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An anti-inflammatory role for C/EBPδ in human brain pericytes.

Rustenhoven J, Scotter EL, Jansson D, Kho DT, Oldfield RL, Bergin PS, Mee EW, Faull RL, Curtis MA, Graham SE, Park TI, Dragunow M - Sci Rep (2015)

Bottom Line: Our results reveal that C/EBPδ is induced in a concentration- and time-dependent fashion in human brain pericytes by interleukin-1β (IL-1β).C/EBPδ knockdown enhanced IL-1β-induced production of intracellular adhesion molecule-1 (ICAM-1), interleukin-8, monocyte chemoattractant protein-1 (MCP-1) and IL-1β, whilst attenuating cyclooxygenase-2 and superoxide dismutase-2 gene expression.Altered ICAM-1 and MCP-1 protein expression were confirmed by cytometric bead array and immunocytochemistry.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Pharmacology and Clinical Pharmacology [2] Centre for Brain Research.

ABSTRACT
Neuroinflammation contributes to the pathogenesis of several neurological disorders and pericytes are implicated in brain inflammatory processes. Cellular inflammatory responses are orchestrated by transcription factors but information on transcriptional control in pericytes is lacking. Because the transcription factor CCAAT/enhancer binding protein delta (C/EBPδ) is induced in a number of inflammatory brain disorders, we sought to investigate its role in regulating pericyte immune responses. Our results reveal that C/EBPδ is induced in a concentration- and time-dependent fashion in human brain pericytes by interleukin-1β (IL-1β). To investigate the function of the induced C/EBPδ in pericytes we used siRNA to knockdown IL-1β-induced C/EBPδ expression. C/EBPδ knockdown enhanced IL-1β-induced production of intracellular adhesion molecule-1 (ICAM-1), interleukin-8, monocyte chemoattractant protein-1 (MCP-1) and IL-1β, whilst attenuating cyclooxygenase-2 and superoxide dismutase-2 gene expression. Altered ICAM-1 and MCP-1 protein expression were confirmed by cytometric bead array and immunocytochemistry. Our results show that knock-down of C/EBPδ expression in pericytes following immune stimulation increased chemokine and adhesion molecule expression, thus modifying the human brain pericyte inflammatory response. The induction of C/EBPδ following immune stimulation may act to limit infiltration of peripheral immune cells, thereby preventing further inflammatory responses in the brain.

No MeSH data available.


Related in: MedlinePlus

C/EBPδ knockdown modifies pericyte secretion of inflammatory mediators.Human brain pericytes were transfected with 50 nM of control or C/EBPδ siRNA for 48 hours. Following transfection cells were treated with vehicle or 10 ng/mL IL-1β for 24 hours and conditioned media collected. Concentration of sICAM-1 (a), MCP-1 (b), IL-8 (c) and IL-6 (d) in media was determined using a multiplex cytometric bead array. Data represent mean ± SEM (n = 4). *** = p < 0.001.
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f8: C/EBPδ knockdown modifies pericyte secretion of inflammatory mediators.Human brain pericytes were transfected with 50 nM of control or C/EBPδ siRNA for 48 hours. Following transfection cells were treated with vehicle or 10 ng/mL IL-1β for 24 hours and conditioned media collected. Concentration of sICAM-1 (a), MCP-1 (b), IL-8 (c) and IL-6 (d) in media was determined using a multiplex cytometric bead array. Data represent mean ± SEM (n = 4). *** = p < 0.001.

Mentions: Secreted cytokines and chemokines are essential in modulating cellular cross-talk and inflammatory responses. In order to determine whether the observed changed in mRNA and protein expression correlated with increased secretion, cytokine concentrations in the pericytes conditioned media were measured using a cytometric bead array (CBA). Unstimulated pericytes demonstrated basal expression of soluble ICAM-1 (sICAM-1; 3.13 ± 0.75 pg/mL), MCP-1 (4,560.79 ± 143.38 pg/mL) , IL-8 (2,483.51 ± 202.58 pg/mL) and IL-6 (22.75 ± 2.80 pg/mL), which were not significantly altered by C/EBPδ siRNA (ICAM-1 4.15 ± 0.3 pg/mL; p > 0.05, MCP-1 4,837.74 ± 188.27 pg/mL ; p > 0.05, IL-8 2,345.53 ± 206.12 pg/mL ; p > 0.05 and IL-6 21.25 ± 2.66 pg/mL; p > 0.05; Fig. 8a–d). IL-1β treatment significantly increased the concentration of all measured cytokines in the media (ICAM-1 139.90 ± 11.85 pg/mL; p < 0.001, MCP-1 11,755.35 ± 69.58 pg/mL; p < 0.001, IL-8 37,616.57 ± 10,815.63 pg/mL; p < 0.001 and IL-6 6,040.13 ± 885.67 pg/mL; p < 0.001; Fig. 8a–d). Compared to the control siRNA condition, C/EBPδ siRNA significantly enhanced the IL-1β induced secretion of ICAM-1 (246.5 ± 10.93 pg/mL; p < 0.001; Fig. 8a) and MCP-1 (14,551.65 ± 205.73 pg/mL; p < 0.001; Fig. 8b); however, did not alter IL-8 (50,833.75 ± 2,719.64 pg/mL; p > 0.05; Fig. 8c) or IL-6 secretion (7,724.97 ± 705.35 pg/mL; p > 0.05; Fig. 8d).


An anti-inflammatory role for C/EBPδ in human brain pericytes.

Rustenhoven J, Scotter EL, Jansson D, Kho DT, Oldfield RL, Bergin PS, Mee EW, Faull RL, Curtis MA, Graham SE, Park TI, Dragunow M - Sci Rep (2015)

C/EBPδ knockdown modifies pericyte secretion of inflammatory mediators.Human brain pericytes were transfected with 50 nM of control or C/EBPδ siRNA for 48 hours. Following transfection cells were treated with vehicle or 10 ng/mL IL-1β for 24 hours and conditioned media collected. Concentration of sICAM-1 (a), MCP-1 (b), IL-8 (c) and IL-6 (d) in media was determined using a multiplex cytometric bead array. Data represent mean ± SEM (n = 4). *** = p < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4499812&req=5

f8: C/EBPδ knockdown modifies pericyte secretion of inflammatory mediators.Human brain pericytes were transfected with 50 nM of control or C/EBPδ siRNA for 48 hours. Following transfection cells were treated with vehicle or 10 ng/mL IL-1β for 24 hours and conditioned media collected. Concentration of sICAM-1 (a), MCP-1 (b), IL-8 (c) and IL-6 (d) in media was determined using a multiplex cytometric bead array. Data represent mean ± SEM (n = 4). *** = p < 0.001.
Mentions: Secreted cytokines and chemokines are essential in modulating cellular cross-talk and inflammatory responses. In order to determine whether the observed changed in mRNA and protein expression correlated with increased secretion, cytokine concentrations in the pericytes conditioned media were measured using a cytometric bead array (CBA). Unstimulated pericytes demonstrated basal expression of soluble ICAM-1 (sICAM-1; 3.13 ± 0.75 pg/mL), MCP-1 (4,560.79 ± 143.38 pg/mL) , IL-8 (2,483.51 ± 202.58 pg/mL) and IL-6 (22.75 ± 2.80 pg/mL), which were not significantly altered by C/EBPδ siRNA (ICAM-1 4.15 ± 0.3 pg/mL; p > 0.05, MCP-1 4,837.74 ± 188.27 pg/mL ; p > 0.05, IL-8 2,345.53 ± 206.12 pg/mL ; p > 0.05 and IL-6 21.25 ± 2.66 pg/mL; p > 0.05; Fig. 8a–d). IL-1β treatment significantly increased the concentration of all measured cytokines in the media (ICAM-1 139.90 ± 11.85 pg/mL; p < 0.001, MCP-1 11,755.35 ± 69.58 pg/mL; p < 0.001, IL-8 37,616.57 ± 10,815.63 pg/mL; p < 0.001 and IL-6 6,040.13 ± 885.67 pg/mL; p < 0.001; Fig. 8a–d). Compared to the control siRNA condition, C/EBPδ siRNA significantly enhanced the IL-1β induced secretion of ICAM-1 (246.5 ± 10.93 pg/mL; p < 0.001; Fig. 8a) and MCP-1 (14,551.65 ± 205.73 pg/mL; p < 0.001; Fig. 8b); however, did not alter IL-8 (50,833.75 ± 2,719.64 pg/mL; p > 0.05; Fig. 8c) or IL-6 secretion (7,724.97 ± 705.35 pg/mL; p > 0.05; Fig. 8d).

Bottom Line: Our results reveal that C/EBPδ is induced in a concentration- and time-dependent fashion in human brain pericytes by interleukin-1β (IL-1β).C/EBPδ knockdown enhanced IL-1β-induced production of intracellular adhesion molecule-1 (ICAM-1), interleukin-8, monocyte chemoattractant protein-1 (MCP-1) and IL-1β, whilst attenuating cyclooxygenase-2 and superoxide dismutase-2 gene expression.Altered ICAM-1 and MCP-1 protein expression were confirmed by cytometric bead array and immunocytochemistry.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Pharmacology and Clinical Pharmacology [2] Centre for Brain Research.

ABSTRACT
Neuroinflammation contributes to the pathogenesis of several neurological disorders and pericytes are implicated in brain inflammatory processes. Cellular inflammatory responses are orchestrated by transcription factors but information on transcriptional control in pericytes is lacking. Because the transcription factor CCAAT/enhancer binding protein delta (C/EBPδ) is induced in a number of inflammatory brain disorders, we sought to investigate its role in regulating pericyte immune responses. Our results reveal that C/EBPδ is induced in a concentration- and time-dependent fashion in human brain pericytes by interleukin-1β (IL-1β). To investigate the function of the induced C/EBPδ in pericytes we used siRNA to knockdown IL-1β-induced C/EBPδ expression. C/EBPδ knockdown enhanced IL-1β-induced production of intracellular adhesion molecule-1 (ICAM-1), interleukin-8, monocyte chemoattractant protein-1 (MCP-1) and IL-1β, whilst attenuating cyclooxygenase-2 and superoxide dismutase-2 gene expression. Altered ICAM-1 and MCP-1 protein expression were confirmed by cytometric bead array and immunocytochemistry. Our results show that knock-down of C/EBPδ expression in pericytes following immune stimulation increased chemokine and adhesion molecule expression, thus modifying the human brain pericyte inflammatory response. The induction of C/EBPδ following immune stimulation may act to limit infiltration of peripheral immune cells, thereby preventing further inflammatory responses in the brain.

No MeSH data available.


Related in: MedlinePlus