BIO alleviates inflammation through inhibition of GSK-3β in a rat model of intracerebral hemorrhage

Sha Zhao BS, Zhen Liu MM, Zihan Yu BS, Xinran Wu BS, Rui Li BS and Xiaobo Tang MD, PhD
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  • Department of Biopharmaceutical Sciences (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China), College of Pharmacy, Harbin Medical University, Heilongjiang, China
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OBJECTIVE

Inflammation plays a key role in secondary brain damage following intracerebral hemorrhage (ICH). Glycogen synthase kinase–3β (GSK-3β) plays a strong proinflammatory role in many CNS diseases, including stroke. The present study was undertaken to examine the effects of 6-bromoindirubin-3ʹ-oxime (BIO), a specific inhibitor of GSK-3β, on inflammation in ICH rats.

METHODS

An ICH rat model was induced by autologous whole-blood injection into the striatum. First, 10, 20, 40, 60, 80, or 100 μg/kg BIO was applied to ICH animals to determine an optimal dosage for producing sufficient GSK-3β inhibition in rat ipsilateral hippocampus by Western blotting. Second, 40 μg/kg BIO was applied to ICH rats for 1, 3, 7, or 14 days, respectively, to determine a suitable intervention time course of BIO by Western blotting analysis on GSK-3β. Third, Western blotting and enzyme-linked immunosorbent assay were used for quantification of inflammation-related factors upstream or downstream of GSK-3β in rat ipsilateral hippocampus. Then, immunohistochemical staining was applied to detect activated microglia and apoptotic cells in rat ipsilateral hippocampus. Last, neurobehavioral tests were performed to assess the sensorimotor impairments in the ICH rats.

RESULTS

The results show that BIO 1) blocked GSK-3βTyr216 phosphorylation/activation, thus stabilizing β-catenin, increasing upstream brain-derived neurotrophic factor and downstream heat shock protein 70 levels, and decreasing the levels of nuclear factor–κB p65 and cyclooxygenase 2; 2) decreased the levels of the proinflammatory cytokines tumor necrosis factor–α and interleukin (IL)–1β and IL-6 and elevated the level of antiinflammatory cytokine IL-10; 3) inhibited microglia activation and cell apoptosis; and 4) improved the sensorimotor deficits of ICH rats.

CONCLUSIONS

BIO posttreatment inhibited microglia activation, prevented inflammation and hippocampal cell death, and ameliorated functional and morphological outcomes in a rat ICH model through inactivation of GSK-3β.

ABBREVIATIONS BDNF = brain-derived neurotrophic factor; BIO = 6-bromoindirubin-3ʹ-oxime; BSA = bovine serum albumin; CA1 = cornu ammonis 1; COX-2 = cyclooxygenase 2; DG = dentate gyrus; DMSO = dimethyl sulfoxide; ELISA = enzyme-linked immunosorbent assay; GSK-3β = glycogen synthase kinase–3β; HSP 70 = heat shock protein 70; ICH = intracerebral hemorrhage; IL = interleukin; NF-κB = nuclear factor–κB; NS = no statistical difference; p-GSK-3βTyr216 = phosphorylated GSK-3β; TNF-α = tumor necrosis factor–α.

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Contributor Notes

Correspondence Xiaobo Tang: College of Pharmacy, Harbin Medical University, Heilongjiang, China. ty6163@aliyun.com.

INCLUDE WHEN CITING Published online June 21, 2019; DOI: 10.3171/2019.4.JNS183501.

S.Z. and Z.L. contributed equally to this work.

Disclosures The authors report no conflict of interest concerning the materials or methods used in this study or the findings specified in this paper.

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