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Combustion smoke-induced inflammation in the olfactory bulb of adult rats.

Zou YY, Yuan Y, Kan EM, Lu J, Ling EA - J Neuroinflammation (2014)

Bottom Line: The results showed a significant increase in VEGF, iNOS, eNOS, nNOS, NKCC1, and GFAP expression in the bulb tissues, with corresponding increases in inflammatory cytokines and chemokines after smoke inhalation.Concurrent to this was a drastic increase in AQP4 expression and RITC permeability.This was coupled with a significant reduction in incidence of TUNEL + cells that was not altered with administration of L-NG-nitroarginine methyl ester (L-NAME).

View Article: PubMed Central - PubMed

ABSTRACT

Background: The damaging effect of combustion smoke inhalation on the lung is widely reported but information on its effects on the olfactory bulb is lacking. This study sought to determine the effects of smoke inhalation on the olfactory bulb, whose afferent input neurons in the nasal mucosa are directly exposed to external stimuli, such as smoke.

Methods: Adult male Sprague-Dawley rats were subjected to combustion smoke inhalation and sacrificed at different time points. Changes in olfactory bulb proteins including vascular endothelial growth factor (VEGF), inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), neuronal nitric oxide synthase (nNOS), Na+-K+-Cl- cotransporter 1 (NKCC1), glial fibrillary acidic protein (GFAP), and aquaporin-4 (AQP4) were evaluated by Western blot analysis. In addition, ELISA was conducted for cytokine and chemokine levels, and double immunofluorescence labeling was carried out for GFAP/VEGF, GFAP/AQP4, NeuN/nNOS, GFAP/NKCC1, NeuN/NKCC1, GFAP/Rhodamine isothiocyanate (RITC), and transferase dUTP nick end labeling (TUNEL). Aminoguanidine was administered to determine the effects of iNOS inhibition on the targets probed after smoke inhalation.

Results: The results showed a significant increase in VEGF, iNOS, eNOS, nNOS, NKCC1, and GFAP expression in the bulb tissues, with corresponding increases in inflammatory cytokines and chemokines after smoke inhalation. Concurrent to this was a drastic increase in AQP4 expression and RITC permeability. Aminoguanidine administration decreased the expression of iNOS and RITC extravasation after smoke inhalation. This was coupled with a significant reduction in incidence of TUNEL + cells that was not altered with administration of L-NG-nitroarginine methyl ester (L-NAME).

Conclusions: These findings suggest that the upregulation of iNOS in response to smoke inhalation plays a major role in the olfactory bulb inflammatory pathophysiology, along with a concomitant increase in pro-inflammatory molecules, vascular permeability, and edema. Overall, these findings indicate that the olfactory bulb is vulnerable to smoke inhalation.

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VEGF (red) and GFAP (green) immunopositive astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer in the olfactory bulb of the C group (A-C), SI + S group at 24 and 72 h (D-I), and SI + AG group at 72 h (J-L). In the control, moderate VEGF immunofluorescence is detected in GFAP-labeled astrocytes. In the SI + S group, VEGF immunofluorescence is evidently enhanced in hypertrophic astrocytes at 24 and 72 h (D-I), notably at 24 h (E). Colocalized expression of VEGF with GFAP immunoreactive astrocytes (arrows) and the lumen of blood vessels (asterisks) can be seen in C, FI, and L. Immunoreactivity for both VEGF and GFAP is attenuated in the SI + AG group at 72 h, as compared with the SI + S group at corresponding time point (J-L). Scale bar =20.0 μm.
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Fig5: VEGF (red) and GFAP (green) immunopositive astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer in the olfactory bulb of the C group (A-C), SI + S group at 24 and 72 h (D-I), and SI + AG group at 72 h (J-L). In the control, moderate VEGF immunofluorescence is detected in GFAP-labeled astrocytes. In the SI + S group, VEGF immunofluorescence is evidently enhanced in hypertrophic astrocytes at 24 and 72 h (D-I), notably at 24 h (E). Colocalized expression of VEGF with GFAP immunoreactive astrocytes (arrows) and the lumen of blood vessels (asterisks) can be seen in C, FI, and L. Immunoreactivity for both VEGF and GFAP is attenuated in the SI + AG group at 72 h, as compared with the SI + S group at corresponding time point (J-L). Scale bar =20.0 μm.

Mentions: Expression of VEGF was observed in widely distributed branched cells identified as GFAP-expressing astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer (Figure 5). In the C group (Figure 5A,B), GFAP and VEGF colocalization was identifiable at moderate levels (Figure 5C). In the SI + S group at 24 and 72 h, both GFAP and VEGF immunoreactivity was markedly enhanced (Figure 5D-I), being most conspicuous at 24 h. The vascular profiles appeared dilated with associated GFAP + VEGF-positive cell processes (Figure 5F,I), notably at 24 h. Both GFAP and VEGF immunofluorescence was moderately attenuated at 72 h in the SI + AG group (Figure 5J-L).Figure 5


Combustion smoke-induced inflammation in the olfactory bulb of adult rats.

Zou YY, Yuan Y, Kan EM, Lu J, Ling EA - J Neuroinflammation (2014)

VEGF (red) and GFAP (green) immunopositive astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer in the olfactory bulb of the C group (A-C), SI + S group at 24 and 72 h (D-I), and SI + AG group at 72 h (J-L). In the control, moderate VEGF immunofluorescence is detected in GFAP-labeled astrocytes. In the SI + S group, VEGF immunofluorescence is evidently enhanced in hypertrophic astrocytes at 24 and 72 h (D-I), notably at 24 h (E). Colocalized expression of VEGF with GFAP immunoreactive astrocytes (arrows) and the lumen of blood vessels (asterisks) can be seen in C, FI, and L. Immunoreactivity for both VEGF and GFAP is attenuated in the SI + AG group at 72 h, as compared with the SI + S group at corresponding time point (J-L). Scale bar =20.0 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4197300&req=5

Fig5: VEGF (red) and GFAP (green) immunopositive astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer in the olfactory bulb of the C group (A-C), SI + S group at 24 and 72 h (D-I), and SI + AG group at 72 h (J-L). In the control, moderate VEGF immunofluorescence is detected in GFAP-labeled astrocytes. In the SI + S group, VEGF immunofluorescence is evidently enhanced in hypertrophic astrocytes at 24 and 72 h (D-I), notably at 24 h (E). Colocalized expression of VEGF with GFAP immunoreactive astrocytes (arrows) and the lumen of blood vessels (asterisks) can be seen in C, FI, and L. Immunoreactivity for both VEGF and GFAP is attenuated in the SI + AG group at 72 h, as compared with the SI + S group at corresponding time point (J-L). Scale bar =20.0 μm.
Mentions: Expression of VEGF was observed in widely distributed branched cells identified as GFAP-expressing astrocytes in the external plexiform layer, mitral cell layer, and internal plexiform layer (Figure 5). In the C group (Figure 5A,B), GFAP and VEGF colocalization was identifiable at moderate levels (Figure 5C). In the SI + S group at 24 and 72 h, both GFAP and VEGF immunoreactivity was markedly enhanced (Figure 5D-I), being most conspicuous at 24 h. The vascular profiles appeared dilated with associated GFAP + VEGF-positive cell processes (Figure 5F,I), notably at 24 h. Both GFAP and VEGF immunofluorescence was moderately attenuated at 72 h in the SI + AG group (Figure 5J-L).Figure 5

Bottom Line: The results showed a significant increase in VEGF, iNOS, eNOS, nNOS, NKCC1, and GFAP expression in the bulb tissues, with corresponding increases in inflammatory cytokines and chemokines after smoke inhalation.Concurrent to this was a drastic increase in AQP4 expression and RITC permeability.This was coupled with a significant reduction in incidence of TUNEL + cells that was not altered with administration of L-NG-nitroarginine methyl ester (L-NAME).

View Article: PubMed Central - PubMed

ABSTRACT

Background: The damaging effect of combustion smoke inhalation on the lung is widely reported but information on its effects on the olfactory bulb is lacking. This study sought to determine the effects of smoke inhalation on the olfactory bulb, whose afferent input neurons in the nasal mucosa are directly exposed to external stimuli, such as smoke.

Methods: Adult male Sprague-Dawley rats were subjected to combustion smoke inhalation and sacrificed at different time points. Changes in olfactory bulb proteins including vascular endothelial growth factor (VEGF), inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), neuronal nitric oxide synthase (nNOS), Na+-K+-Cl- cotransporter 1 (NKCC1), glial fibrillary acidic protein (GFAP), and aquaporin-4 (AQP4) were evaluated by Western blot analysis. In addition, ELISA was conducted for cytokine and chemokine levels, and double immunofluorescence labeling was carried out for GFAP/VEGF, GFAP/AQP4, NeuN/nNOS, GFAP/NKCC1, NeuN/NKCC1, GFAP/Rhodamine isothiocyanate (RITC), and transferase dUTP nick end labeling (TUNEL). Aminoguanidine was administered to determine the effects of iNOS inhibition on the targets probed after smoke inhalation.

Results: The results showed a significant increase in VEGF, iNOS, eNOS, nNOS, NKCC1, and GFAP expression in the bulb tissues, with corresponding increases in inflammatory cytokines and chemokines after smoke inhalation. Concurrent to this was a drastic increase in AQP4 expression and RITC permeability. Aminoguanidine administration decreased the expression of iNOS and RITC extravasation after smoke inhalation. This was coupled with a significant reduction in incidence of TUNEL + cells that was not altered with administration of L-NG-nitroarginine methyl ester (L-NAME).

Conclusions: These findings suggest that the upregulation of iNOS in response to smoke inhalation plays a major role in the olfactory bulb inflammatory pathophysiology, along with a concomitant increase in pro-inflammatory molecules, vascular permeability, and edema. Overall, these findings indicate that the olfactory bulb is vulnerable to smoke inhalation.

Show MeSH
Related in: MedlinePlus