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Monocyte chemoattractant protein-1-deficiency impairs the expression of IL-6, IL-1β and G-CSF after transient focal ischemia in mice.

Strecker JK, Minnerup J, Gess B, Ringelstein EB, Schäbitz WR, Schilling M - PLoS ONE (2011)

Bottom Line: Here we report that MCP-1-deficiency leads to a shift towards a less inflammatory state following experimental occlusion of the middle cerebral artery including an impaired induction of interleukin-6, interleukin-1β and granulocyte-colony stimulating factor expression as well as a subsequent diminished influx of hematogenous cells.Investigations revealed no differences in transcription of tumor necrosis factor-α and astrogliosis 12 and 36 hours after onset of ischemia.These novel results help to understand post ischemic, inflammatory mechanisms and might give further arguments towards therapeutical interventions by modulation of MCP-1 expression in post stroke inflammation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of Münster, Münster, Germany. strecker.jan@gmx.de

ABSTRACT
Monocyte chemoattractant protein-1 (MCP-1), a chemokine secreted by neurons and astrocytes following stroke is known to aggravate ischemia-related damage. Previous studies revealed that MCP-1-deficient mice develop smaller infarcts and have an improved neurological outcome, whereas mice overexpressing MCP-1 show worsened brain damage and impaired neurological function. The aim of the present study was to elucidate the molecular background of the enhanced recovery in MCP-1-deficient mice after stroke. For this purpose, we (1) performed expression analyses on crucial post-stroke related inflammatory genes in MCP-1-deficient mice compared to wildtype controls, (2) analyzed a possible impact of MCP-1 on astrocyte activation (3) investigated the cellular origin of respective inflammatory cytokines and (4) analyzed the impact of MCP-1 secretion on the migration of both neutrophil granulocytes and T-cells. Here we report that MCP-1-deficiency leads to a shift towards a less inflammatory state following experimental occlusion of the middle cerebral artery including an impaired induction of interleukin-6, interleukin-1β and granulocyte-colony stimulating factor expression as well as a subsequent diminished influx of hematogenous cells. Additionally, MCP-1-deficient mice developed smaller infarcts 36 hours after experimental stroke. Investigations revealed no differences in transcription of tumor necrosis factor-α and astrogliosis 12 and 36 hours after onset of ischemia. These novel results help to understand post ischemic, inflammatory mechanisms and might give further arguments towards therapeutical interventions by modulation of MCP-1 expression in post stroke inflammation.

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Immunofluorescence-staining within the ipsilateral cerebral cortex revealed astrocytes (A−C) and neurons (E−G) as the main source of interleukin-6-synthesis.In MCP-1-deficient mice, IL-6 signal pattern also showed double positive signals colocalized with astroctyes (D) and neurons (H) but with reduced intensity indicating a diminished IL-6 concentration on the translational level. Expression of IL-1β within the ischemic core was mainly restricted to neurons (I−K) in both wildtype and MCP-1-deficient mice, the latter displaying a reduced IL-1β-fluorescence intensity confirming the results of the expression analyses (L). Shrinked cell morphology indicates apoptotic processes within the infarcted core. Double immunofluorescence-staining also showed co-localization of G-CSF-positive cells and astrocytes (M−O), microglia (Q−S) and occasionally, neurons (T). In MCP-1-deficient mice astrocytic G-CSF secretion was clearly diminished, visualized by the reduction of fluorescence signal intensity reveals (P). Scale bar = 25 µm.
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pone-0025863-g004: Immunofluorescence-staining within the ipsilateral cerebral cortex revealed astrocytes (A−C) and neurons (E−G) as the main source of interleukin-6-synthesis.In MCP-1-deficient mice, IL-6 signal pattern also showed double positive signals colocalized with astroctyes (D) and neurons (H) but with reduced intensity indicating a diminished IL-6 concentration on the translational level. Expression of IL-1β within the ischemic core was mainly restricted to neurons (I−K) in both wildtype and MCP-1-deficient mice, the latter displaying a reduced IL-1β-fluorescence intensity confirming the results of the expression analyses (L). Shrinked cell morphology indicates apoptotic processes within the infarcted core. Double immunofluorescence-staining also showed co-localization of G-CSF-positive cells and astrocytes (M−O), microglia (Q−S) and occasionally, neurons (T). In MCP-1-deficient mice astrocytic G-CSF secretion was clearly diminished, visualized by the reduction of fluorescence signal intensity reveals (P). Scale bar = 25 µm.

Mentions: To investigate whether MCP-1 expression is associated with an induction of IL-6, we measured and compared IL-6-mRNA concentrations within three different brain areas of wildtype and MCP-1-deficient mice. Twelve hours after onset of MCAO a considerable induction of IL-6 was found within the cortex and infarct core of wildtype animals. In contrast, MCP-1-deficient animals showed a severely impaired IL-6 expression in both ipsilateral cortex (p<0.01) and infarcted core (p<0.01, Fig.3 A, B, C). Thirty-six hours after MCAO IL-6 expression patterns showed no significant change compared to the earlier time-point in both wildtype and MCP-1-deficient animals. IL-6 expression remained elevated in both wildtype cerebral cortex (p<0.05) and infarcted core, whereas samples of MCP-1-deficient mice showed almost no IL-6 expression in all investigated areas (Fig.3 D, E, F-). Next, we performed dual immunohistochemical analysis of IL-6 and cell markers for neurons, astrocytes, microglia/macrophages and neutrophil granulocytes. Double fluorescence analysis revealed colocalizations of IL-6 with GFAP and NeuN within the ipsilateral cerebral cortex and infarcted core indicating that IL-6 is particularly expressed by astrocytes and neurons after cerebral ischemia (Fig.4 A, B, C, D, E, F, G, H). In concordance with the results of mRNA expression we found significantly reduced cell numbers positive for IL-6 in MCP-1 deficient mice within the ipsilateral cortex and the ischemic core at both investigated time-points (Fig.5). Regarding the contralateral hemisphere, IL-6 positive cells increased in wildtype animals 12 hours after MCAO, whereas cell counts remained on the level of the control group in MCP-1-deficient mice at both time-points.


Monocyte chemoattractant protein-1-deficiency impairs the expression of IL-6, IL-1β and G-CSF after transient focal ischemia in mice.

Strecker JK, Minnerup J, Gess B, Ringelstein EB, Schäbitz WR, Schilling M - PLoS ONE (2011)

Immunofluorescence-staining within the ipsilateral cerebral cortex revealed astrocytes (A−C) and neurons (E−G) as the main source of interleukin-6-synthesis.In MCP-1-deficient mice, IL-6 signal pattern also showed double positive signals colocalized with astroctyes (D) and neurons (H) but with reduced intensity indicating a diminished IL-6 concentration on the translational level. Expression of IL-1β within the ischemic core was mainly restricted to neurons (I−K) in both wildtype and MCP-1-deficient mice, the latter displaying a reduced IL-1β-fluorescence intensity confirming the results of the expression analyses (L). Shrinked cell morphology indicates apoptotic processes within the infarcted core. Double immunofluorescence-staining also showed co-localization of G-CSF-positive cells and astrocytes (M−O), microglia (Q−S) and occasionally, neurons (T). In MCP-1-deficient mice astrocytic G-CSF secretion was clearly diminished, visualized by the reduction of fluorescence signal intensity reveals (P). Scale bar = 25 µm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198727&req=5

pone-0025863-g004: Immunofluorescence-staining within the ipsilateral cerebral cortex revealed astrocytes (A−C) and neurons (E−G) as the main source of interleukin-6-synthesis.In MCP-1-deficient mice, IL-6 signal pattern also showed double positive signals colocalized with astroctyes (D) and neurons (H) but with reduced intensity indicating a diminished IL-6 concentration on the translational level. Expression of IL-1β within the ischemic core was mainly restricted to neurons (I−K) in both wildtype and MCP-1-deficient mice, the latter displaying a reduced IL-1β-fluorescence intensity confirming the results of the expression analyses (L). Shrinked cell morphology indicates apoptotic processes within the infarcted core. Double immunofluorescence-staining also showed co-localization of G-CSF-positive cells and astrocytes (M−O), microglia (Q−S) and occasionally, neurons (T). In MCP-1-deficient mice astrocytic G-CSF secretion was clearly diminished, visualized by the reduction of fluorescence signal intensity reveals (P). Scale bar = 25 µm.
Mentions: To investigate whether MCP-1 expression is associated with an induction of IL-6, we measured and compared IL-6-mRNA concentrations within three different brain areas of wildtype and MCP-1-deficient mice. Twelve hours after onset of MCAO a considerable induction of IL-6 was found within the cortex and infarct core of wildtype animals. In contrast, MCP-1-deficient animals showed a severely impaired IL-6 expression in both ipsilateral cortex (p<0.01) and infarcted core (p<0.01, Fig.3 A, B, C). Thirty-six hours after MCAO IL-6 expression patterns showed no significant change compared to the earlier time-point in both wildtype and MCP-1-deficient animals. IL-6 expression remained elevated in both wildtype cerebral cortex (p<0.05) and infarcted core, whereas samples of MCP-1-deficient mice showed almost no IL-6 expression in all investigated areas (Fig.3 D, E, F-). Next, we performed dual immunohistochemical analysis of IL-6 and cell markers for neurons, astrocytes, microglia/macrophages and neutrophil granulocytes. Double fluorescence analysis revealed colocalizations of IL-6 with GFAP and NeuN within the ipsilateral cerebral cortex and infarcted core indicating that IL-6 is particularly expressed by astrocytes and neurons after cerebral ischemia (Fig.4 A, B, C, D, E, F, G, H). In concordance with the results of mRNA expression we found significantly reduced cell numbers positive for IL-6 in MCP-1 deficient mice within the ipsilateral cortex and the ischemic core at both investigated time-points (Fig.5). Regarding the contralateral hemisphere, IL-6 positive cells increased in wildtype animals 12 hours after MCAO, whereas cell counts remained on the level of the control group in MCP-1-deficient mice at both time-points.

Bottom Line: Here we report that MCP-1-deficiency leads to a shift towards a less inflammatory state following experimental occlusion of the middle cerebral artery including an impaired induction of interleukin-6, interleukin-1β and granulocyte-colony stimulating factor expression as well as a subsequent diminished influx of hematogenous cells.Investigations revealed no differences in transcription of tumor necrosis factor-α and astrogliosis 12 and 36 hours after onset of ischemia.These novel results help to understand post ischemic, inflammatory mechanisms and might give further arguments towards therapeutical interventions by modulation of MCP-1 expression in post stroke inflammation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of Münster, Münster, Germany. strecker.jan@gmx.de

ABSTRACT
Monocyte chemoattractant protein-1 (MCP-1), a chemokine secreted by neurons and astrocytes following stroke is known to aggravate ischemia-related damage. Previous studies revealed that MCP-1-deficient mice develop smaller infarcts and have an improved neurological outcome, whereas mice overexpressing MCP-1 show worsened brain damage and impaired neurological function. The aim of the present study was to elucidate the molecular background of the enhanced recovery in MCP-1-deficient mice after stroke. For this purpose, we (1) performed expression analyses on crucial post-stroke related inflammatory genes in MCP-1-deficient mice compared to wildtype controls, (2) analyzed a possible impact of MCP-1 on astrocyte activation (3) investigated the cellular origin of respective inflammatory cytokines and (4) analyzed the impact of MCP-1 secretion on the migration of both neutrophil granulocytes and T-cells. Here we report that MCP-1-deficiency leads to a shift towards a less inflammatory state following experimental occlusion of the middle cerebral artery including an impaired induction of interleukin-6, interleukin-1β and granulocyte-colony stimulating factor expression as well as a subsequent diminished influx of hematogenous cells. Additionally, MCP-1-deficient mice developed smaller infarcts 36 hours after experimental stroke. Investigations revealed no differences in transcription of tumor necrosis factor-α and astrogliosis 12 and 36 hours after onset of ischemia. These novel results help to understand post ischemic, inflammatory mechanisms and might give further arguments towards therapeutical interventions by modulation of MCP-1 expression in post stroke inflammation.

Show MeSH
Related in: MedlinePlus