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Involvement of B2 receptor in bradykinin-induced proliferation and proinflammatory effects in human nasal mucosa-derived fibroblasts isolated from chronic rhinosinusitis patients.

Tsai YJ, Hao SP, Chen CL, Lin BJ, Wu WB - PLoS ONE (2015)

Bottom Line: Accordingly, the B2R was preferentially expressed in the NMDFs than B1R.Collectively, we report here for the first time that fibroblasts, KNG/BK, and BKRs are overexpressed in CRSsNP mucosa and BK upregulates chemokine expression, proliferation, and proinflammatory molecule expression in NMDFs via B2R activation, which lead to a functional increase in monocyte-fibroblast interaction.Our findings reveal a critical role of fibroblast, KNG/BK, and BKRs in the development of CRSsNP.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology Head and Neck Surgery, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; Department of Chemistry, Graduate Institute of Applied Science and Engineering, College of Science and Engineering, Fu-Jen Catholic University, New Taipei City, Taiwan; Graduate Institute of Basic Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan; School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan.

ABSTRACT
Chronic rhinosinusitis (CRS) is a chronic inflammatory disease of the sinonasal mucosa either accompanied by polyp formation (CRSwNP) or without polyps (CRSsNP). CRSsNP accounts for the majority of CRS cases and is characterized by fibrosis and neutrophilic inflammation. However, the pathogenesis of CRS, especially CRSsNP, remains unclear. Immunohistochemistry of CRSsNP specimens in the present study showed that the submucosa, perivascular areas, and the mucous glands were abundant in fibroblasts. Therefore, we investigated the effects bradykinin (BK), an autacoid known to participate in inflammation, on human CRSsNP nasal mucosa-derived fibroblasts (NMDFs). BK increased CXCL1 and -8 secretion and mRNA expression with EC50 ranging from 0.15~0.35 μM. Moreover, BK enhanced cell proliferation and upregulated the expressions of proinflammatory molecules, including cell adhesion molecules (CAMs) and cyclooxygenase (COX)-1 and -2. These functionally caused an increase in monocyte adhesion to fibroblast monolayer. Using pharmacological intervention and BKR siRNA knockdown, we demonstrated that the BK-induced CXCL chemokine release, cell proliferation and COX and CAM expressions were mainly through the B2 receptor (B2R). Accordingly, the B2R was preferentially expressed in the NMDFs than B1R. The B2R was highly expressed in the CRSsNP than the control specimens, while the B1R and kininogen (KNG)/BK expression slightly increased in the CRSsNP mucosa. Collectively, we report here for the first time that fibroblasts, KNG/BK, and BKRs are overexpressed in CRSsNP mucosa and BK upregulates chemokine expression, proliferation, and proinflammatory molecule expression in NMDFs via B2R activation, which lead to a functional increase in monocyte-fibroblast interaction. Our findings reveal a critical role of fibroblast, KNG/BK, and BKRs in the development of CRSsNP.

No MeSH data available.


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Effects of BK on cell proliferation, proinflammatory molecule expression, and monocyte adhesion.NMDFs were treated with the indicated concentrations of BK for 16 h, and (A) cell proliferation was determined by the MTT assay and direct cell counting assay (n = 3–4). (B) The expression of CAMs and COXs was determined by western blotting (n = 4). (C) Effect of BK on monocytes adhesion to fibroblasts. Twenty-four well plates seeded with or without NMDFs were treated with BK (1 μM) for 16 h. Then, BCECF/AM-labeled monocytes were allowed to adhere to the wells for 1 h. After a brief wash, monocytes adhered to wells (green spots) were determined by direct cell counting under the high-powered field (HPF) (n = 4). *P<0.05, **P<0.01, ***p < 0.001 versus basal (BK 0 μM).
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pone.0126853.g003: Effects of BK on cell proliferation, proinflammatory molecule expression, and monocyte adhesion.NMDFs were treated with the indicated concentrations of BK for 16 h, and (A) cell proliferation was determined by the MTT assay and direct cell counting assay (n = 3–4). (B) The expression of CAMs and COXs was determined by western blotting (n = 4). (C) Effect of BK on monocytes adhesion to fibroblasts. Twenty-four well plates seeded with or without NMDFs were treated with BK (1 μM) for 16 h. Then, BCECF/AM-labeled monocytes were allowed to adhere to the wells for 1 h. After a brief wash, monocytes adhered to wells (green spots) were determined by direct cell counting under the high-powered field (HPF) (n = 4). *P<0.05, **P<0.01, ***p < 0.001 versus basal (BK 0 μM).

Mentions: To examine whether BK affects fibroblast proliferation, NMDFs were treated with the indicated concentrations of BK for 16 h and cell proliferation was determined by the MTT assay and by direct cell counting using flow cytometry. As shown in Fig 3A,0.05μM BK effectively induced significant cell growth. The cell growth reached a maximum with 0.2 μM of BK (left panel). Flow cytometric analysis of cell number showed similar results to the MTT assay (right panel). Next, NMDFs were treated with the indicated concentrations of BK for 16 h, the expression of CAMs and COXs was then determined by western blot analysis. As shown in Fig 3B, 005μM BK could induce the expression of ICAM-1 and VCAM-1. The VCAM-1 expression by BK was apparent at 0.05 μM of BK but slightly declined when the concentrations increased. Strikingly, BK induced COX-2 and -1 expression in the NMDFs.


Involvement of B2 receptor in bradykinin-induced proliferation and proinflammatory effects in human nasal mucosa-derived fibroblasts isolated from chronic rhinosinusitis patients.

Tsai YJ, Hao SP, Chen CL, Lin BJ, Wu WB - PLoS ONE (2015)

Effects of BK on cell proliferation, proinflammatory molecule expression, and monocyte adhesion.NMDFs were treated with the indicated concentrations of BK for 16 h, and (A) cell proliferation was determined by the MTT assay and direct cell counting assay (n = 3–4). (B) The expression of CAMs and COXs was determined by western blotting (n = 4). (C) Effect of BK on monocytes adhesion to fibroblasts. Twenty-four well plates seeded with or without NMDFs were treated with BK (1 μM) for 16 h. Then, BCECF/AM-labeled monocytes were allowed to adhere to the wells for 1 h. After a brief wash, monocytes adhered to wells (green spots) were determined by direct cell counting under the high-powered field (HPF) (n = 4). *P<0.05, **P<0.01, ***p < 0.001 versus basal (BK 0 μM).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4430235&req=5

pone.0126853.g003: Effects of BK on cell proliferation, proinflammatory molecule expression, and monocyte adhesion.NMDFs were treated with the indicated concentrations of BK for 16 h, and (A) cell proliferation was determined by the MTT assay and direct cell counting assay (n = 3–4). (B) The expression of CAMs and COXs was determined by western blotting (n = 4). (C) Effect of BK on monocytes adhesion to fibroblasts. Twenty-four well plates seeded with or without NMDFs were treated with BK (1 μM) for 16 h. Then, BCECF/AM-labeled monocytes were allowed to adhere to the wells for 1 h. After a brief wash, monocytes adhered to wells (green spots) were determined by direct cell counting under the high-powered field (HPF) (n = 4). *P<0.05, **P<0.01, ***p < 0.001 versus basal (BK 0 μM).
Mentions: To examine whether BK affects fibroblast proliferation, NMDFs were treated with the indicated concentrations of BK for 16 h and cell proliferation was determined by the MTT assay and by direct cell counting using flow cytometry. As shown in Fig 3A,0.05μM BK effectively induced significant cell growth. The cell growth reached a maximum with 0.2 μM of BK (left panel). Flow cytometric analysis of cell number showed similar results to the MTT assay (right panel). Next, NMDFs were treated with the indicated concentrations of BK for 16 h, the expression of CAMs and COXs was then determined by western blot analysis. As shown in Fig 3B, 005μM BK could induce the expression of ICAM-1 and VCAM-1. The VCAM-1 expression by BK was apparent at 0.05 μM of BK but slightly declined when the concentrations increased. Strikingly, BK induced COX-2 and -1 expression in the NMDFs.

Bottom Line: Accordingly, the B2R was preferentially expressed in the NMDFs than B1R.Collectively, we report here for the first time that fibroblasts, KNG/BK, and BKRs are overexpressed in CRSsNP mucosa and BK upregulates chemokine expression, proliferation, and proinflammatory molecule expression in NMDFs via B2R activation, which lead to a functional increase in monocyte-fibroblast interaction.Our findings reveal a critical role of fibroblast, KNG/BK, and BKRs in the development of CRSsNP.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology Head and Neck Surgery, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; Department of Chemistry, Graduate Institute of Applied Science and Engineering, College of Science and Engineering, Fu-Jen Catholic University, New Taipei City, Taiwan; Graduate Institute of Basic Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan; School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan.

ABSTRACT
Chronic rhinosinusitis (CRS) is a chronic inflammatory disease of the sinonasal mucosa either accompanied by polyp formation (CRSwNP) or without polyps (CRSsNP). CRSsNP accounts for the majority of CRS cases and is characterized by fibrosis and neutrophilic inflammation. However, the pathogenesis of CRS, especially CRSsNP, remains unclear. Immunohistochemistry of CRSsNP specimens in the present study showed that the submucosa, perivascular areas, and the mucous glands were abundant in fibroblasts. Therefore, we investigated the effects bradykinin (BK), an autacoid known to participate in inflammation, on human CRSsNP nasal mucosa-derived fibroblasts (NMDFs). BK increased CXCL1 and -8 secretion and mRNA expression with EC50 ranging from 0.15~0.35 μM. Moreover, BK enhanced cell proliferation and upregulated the expressions of proinflammatory molecules, including cell adhesion molecules (CAMs) and cyclooxygenase (COX)-1 and -2. These functionally caused an increase in monocyte adhesion to fibroblast monolayer. Using pharmacological intervention and BKR siRNA knockdown, we demonstrated that the BK-induced CXCL chemokine release, cell proliferation and COX and CAM expressions were mainly through the B2 receptor (B2R). Accordingly, the B2R was preferentially expressed in the NMDFs than B1R. The B2R was highly expressed in the CRSsNP than the control specimens, while the B1R and kininogen (KNG)/BK expression slightly increased in the CRSsNP mucosa. Collectively, we report here for the first time that fibroblasts, KNG/BK, and BKRs are overexpressed in CRSsNP mucosa and BK upregulates chemokine expression, proliferation, and proinflammatory molecule expression in NMDFs via B2R activation, which lead to a functional increase in monocyte-fibroblast interaction. Our findings reveal a critical role of fibroblast, KNG/BK, and BKRs in the development of CRSsNP.

No MeSH data available.


Related in: MedlinePlus