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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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Immunoexpression of nNOS (red) and NeuN (green) in the olfactory bulb of the C group (A-C), the SI + S group at 24 and 72 h (D-I), and the SI + AG group at 72 h (J-L). In the normal bulb, nNOS expression is absent (A-C); however, in the SI + S group, it is induced in some neurons at different time points (24 and 72 h, D-I); the incidence of labeled cells is higher at 24 h (E). Aminoguanidine administration did not obviously alter the immunofluorescence at 72 h (J-L). Scale bar =20.0 μm.
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Fig7: Immunoexpression of nNOS (red) and NeuN (green) in the olfactory bulb of the C group (A-C), the SI + S group at 24 and 72 h (D-I), and the SI + AG group at 72 h (J-L). In the normal bulb, nNOS expression is absent (A-C); however, in the SI + S group, it is induced in some neurons at different time points (24 and 72 h, D-I); the incidence of labeled cells is higher at 24 h (E). Aminoguanidine administration did not obviously alter the immunofluorescence at 72 h (J-L). Scale bar =20.0 μm.

Mentions: Expression of nNOS was observed in neurons in the external plexiform layer, mitral cell layer, internal plexiform layer, and granule cell layer (Figure 7). The nNOS-expressing cells were identified as neurons with colocalization with NeuN by double immunofluorescence labeling (Figure 7). In the C group (Figure 7A,B), nNOS and NeuN colocalization could hardly be detected (Figure 7C). In the SI + S group at 24 and 72 h, nNOS intense immunoreactivity was detected in some NeuN-positive neurons (Figure 7D-I) with a higher incidence at 24 h. Intense nNOS immunoreactivity was localized in the nucleus, whose outlined is well defined (Figure 7F,I,L). Aminoguanidine did not obviously suppress the immunofluorescence of nNOS at 72 h (Figure 7J-L).Figure 7


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

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

Immunoexpression of nNOS (red) and NeuN (green) in the olfactory bulb of the C group (A-C), the SI + S group at 24 and 72 h (D-I), and the SI + AG group at 72 h (J-L). In the normal bulb, nNOS expression is absent (A-C); however, in the SI + S group, it is induced in some neurons at different time points (24 and 72 h, D-I); the incidence of labeled cells is higher at 24 h (E). Aminoguanidine administration did not obviously alter the immunofluorescence at 72 h (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

Fig7: Immunoexpression of nNOS (red) and NeuN (green) in the olfactory bulb of the C group (A-C), the SI + S group at 24 and 72 h (D-I), and the SI + AG group at 72 h (J-L). In the normal bulb, nNOS expression is absent (A-C); however, in the SI + S group, it is induced in some neurons at different time points (24 and 72 h, D-I); the incidence of labeled cells is higher at 24 h (E). Aminoguanidine administration did not obviously alter the immunofluorescence at 72 h (J-L). Scale bar =20.0 μm.
Mentions: Expression of nNOS was observed in neurons in the external plexiform layer, mitral cell layer, internal plexiform layer, and granule cell layer (Figure 7). The nNOS-expressing cells were identified as neurons with colocalization with NeuN by double immunofluorescence labeling (Figure 7). In the C group (Figure 7A,B), nNOS and NeuN colocalization could hardly be detected (Figure 7C). In the SI + S group at 24 and 72 h, nNOS intense immunoreactivity was detected in some NeuN-positive neurons (Figure 7D-I) with a higher incidence at 24 h. Intense nNOS immunoreactivity was localized in the nucleus, whose outlined is well defined (Figure 7F,I,L). Aminoguanidine did not obviously suppress the immunofluorescence of nNOS at 72 h (Figure 7J-L).Figure 7

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