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Klotho expression is reduced in COPD airway epithelial cells: effects on inflammation and oxidant injury.

Gao W, Yuan C, Zhang J, Li L, Yu L, Wiegman CH, Barnes PJ, Adcock IM, Huang M, Yao X - Clin. Sci. (2015)

Bottom Line: Moreover, the effect of KL on CSE-mediated inflammation and hydrogen peroxide-induced cellular injury/apoptosis was determined using siRNAs.CSE and TNFα (tumour necrosis factor α) decreased KL expression and release from airway epithelial cells, which was associated with enhanced pro-inflammatory cytokine expression.These effects involved the NF-κB (nuclear factor κB), MAPK (mitogen-activated protein kinase) and Nrf2 (nuclear factor erythroid 2-related factor 2) pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China.

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Klotho knockdown modulates epithelial cell survival and inflammatory gene expression(A) Transfection of 16HBE cells with KL siRNAs (siRNA1–siRNA3) significantly down-regulated KL protein and mRNA expression at 24 h compared with control siRNA (siRNAc) treatment. (B) CCK-8 staining shows that KL knockdown slightly, but significantly, reduced cell viability compared with control siRNA. (C) KL-directed siRNAs (siRNA1–siRNA3) induced apoptosis 24 h after transfection as determined by flow cytometry. Representative scatter plots of untreated control cells (CON), siRNAc-treated cells (NC) and siRNA1–siRNA3-treated cells (KL1019, KL1278 and KL2071) are shown. Results in the histogram are means±S.E.M. for three independent experiments. #P<0.05 compared with the control group, *P<0.05 compared with control siRNAc. (D) IL6, MCP1 and IL8 mRNA expression significantly increased after KL knockdown. Results are means±S.E.M. from three independent experiments. (*P<0.05, **P<0.01, ***P<0.001).
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Figure 5: Klotho knockdown modulates epithelial cell survival and inflammatory gene expression(A) Transfection of 16HBE cells with KL siRNAs (siRNA1–siRNA3) significantly down-regulated KL protein and mRNA expression at 24 h compared with control siRNA (siRNAc) treatment. (B) CCK-8 staining shows that KL knockdown slightly, but significantly, reduced cell viability compared with control siRNA. (C) KL-directed siRNAs (siRNA1–siRNA3) induced apoptosis 24 h after transfection as determined by flow cytometry. Representative scatter plots of untreated control cells (CON), siRNAc-treated cells (NC) and siRNA1–siRNA3-treated cells (KL1019, KL1278 and KL2071) are shown. Results in the histogram are means±S.E.M. for three independent experiments. #P<0.05 compared with the control group, *P<0.05 compared with control siRNAc. (D) IL6, MCP1 and IL8 mRNA expression significantly increased after KL knockdown. Results are means±S.E.M. from three independent experiments. (*P<0.05, **P<0.01, ***P<0.001).

Mentions: In reverse experiments, we examined the effect of KL knockdown on epithelial cell function by using selective siRNAs. KL knockdown resulted in an 80–95% suppression of KL protein and mRNA expression, with siRNA3 being the most effective (Figure 5A). KL knockdown showed a small, but significant, decrease in cell viability compared with the control siRNA (Figure 5B), and was associated with an increase in apoptosis (Figure 5C). The apoptosis level induced by control siRNA transfection alone (9.6±1.3% compared with 18.5±1.6%, P<0.05) was enhanced further with individual KL-directed siRNAs (siRNA2: 29.1±2.5%, P<0.05 compared with control siRNA; Figure 5C). In addition, the mRNA expression of inflammatory cytokines [IL-6, MCP-1 (monocyte chemoattractant protein 1) and IL-8] was significantly upregulated by KL knockdown (Figure 5D). However, no effect on protein secretion was observed (results not shown).


Klotho expression is reduced in COPD airway epithelial cells: effects on inflammation and oxidant injury.

Gao W, Yuan C, Zhang J, Li L, Yu L, Wiegman CH, Barnes PJ, Adcock IM, Huang M, Yao X - Clin. Sci. (2015)

Klotho knockdown modulates epithelial cell survival and inflammatory gene expression(A) Transfection of 16HBE cells with KL siRNAs (siRNA1–siRNA3) significantly down-regulated KL protein and mRNA expression at 24 h compared with control siRNA (siRNAc) treatment. (B) CCK-8 staining shows that KL knockdown slightly, but significantly, reduced cell viability compared with control siRNA. (C) KL-directed siRNAs (siRNA1–siRNA3) induced apoptosis 24 h after transfection as determined by flow cytometry. Representative scatter plots of untreated control cells (CON), siRNAc-treated cells (NC) and siRNA1–siRNA3-treated cells (KL1019, KL1278 and KL2071) are shown. Results in the histogram are means±S.E.M. for three independent experiments. #P<0.05 compared with the control group, *P<0.05 compared with control siRNAc. (D) IL6, MCP1 and IL8 mRNA expression significantly increased after KL knockdown. Results are means±S.E.M. from three independent experiments. (*P<0.05, **P<0.01, ***P<0.001).
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Figure 5: Klotho knockdown modulates epithelial cell survival and inflammatory gene expression(A) Transfection of 16HBE cells with KL siRNAs (siRNA1–siRNA3) significantly down-regulated KL protein and mRNA expression at 24 h compared with control siRNA (siRNAc) treatment. (B) CCK-8 staining shows that KL knockdown slightly, but significantly, reduced cell viability compared with control siRNA. (C) KL-directed siRNAs (siRNA1–siRNA3) induced apoptosis 24 h after transfection as determined by flow cytometry. Representative scatter plots of untreated control cells (CON), siRNAc-treated cells (NC) and siRNA1–siRNA3-treated cells (KL1019, KL1278 and KL2071) are shown. Results in the histogram are means±S.E.M. for three independent experiments. #P<0.05 compared with the control group, *P<0.05 compared with control siRNAc. (D) IL6, MCP1 and IL8 mRNA expression significantly increased after KL knockdown. Results are means±S.E.M. from three independent experiments. (*P<0.05, **P<0.01, ***P<0.001).
Mentions: In reverse experiments, we examined the effect of KL knockdown on epithelial cell function by using selective siRNAs. KL knockdown resulted in an 80–95% suppression of KL protein and mRNA expression, with siRNA3 being the most effective (Figure 5A). KL knockdown showed a small, but significant, decrease in cell viability compared with the control siRNA (Figure 5B), and was associated with an increase in apoptosis (Figure 5C). The apoptosis level induced by control siRNA transfection alone (9.6±1.3% compared with 18.5±1.6%, P<0.05) was enhanced further with individual KL-directed siRNAs (siRNA2: 29.1±2.5%, P<0.05 compared with control siRNA; Figure 5C). In addition, the mRNA expression of inflammatory cytokines [IL-6, MCP-1 (monocyte chemoattractant protein 1) and IL-8] was significantly upregulated by KL knockdown (Figure 5D). However, no effect on protein secretion was observed (results not shown).

Bottom Line: Moreover, the effect of KL on CSE-mediated inflammation and hydrogen peroxide-induced cellular injury/apoptosis was determined using siRNAs.CSE and TNFα (tumour necrosis factor α) decreased KL expression and release from airway epithelial cells, which was associated with enhanced pro-inflammatory cytokine expression.These effects involved the NF-κB (nuclear factor κB), MAPK (mitogen-activated protein kinase) and Nrf2 (nuclear factor erythroid 2-related factor 2) pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China.

Show MeSH
Related in: MedlinePlus