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Inhibition of the IgE-mediated activation of RBL-2H3 cells by TIPP, a novel thymic immunosuppressive pentapeptide.

Lian Q, Cheng Y, Zhong C, Wang F - Int J Mol Sci (2015)

Bottom Line: TIPP is a novel thymic immunosuppressive pentapeptide originally obtained from calf thymic immunosuppressive extract.TIPP significantly suppressed the increase of intracellular calcium and the rearrangement of F-actin, attenuated the transcription of pro-inflammatory cytokines (IL-3, -4, -6, -13, TNF-α, and monocyte chemotactic protein-1 (MCP-1)), and decreased the expression of COX-2.Western blot analysis showed that TIPP had an inhibitory activity on the phosphorylation of extracellular signal-regulated protein kinase 1/2 (ERK1/2) and ERK kinase 1/2 (MEK1/2), and inhibited the activation of NF-κB.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Chemical Biology of Natural Products (Ministry of Education), Institute of Biochemical and Biotechnological Drugs, School of Pharmaceutical Sciences, Shandong University, Jinan 250012, China. xhrg_1042@126.com.

ABSTRACT
TIPP is a novel thymic immunosuppressive pentapeptide originally obtained from calf thymic immunosuppressive extract. The present study aimed to investigate the inhibitory activity of TIPP on IgE-mediated activation of RBL-2H3 cells. Release of β-hexosaminidase and histamine, intracellular calcium, membrane ruffling, mRNA levels of cytokines, cyclooxygenase-2 (COX-2) expression, and activation of mitogen-activated protein kinases (MAP kinases) and NF-κB were determined by colorimetric assay, fluorescence spectrophotometer, confocal fluorescence microscope, quantification PCR, and Western blot, respectively. The results showed that TIPP significantly inhibited the degranulation in IgE-antigen complex-stimulated RBL-2H3 cells without cytotoxicity. TIPP significantly suppressed the increase of intracellular calcium and the rearrangement of F-actin, attenuated the transcription of pro-inflammatory cytokines (IL-3, -4, -6, -13, TNF-α, and monocyte chemotactic protein-1 (MCP-1)), and decreased the expression of COX-2. Western blot analysis showed that TIPP had an inhibitory activity on the phosphorylation of extracellular signal-regulated protein kinase 1/2 (ERK1/2) and ERK kinase 1/2 (MEK1/2), and inhibited the activation of NF-κB. The data suggested that TIPP effectively suppressed IgE-mediated activation of RBL-2H3 cells via blocking MEK/ERK and NF-κB signaling pathways.

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Related in: MedlinePlus

Confocal fluorescence microscope observation of F-actin (labeled with rhodamin-phalloidin) in RBL-2H3 cells. The arrow represents membrane ruffling caused by F-actin rearrangement. (a) IgE-sensitized RBL-2H3 cells stimulated with PBS; (b) IgE-sensitized RBL-2H3 cells stimulated with DNP-HSA for 30 min; (c–f), IgE-sensitized RBL-2H3 cells, pretreated with 50, 200, and 800 μg/mL of TIPP or 20 μg/mL of ketotifen, then stimulated with DNP-HSA for 30 min. Magnification: 63×.
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ijms-16-02252-f005: Confocal fluorescence microscope observation of F-actin (labeled with rhodamin-phalloidin) in RBL-2H3 cells. The arrow represents membrane ruffling caused by F-actin rearrangement. (a) IgE-sensitized RBL-2H3 cells stimulated with PBS; (b) IgE-sensitized RBL-2H3 cells stimulated with DNP-HSA for 30 min; (c–f), IgE-sensitized RBL-2H3 cells, pretreated with 50, 200, and 800 μg/mL of TIPP or 20 μg/mL of ketotifen, then stimulated with DNP-HSA for 30 min. Magnification: 63×.

Mentions: As shown in Figure 5, F-actin is neatly arranged around the periphery of the normal RBL-2H3 cells (Figure 5a). Stimulation with DNP-HSA antigen caused a remarkable rearrangement of F-actin, which resulted in a membrane ruffling (Figure 5b). Membrane ruffling was decreased apparently and F-actin tended to be neatly arranged when RBL-2H3 cells were pre-treated with 800 μg/mL of TIPP or 20 μg/mL of ketotifen (Figure 5e,f).


Inhibition of the IgE-mediated activation of RBL-2H3 cells by TIPP, a novel thymic immunosuppressive pentapeptide.

Lian Q, Cheng Y, Zhong C, Wang F - Int J Mol Sci (2015)

Confocal fluorescence microscope observation of F-actin (labeled with rhodamin-phalloidin) in RBL-2H3 cells. The arrow represents membrane ruffling caused by F-actin rearrangement. (a) IgE-sensitized RBL-2H3 cells stimulated with PBS; (b) IgE-sensitized RBL-2H3 cells stimulated with DNP-HSA for 30 min; (c–f), IgE-sensitized RBL-2H3 cells, pretreated with 50, 200, and 800 μg/mL of TIPP or 20 μg/mL of ketotifen, then stimulated with DNP-HSA for 30 min. Magnification: 63×.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4307361&req=5

ijms-16-02252-f005: Confocal fluorescence microscope observation of F-actin (labeled with rhodamin-phalloidin) in RBL-2H3 cells. The arrow represents membrane ruffling caused by F-actin rearrangement. (a) IgE-sensitized RBL-2H3 cells stimulated with PBS; (b) IgE-sensitized RBL-2H3 cells stimulated with DNP-HSA for 30 min; (c–f), IgE-sensitized RBL-2H3 cells, pretreated with 50, 200, and 800 μg/mL of TIPP or 20 μg/mL of ketotifen, then stimulated with DNP-HSA for 30 min. Magnification: 63×.
Mentions: As shown in Figure 5, F-actin is neatly arranged around the periphery of the normal RBL-2H3 cells (Figure 5a). Stimulation with DNP-HSA antigen caused a remarkable rearrangement of F-actin, which resulted in a membrane ruffling (Figure 5b). Membrane ruffling was decreased apparently and F-actin tended to be neatly arranged when RBL-2H3 cells were pre-treated with 800 μg/mL of TIPP or 20 μg/mL of ketotifen (Figure 5e,f).

Bottom Line: TIPP is a novel thymic immunosuppressive pentapeptide originally obtained from calf thymic immunosuppressive extract.TIPP significantly suppressed the increase of intracellular calcium and the rearrangement of F-actin, attenuated the transcription of pro-inflammatory cytokines (IL-3, -4, -6, -13, TNF-α, and monocyte chemotactic protein-1 (MCP-1)), and decreased the expression of COX-2.Western blot analysis showed that TIPP had an inhibitory activity on the phosphorylation of extracellular signal-regulated protein kinase 1/2 (ERK1/2) and ERK kinase 1/2 (MEK1/2), and inhibited the activation of NF-κB.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Chemical Biology of Natural Products (Ministry of Education), Institute of Biochemical and Biotechnological Drugs, School of Pharmaceutical Sciences, Shandong University, Jinan 250012, China. xhrg_1042@126.com.

ABSTRACT
TIPP is a novel thymic immunosuppressive pentapeptide originally obtained from calf thymic immunosuppressive extract. The present study aimed to investigate the inhibitory activity of TIPP on IgE-mediated activation of RBL-2H3 cells. Release of β-hexosaminidase and histamine, intracellular calcium, membrane ruffling, mRNA levels of cytokines, cyclooxygenase-2 (COX-2) expression, and activation of mitogen-activated protein kinases (MAP kinases) and NF-κB were determined by colorimetric assay, fluorescence spectrophotometer, confocal fluorescence microscope, quantification PCR, and Western blot, respectively. The results showed that TIPP significantly inhibited the degranulation in IgE-antigen complex-stimulated RBL-2H3 cells without cytotoxicity. TIPP significantly suppressed the increase of intracellular calcium and the rearrangement of F-actin, attenuated the transcription of pro-inflammatory cytokines (IL-3, -4, -6, -13, TNF-α, and monocyte chemotactic protein-1 (MCP-1)), and decreased the expression of COX-2. Western blot analysis showed that TIPP had an inhibitory activity on the phosphorylation of extracellular signal-regulated protein kinase 1/2 (ERK1/2) and ERK kinase 1/2 (MEK1/2), and inhibited the activation of NF-κB. The data suggested that TIPP effectively suppressed IgE-mediated activation of RBL-2H3 cells via blocking MEK/ERK and NF-κB signaling pathways.

Show MeSH
Related in: MedlinePlus