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Activation of the inflammatory transcription factor nuclear factor interleukin-6 during inflammatory and psychological stress in the brain.

Fuchs F, Damm J, Gerstberger R, Roth J, Rummel C - J Neuroinflammation (2013)

Bottom Line: In vitro NF-IL6-activation was associated with induction and secretion of TNFα in folliculostellate cells, which could be antagonized by the JAK-STAT-inhibitor AG490.Both stressors were associated with time-dependent activation of NF-IL6 in different cell types of the brain and the pituitary.Moreover, while NF-IL6-IR was partially linked to STAT3 and NFκB activation, TNFα production, and ACTH-IR after LPS stimulation; this was not the case after exposure to a novel environment, suggesting distinct underlying signaling pathways.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary-Physiology and -Biochemistry, Justus-Liebig-University Giessen, Frankfurter Strasse 100, Giessen D-35392, Germany. Christoph.D.Rummel@vetmed.uni-giessen.de.

ABSTRACT

Background: The transcription factor nuclear factor interleukin 6 (NF-IL6) is known to be activated by various inflammatory stimuli in the brain. Interestingly, we recently detected NF-IL6-activation within the hypothalamus-pituitary-adrenal (HPA)-axis of rats after systemic lipopolysaccharide (LPS)-injection. Thus, the aim of the present study was to investigate whether NF-IL6 is activated during either, inflammatory, or psychological stress in the rat brain.

Methods: Rats were challenged with either the inflammatory stimulus LPS (100 μg/kg, i.p.) or exposed to a novel environment. Core body temperature (Tb) and motor activity were monitored using telemetry, animals were killed at different time points, brains and blood removed, and primary cell cultures of the anterior pituitary lobe (AL) were investigated. Analyses were performed using immunohistochemistry, RT-PCR, and cytokine-specific bioassays.

Results: Stress stimulation by a novel environment increased NF-IL6-immunoreactivity (IR) in the pituitary's perivascular macrophages and hypothalamic paraventricular cells and a rise in Tb lasting approximately 2 h. LPS stimulation lead to NF-IL6-IR in several additional cell types including ACTH-IR-positive corticotrope cells in vivo and in vitro. Two other proinflammatory transcription factors, namely signal transducer and activator of transcription (STAT)3 and NFκB, were significantly activated and partially colocalized with NF-IL6-IR in cells of the AL only after LPS-stimulation, but not following psychological stress. In vitro NF-IL6-activation was associated with induction and secretion of TNFα in folliculostellate cells, which could be antagonized by the JAK-STAT-inhibitor AG490.

Conclusions: We revealed, for the first time, that NF-IL6 activation occurs not only during inflammatory LPS stimulation, but also during psychological stress, that is, a novel environment. Both stressors were associated with time-dependent activation of NF-IL6 in different cell types of the brain and the pituitary. Moreover, while NF-IL6-IR was partially linked to STAT3 and NFκB activation, TNFα production, and ACTH-IR after LPS stimulation; this was not the case after exposure to a novel environment, suggesting distinct underlying signaling pathways. Overall, NF-IL6 can be used as a broad activation marker in the brain and might be of interest for therapeutic approaches not only during inflammatory but also psychological stress.

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Novel environment stress (stress) induced a significant increase in nuclear NF-IL6-IR in the pituitary of rats. (A, B) Some nuclear (DAPI, blue) NF-IL6-IR (red) can be observed in unstimulated control animals. (C-F) After novel environment-stress was introduced, nuclear NF-IL6-IR peaked (D, E) 60 to 90 min later and (G) declined to control levels at 240 min. GFAP detection (green) was used to better visualize the posterior pituitary lobe. (H) The schematic overview clearly depicts the substructure of microphotographs, containing all pituitary lobes (anterior lobe (AL), intermediate lobe (IL), and posterior lobe (PL)). Brightness, contrast, and color balance were adjusted for better representation of the actual data. The scale bar in A represents 100 μm (applies to A-G).
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Figure 2: Novel environment stress (stress) induced a significant increase in nuclear NF-IL6-IR in the pituitary of rats. (A, B) Some nuclear (DAPI, blue) NF-IL6-IR (red) can be observed in unstimulated control animals. (C-F) After novel environment-stress was introduced, nuclear NF-IL6-IR peaked (D, E) 60 to 90 min later and (G) declined to control levels at 240 min. GFAP detection (green) was used to better visualize the posterior pituitary lobe. (H) The schematic overview clearly depicts the substructure of microphotographs, containing all pituitary lobes (anterior lobe (AL), intermediate lobe (IL), and posterior lobe (PL)). Brightness, contrast, and color balance were adjusted for better representation of the actual data. The scale bar in A represents 100 μm (applies to A-G).

Mentions: These effects were accompanied by increased novel environment stress-induced nuclear NF-IL6-IR that peaked between 90 and 120 min in the PVN (Additional file 1E,F) and between 60 and 90 min in the pituitary (Figure 2D,E) but was unchanged in other brain structures such as the SFO (Table 2). NF-IL6-IR appeared to be particularly strong lining the intermediate lobe and in the anterior pituitary lobe compared to untreated controls (Figure 2A-G). A semi-quantitative five-point scale evaluation of two sections per rat for three or six (for 60 and 240 min) animals per group confirmed these qualitative observations (Table 2). In this stress paradigm no NF-IL6-positive corticotropes (Figure 3A,B), pituicytes (Figure 3C,D, GFAP), or endothelial cells (Figure 3E,F, VWF) could be observed. Nonetheless, some NF-IL6-IR positive cells co-localized with CD163-stained perivascular macrophages in both the anterior and the posterior pituitary lobe in stressed (Figure 3H) but not control animals (Figure 3G). Neither increased activation of STAT3 nor NFκB could be observed in brains and pituitaries of these animals (data not shown).


Activation of the inflammatory transcription factor nuclear factor interleukin-6 during inflammatory and psychological stress in the brain.

Fuchs F, Damm J, Gerstberger R, Roth J, Rummel C - J Neuroinflammation (2013)

Novel environment stress (stress) induced a significant increase in nuclear NF-IL6-IR in the pituitary of rats. (A, B) Some nuclear (DAPI, blue) NF-IL6-IR (red) can be observed in unstimulated control animals. (C-F) After novel environment-stress was introduced, nuclear NF-IL6-IR peaked (D, E) 60 to 90 min later and (G) declined to control levels at 240 min. GFAP detection (green) was used to better visualize the posterior pituitary lobe. (H) The schematic overview clearly depicts the substructure of microphotographs, containing all pituitary lobes (anterior lobe (AL), intermediate lobe (IL), and posterior lobe (PL)). Brightness, contrast, and color balance were adjusted for better representation of the actual data. The scale bar in A represents 100 μm (applies to A-G).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4222273&req=5

Figure 2: Novel environment stress (stress) induced a significant increase in nuclear NF-IL6-IR in the pituitary of rats. (A, B) Some nuclear (DAPI, blue) NF-IL6-IR (red) can be observed in unstimulated control animals. (C-F) After novel environment-stress was introduced, nuclear NF-IL6-IR peaked (D, E) 60 to 90 min later and (G) declined to control levels at 240 min. GFAP detection (green) was used to better visualize the posterior pituitary lobe. (H) The schematic overview clearly depicts the substructure of microphotographs, containing all pituitary lobes (anterior lobe (AL), intermediate lobe (IL), and posterior lobe (PL)). Brightness, contrast, and color balance were adjusted for better representation of the actual data. The scale bar in A represents 100 μm (applies to A-G).
Mentions: These effects were accompanied by increased novel environment stress-induced nuclear NF-IL6-IR that peaked between 90 and 120 min in the PVN (Additional file 1E,F) and between 60 and 90 min in the pituitary (Figure 2D,E) but was unchanged in other brain structures such as the SFO (Table 2). NF-IL6-IR appeared to be particularly strong lining the intermediate lobe and in the anterior pituitary lobe compared to untreated controls (Figure 2A-G). A semi-quantitative five-point scale evaluation of two sections per rat for three or six (for 60 and 240 min) animals per group confirmed these qualitative observations (Table 2). In this stress paradigm no NF-IL6-positive corticotropes (Figure 3A,B), pituicytes (Figure 3C,D, GFAP), or endothelial cells (Figure 3E,F, VWF) could be observed. Nonetheless, some NF-IL6-IR positive cells co-localized with CD163-stained perivascular macrophages in both the anterior and the posterior pituitary lobe in stressed (Figure 3H) but not control animals (Figure 3G). Neither increased activation of STAT3 nor NFκB could be observed in brains and pituitaries of these animals (data not shown).

Bottom Line: In vitro NF-IL6-activation was associated with induction and secretion of TNFα in folliculostellate cells, which could be antagonized by the JAK-STAT-inhibitor AG490.Both stressors were associated with time-dependent activation of NF-IL6 in different cell types of the brain and the pituitary.Moreover, while NF-IL6-IR was partially linked to STAT3 and NFκB activation, TNFα production, and ACTH-IR after LPS stimulation; this was not the case after exposure to a novel environment, suggesting distinct underlying signaling pathways.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary-Physiology and -Biochemistry, Justus-Liebig-University Giessen, Frankfurter Strasse 100, Giessen D-35392, Germany. Christoph.D.Rummel@vetmed.uni-giessen.de.

ABSTRACT

Background: The transcription factor nuclear factor interleukin 6 (NF-IL6) is known to be activated by various inflammatory stimuli in the brain. Interestingly, we recently detected NF-IL6-activation within the hypothalamus-pituitary-adrenal (HPA)-axis of rats after systemic lipopolysaccharide (LPS)-injection. Thus, the aim of the present study was to investigate whether NF-IL6 is activated during either, inflammatory, or psychological stress in the rat brain.

Methods: Rats were challenged with either the inflammatory stimulus LPS (100 μg/kg, i.p.) or exposed to a novel environment. Core body temperature (Tb) and motor activity were monitored using telemetry, animals were killed at different time points, brains and blood removed, and primary cell cultures of the anterior pituitary lobe (AL) were investigated. Analyses were performed using immunohistochemistry, RT-PCR, and cytokine-specific bioassays.

Results: Stress stimulation by a novel environment increased NF-IL6-immunoreactivity (IR) in the pituitary's perivascular macrophages and hypothalamic paraventricular cells and a rise in Tb lasting approximately 2 h. LPS stimulation lead to NF-IL6-IR in several additional cell types including ACTH-IR-positive corticotrope cells in vivo and in vitro. Two other proinflammatory transcription factors, namely signal transducer and activator of transcription (STAT)3 and NFκB, were significantly activated and partially colocalized with NF-IL6-IR in cells of the AL only after LPS-stimulation, but not following psychological stress. In vitro NF-IL6-activation was associated with induction and secretion of TNFα in folliculostellate cells, which could be antagonized by the JAK-STAT-inhibitor AG490.

Conclusions: We revealed, for the first time, that NF-IL6 activation occurs not only during inflammatory LPS stimulation, but also during psychological stress, that is, a novel environment. Both stressors were associated with time-dependent activation of NF-IL6 in different cell types of the brain and the pituitary. Moreover, while NF-IL6-IR was partially linked to STAT3 and NFκB activation, TNFα production, and ACTH-IR after LPS stimulation; this was not the case after exposure to a novel environment, suggesting distinct underlying signaling pathways. Overall, NF-IL6 can be used as a broad activation marker in the brain and might be of interest for therapeutic approaches not only during inflammatory but also psychological stress.

Show MeSH
Related in: MedlinePlus