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Hypertonic saline alleviates experimentally induced cerebral oedema through suppression of vascular endothelial growth factor and its receptor VEGFR2 expression in astrocytes - PubMed

  • ️Fri Jan 01 2016

Hypertonic saline alleviates experimentally induced cerebral oedema through suppression of vascular endothelial growth factor and its receptor VEGFR2 expression in astrocytes

Linqiang Huang et al. BMC Neurosci. 2016.

Abstract

Background: Cerebral oedema is closely related to the permeability of blood-brain barrier, vascular endothelial growth factor (VEGF) and its receptor vascular endothelial growth factor receptor 2 (VEGFR2) all of which are important blood-brain barrier (BBB) permeability regulatory factors. Zonula occludens 1 (ZO-1) and claudin-5 are also the key components of BBB. Hypertonic saline is widely used to alleviate cerebral oedema. This study aimed to explore the possible mechanisms underlying hypertonic saline that ameliorates cerebral oedema effectively.

Methods: Middle cerebral artery occlusion (MCAO) model in Sprague-Dawley (SD) rats and of oxygen-glucose deprivation model in primary astrocytes were used in this study. The brain water content (BWC) was used to assess the effect of 10 % HS on cerebral oedema. The assessment of Evans blue (EB) extravasation was performed to evaluate the protective effect of 10 % HS on blood-brain barrier. The quantification of VEGF, VEGFR2, ZO-1 and claudin-5 was used to illustrate the mechanism of 10 % HS ameliorating cerebral oedema.

Results: BWC was analysed by wet-to-dry ratios in the ischemic hemisphere of SD rats; it was significantly decreased after 10 % HS treatment (P < 0.05). We also investigated the blood-brain barrier protective effect by 10 % HS which reduced EB extravasation effectively in the peri-ischemic brain tissue. In parallel to the above notably at 24 h following MCAO, mRNA and protein expression of VEGF and VEGFR2 in the peri-ischemic brain tissue was down-regulated after 10 % HS treatment (P < 0.05). Along with this, in vitro studies showed increased VEGF and VEGFR2 mRNA and protein expression in primary astrocytes under hypoxic condition (P < 0.05), but it was suppressed after HS treatment (P < 0.05). In addition, HS inhibited the down-regulation of ZO-1, claudin-5 effectively.

Conclusions: The results suggest that 10 % HS could alleviate cerebral oedema possibly through reducing the ischemia induced BBB permeability as a consequence of inhibiting VEGF-VEGFR2-mediated down-regulation of ZO-1, claudin-5.

Keywords: Astrocyte; Cerebral oedema; Claudin-5; Hypertonic saline; Vascular endothelial growth factor; ZO-1.

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Figures

Fig. 1
Fig. 1

Assessment of BWC and Evans blue. Bar graph A the neurological score is not significantly different between ischemia group and 10 % HS group at 2 h after MCAO based on Zea-longa scores (ns: P > 0.05). Bar graph B the percentage of BWC in 10 % HS group was significantly decreased as compared with corresponding ischemia groups at 6, 12 and 24 h after MCAO (*P < 0.05). Bar graph C 10 % HS could reduce Evans blue extravasation effectively when compared with corresponding ischemia groups at 6, 12 and 24 h after MCAO (*P < 0.05). #indicates compared with sham group, P < 0.05. D, E and F Evans blue extravasation in each group at 24 h after MCAO. The values are presented as the mean ± SD

Fig. 2
Fig. 2

VEGF mRNA and protein expression in the peri-ischemic brain tissue in each group. A VEGF (45 kDa) and GAPDH (37 kDa) immunoreactive bands, respectively. Bar graph B depicts significant changes in the optical density of VEGF expression when compared with the corresponding ischemia groups (*P < 0.05). Bar graph C the fold change in VEGF mRNA expression. When compared with ischemia group at 6, 12 and 24 h after MCAO, VEGF mRNA expression in corresponding 10 % HS group is evidently reduced (*P < 0.05). #indicates compared with sham group, P < 0.05. The values are presented as the mean ± SD. Confocal images showing the distribution of GFAP labeled astrocytes (D, G, J, red), VEGF (E, H, K, green), and GFAP labeling overlapping VEGF immunofluorescence can be seen in F, I and L. Note that VEGF expression in astrocytes (arrows) is markedly enhanced at 24 h following MCAO. However, after treatment with 10 % HS, it is noticeably reduced. Scale bars (D–L), 50 μm

Fig. 3
Fig. 3

VEGFR2 mRNA and protein expression in the peri-ischemic brain tissue in each group. A VEGFR2 (154 kDa) and GAPDH (37 kDa) immunoreactive bands, respectively. Bar graph B the optical density of VEGFR2 expression in 10 % HS group at 6, 12 and 24 h after MCAO is significantly decreased when compared with the corresponding ischemia group (*P < 0.05). Bar graph C the fold change in VEGFR2 mRNA expression. Significant differences in mRNA level in ischemia group at 6, 12 and 24 h after MCAO is evident when compared with the corresponding 10 % HS groups (*P < 0.05). #indicates compared with sham group, P < 0.05. The values are presented as the mean ± SD. Immunofluorescence images showing the distribution of GFAP (D, G, J, red) and VEGFR2 (E, H, K, green) in astrocytes (arrows) after MCAO for 12 h. Co-localization of GFAP and VEGFR2 can be seen in F, I and L. Note expression of VEGFR2 is down-regulated after treatment with 10 % HS. Scale bars (D–L), 20 μm

Fig. 4
Fig. 4

VEGF mRNA and protein expression in primary astrocytes. Bar graph A VEGF mRNA expression in HS group was significantly decreased in comparison to hypoxia group after hypoxia for 4 h. B VEGF (45 kDa) and GAPDH (37 kDa) immunoreactive bands, respectively. Bar graph C the optical density of VEGF expression in HS group was drastically attenuated when compared with the hypoxia group (*P < 0.05). #indicates compared with sham group, P < 0.05. The values are presented as the mean ± SD. Immunofluorescence images showing GFAP labelled astrocytes (D, G, J, red), double labelled with VEGF (E, H, K, green). Co-localized expression of GFAP and VEGF in astrocytes can be seen in F, I and L. Note that VEGF expression in astrocytes is markedly enhanced at 24 h following MCAO. However, after treatment with 10 % HS, it is noticeably reduced. Scale bars (D–L), 20 μm

Fig. 5
Fig. 5

VEGFR2 mRNA and protein expression in primary astrocytes. Bar graph A VEGFR2 mRNA expression in HS group was significantly decreased in comparison to hypoxia group after hypoxia for 4 h. B VEGFR2 (154 kDa) and GAPDH (37 kDa) immunoreactive bands, respectively. Bar graph C the optical density of VEGFR2 expression in HS group was drastically attenuated when compared with the hypoxia group (*P < 0.05). #indicates compared with sham group, P < 0.05. The values are presented as the mean ± SD. Double immunofluorescence images show that the expression of GFAP (D, G, J, red), VEGFR2 (E, H, K, green). The co-localized expression of GFAP and VEGFR2 can be seen in F, I and L. Note expression of VEGFR2 is markedly enhanced after hypoxia which is reduced by HS. Scale bars (D–L), 20 μm

Fig. 6
Fig. 6

Zo-1, claudin-5 mRNA and protein expression in the peri-ischemic brain tissue in each group. Bar graphs A, B the mRNA expressions of Zo-1, claudin-5 were significantly decreased at 6, 12 and 24 h after MCAO, respectively, as compared with the corresponding sham group (#indicates compared with the corresponding sham group, P < 0.05), but they were increased significantly at corresponding ischemia group after treatment with 10 % HS (*P < 0.05, **P < 0.01). C Zo-1 (225 kDa), claudin-5 (22 kDa) and GAPDH (37 kDa) immunoreactive bands, respectively. As seen in Bar graphs D, E the optical density of Zo-1, claudin-5 was significantly attenuated after MCAO at 6, 12 and 24 h (#indicates compared with sham group, P < 0.05). But 10 % HS could inhibit the down-regulation of Zo-1, claudin-5 protein expression effectively at 6, 12 h after MCAO (*P < 0.05, **P < 0.01, ns non-significant). The values are presented as the mean ± SD

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References

    1. Qureshi AI, Suarez JI. Use of hypertonic saline solutions in treatment of cerebral edema and intracranial hypertension. Crit Care Med. 2000;28(9):3301–3313. doi: 10.1097/00003246-200009000-00032. - DOI - PubMed
    1. Donkin JJ, Vink R. Mechanisms of cerebral edema in traumatic brain injury: therapeutic developments. Curr Opin Neurol. 2010;23(3):293–299. doi: 10.1097/WCO.0b013e328337f451. - DOI - PubMed
    1. Ziai WC, Toung TJ, Bhardwaj A. Hypertonic saline: first-line therapy for cerebral edema? J Neurol Sci. 2007;261(1–2):157–166. doi: 10.1016/j.jns.2007.04.048. - DOI - PubMed
    1. Zeng HK, Wang QS, Deng YY, Jiang WQ, Fang M, Chen CB, Jiang X. A comparative study on the efficacy of 10% hypertonic saline and equal volume of 20% mannitol in the treatment of experimentally induced cerebral edema in adult rats. BMC Neurosci. 2010;11:153. doi: 10.1186/1471-2202-11-153. - DOI - PMC - PubMed
    1. Schwarz S, Georgiadis D, Aschoff A, Schwab S. Effects of hypertonic (10%) saline in patients with raised intracranial pressure after stroke. Stroke. 2002;33(1):136–140. doi: 10.1161/hs0102.100877. - DOI - PubMed

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