1, p-p38 and p-ERK protein levels were significantly inhibited in NCI-H292 cells compared with the control group (Fig. 6C–G, p<0.05). Consistently, MUC5AC mRNA and protein levels were significantly inhibited in NCI-H292 cells compared with the control group (Fig 6H, p<0.05). These findings suggested that IL-17RA and IL-17RC and act1 were required for IL-17A-induced MUC5AC production in airway epithelial cells.

Bottom Line: IL-17A significantly stimulated the expression of IL-17RA, IL-17RC, act1 and MUC5AC, and the activation of the MAPK pathway in cultured PECs and NCI-H292 cells (p<0.05).In addition, IL-17RA, IL-17RC and act1 siRNA significantly blocked IL-17A-induced MUC5AC production in vitro (p<0.05).Our results suggest that IL-17A plays a crucial role in stimulating the production of MUC5AC and goblet cell hyperplasia through the act1-mediated signaling pathway and may suggest a promising strategy for the management of Th17-dominant NP patients.

View Article: PubMed Central - PubMed

Affiliation: Allergy and Cancer Center, Otorhinolarygology Hospital, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China; Department of Otolaryngology, Head and Neck Surgery, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.

ABSTRACT

Background: Recent studies demonstrated that nasal polyps (NP) patients in China and other Asian regions possessed distinct Th17-dominant inflammation and enhanced tissue remodeling. However, the mechanism underlying these observations is not fully understood. This study sought to evaluate the association of interleukin (IL)-17A with MUC5AC expression and goblet cell hyperplasia in Chinese NP patients and to characterize the signaling pathway underlying IL-17A-induced MUC5AC expression in vitro.

Method: We enrolled 25 NP patients and 22 normal controls and examined the expression of IL-17A, MUC5AC and act1 in polyp tissues by immunohistochemical (IHC) staining, quantitative polymerase chain reaction (qPCR) and western blot. Moreover, by using an in vitro culture system of polyp epithelial cells (PECs), IL-17A-induced gene expression was screened in cultured PECs by DNA microarray. The expression of IL-17RA, IL-17RC, act1 and MUC5AC and the activation of the MAPK pathway (ERK, p38 and JNK), were further examined in cultured PECs and NCI-H292 cells by qPCR and western blotting, respectively.

Results: We found that increased IL-17A production was significantly correlated with MUC5AC and act1 expression and goblet cell hyperplasia in polyp tissues (p<0.05). IL-17A significantly stimulated the expression of IL-17RA, IL-17RC, act1 and MUC5AC, and the activation of the MAPK pathway in cultured PECs and NCI-H292 cells (p<0.05). In addition, IL-17RA, IL-17RC and act1 siRNA significantly blocked IL-17A-induced MUC5AC production in vitro (p<0.05).

Conclusion: Our results suggest that IL-17A plays a crucial role in stimulating the production of MUC5AC and goblet cell hyperplasia through the act1-mediated signaling pathway and may suggest a promising strategy for the management of Th17-dominant NP patients.

Show MeSH
Related in: MedlinePlus